KR101896082B1 - Therapeutic target for cardiovascular disease and method for screening therapeutic agent for cardiovascular disease using the same - Google Patents

Abstract

A therapeutic agent for cardiovascular disease can be usefully screened using a novel therapeutic target for cardiovascular disease. An agent for inhibiting the expression or activity of a therapeutic target for cardiovascular disease can be usefully used as a pharmaceutical composition for treating cardiovascular disease. A method for screening the therapeutic agent includes a step of incubating cells with palmitic acid; a step of contacting the cells with a test substance; a step of comparing the expression level of the gene or the activity level of protein with the test substance; and a step of determining the test substance as a candidate agent for cardiovascular disease.

Description

TECHNICAL FIELD The present invention relates to a therapeutic target for a cardiovascular disease and a method for screening a therapeutic agent for cardiovascular disease using the same,

And a method for screening a therapeutic agent for cardiovascular diseases using the target.

Cardiovascular disease (CVD) is the world's highest death rate. According to the World Health Organization report, 23.6 million people are expected to die from cardiovascular disease by 2030. Myocardial infarction is a fatal disease that is the number one cause of sudden death in adults. The causes of cardiovascular disease are smoking, eating, overweight, obesity, and metabolic problems. Studies are continuing to prevent and treat cardiovascular diseases through blood pressure lowering, serum cholesterol lowering, and psychological treatment.

Conventional methods of diagnosing cardiovascular disease include invasive methods such as intravascular ultrasound (IVUS), and biomarkers for the diagnosis of cardiovascular disease include glutamic oxaloacetic transaminase (GOT) Lactate dehydrogenase (LDH), creatine kinase-MB (MB), troponin I, troponin T, and C-reactive protein (CRP) And B-type natriuretic peptide (BNP). However, the mechanisms and pathways that cause cardiovascular disease vary, and there is a continuing need to develop indicators that predict the risk of cardiovascular disease and reflect therapeutic effects. These studies can be useful in the process of discovering new therapies for cardiovascular disease.

One aspect includes contacting the cell with a test substance; Measuring the level of expression or protein activity of at least one gene selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3 in the cell to which the test substance is contacted; Comparing the expression level of the measured gene or the activity level of the protein with the level of the control group; And determining the test substance as a candidate for a therapeutic agent for a cardiovascular disease when the expression level of the gene or the activity level of the protein is lower than the level of the control group.

Another aspect is a method for detecting a test substance comprising contacting at least one protein selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3 with a test substance; Measuring the activity level of the protein contacted with the test substance; Comparing the activity level of the measured protein with the activity level of the control group; And determining the candidate substance as a candidate for a therapeutic agent for a cardiovascular disease when the activity level of the protein measured is lower than that of the control group.

Another aspect provides a pharmaceutical composition for treating cardiovascular diseases comprising, as an active ingredient, an expression of at least one gene selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3 or an inhibitor of protein activity.

One aspect includes contacting the cell with a test substance; Measuring the level of expression or protein activity of at least one gene selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3 in the cell to which the test substance is contacted; Comparing the expression level of the measured gene or the activity level of the protein with the level of the control group; And determining the test substance as a candidate for a therapeutic agent for a cardiovascular disease when the expression level of the gene or the activity level of the protein is lower than the level of the control group.

The Clec4d protein is a C-type lectin domain family member D, which is associated with cell-cell adhesion, immune response, and apoptosis, for example, NCBI Accession number NP_525126.2 The amino acid sequence of SEQ ID NO: 1, or the amino acid sequence of NCBI Accession number NP_034949.3, NP_001156633.1, SEQ ID NO: 3 or SEQ ID NO: 5. The Clec4d gene is a gene encoding Clec4d protein, which may be located in the telomeric region of the NK gene complex on the chromosome 12p13 region, for example, the nucleotide sequence of NCBI Accession number NM_080387.4, or the nucleotide sequence of SEQ ID NO: 2, or NCBI Accession numbers NM_010819.4, NM_001163161.1, SEQ ID NO: 4 or SEQ ID NO: 6.

The Clec4d protein has at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 92%, at least about 95% sequence identity to the amino acid sequence of SEQ ID NO: 1, SEQ ID NO: , At least about 97%, at least about 98%, or at least about 99% sequence homology. SEQ ID NO: 1 is a human Clec4d amino acid sequence, and SEQ ID NO: 3 or SEQ ID NO: 5 is a Clec4d amino acid sequence of a mouse.

The Clec4d gene has at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 92%, at least about 95% sequence identity to the nucleotide sequence of SEQ ID NO: 2, SEQ ID NO: , At least about 97%, at least about 98%, or at least about 99% sequence homology with the polynucleotide. SEQ ID NO: 2 is a human Clec4d nucleotide sequence, and SEQ ID NO: 4 or SEQ ID NO: 6 is a Clec4d nucleotide sequence of a mouse.

The Ctla2b protein is a cytotoxic T lymphocyte-associated protein 2 beta, which is associated with cysteine-type endopeptidase inhibitory activity and protein binding function, for example NCBI Accession number NP_031823.1, NP_001139273 . 1, SEQ ID NO: 7, or SEQ ID NO: 9. The Ctla2b gene may be a gene encoding a Ctla2b protein, for example, having a nucleotide sequence of NCBI Accession number NM_007797.1, NM_001145801.1, SEQ ID NO: 8 or SEQ ID NO: 10.

The Ctla2b protein is at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 92%, at least about 95% , 97% or more, about 98% or more, or about 99% or more. SEQ ID NO: 7 or 9 is the Ctla2b amino acid sequence of mouse.

The Ctla2b gene comprises at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 92%, at least about 95% , 97% or more, about 98% or more, or about 99% or more of the polynucleotide. SEQ ID NO: 8 or 10 is the Ctla2b nucleotide sequence of mouse.

The Ctla4 protein is a cytotoxic T lymphocyte-associated protein 4, which is associated with an immune checkpoint and a downregulation function of the immune response, for example NCBI Accession number NP_001032720.1, NP_005205. 2, SEQ ID NO: 11 or SEQ ID NO: 13. The Ctla4 gene may be a gene encoding a Ctla4 protein, for example, having a nucleotide sequence of NCBI Accession number NM_001037631.2, NM_005214.4, SEQ ID NO: 12 or SEQ ID NO: 14.

The Ctla4 protein comprises at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 92%, at least about 95% , 97% or more, about 98% or more, or about 99% or more. SEQ ID NO: 11 or SEQ ID NO: 13 is the human Ctla4 amino acid sequence.

The Ctla4 gene comprises a nucleotide sequence having at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 92%, at least about 95% , 97% or more, about 98% or more, or about 99% or more of the polynucleotide. SEQ ID NO: 12 or SEQ ID NO: 14 is the human Ctla4 nucleotide sequence.

Although the Ctla2b and Ctla4 genes are derived from different organisms, the protein encoded by Ctla4 may be a protein that exhibits the same or similar activity as the protein encoded by Ctla2b even though the protein name differs from that of the protein encoded by Ctla2b (for example, (isoenzyme or homolog), Ctla2b or Ctla4 can be applied depending on the origin of the cell in contact with the test substance.

The Csf3 protein is a colony-stimulating factor 3, which is a glycoprotein associated with the function of stimulating bone marrow to produce granulocytes and stem cells and release them into the bloodstream, for example NCBI Accession number NP_000750.1, NP_757373. 1, NP_757374.2, NP_001171618.1, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19 or SEQ ID NO: 21, or NCBI Accession number NP_034101.1 or SEQ ID NO: 23. The Csf3 gene is a gene encoding a Csf3 protein, for example, NCBI Accession number NM_000759.3, NM_172219.2, NM_172220.2, NM_001178147.1, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20 or SEQ ID NO: 22 Accession number NM_009971.1 or the nucleotide sequence of SEQ ID NO: 24.

The Csf3 protein is at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90% identical to the amino acid sequence of SEQ ID NO: 15, SEQ ID NO: , About 92% or more, about 95% or more, about 97% or more, about 98% or more, or about 99% or more of the sequence homology. SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 19 or SEQ ID NO: 21 is a human Csf3 amino acid sequence and SEQ ID NO: 23 is a Csf3 amino acid sequence of a mouse.

The Csf3 gene is at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90% identical to the nucleotide sequence of SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20, SEQ ID NO: , About 92% or more, about 95% or more, about 97% or more, about 98% or more, or about 99% or more of a polynucleotide. SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 20 or SEQ ID NO: 22 is a human Csf3 nucleotide sequence and SEQ ID NO: 24 is a Csf3 nucleotide sequence of a mouse.

As used herein, "homology" means the percentage of identity between two polynucleotides or polypeptide mimetics. The homology between sequences from one moiety to another can be determined by known techniques. For example, the algorithm BLAST (Karlin and Altschul, Pro. Natl. Acad. Sci. USA, 90, 5873 (1993)] or FASTA by Pearson (see Methods Enzymol., 183, 63 (1990)).

The method may comprise contacting the cell with a test substance. The test substance may be any substance expected to be effective for preventing or treating cardiovascular diseases. The contact can be appropriately selected depending on the material to be selected. The contacting may be, for example, administration of the test substance during culturing of the cells, or culturing of the cells in a medium containing the test substance.

The cell may comprise one or more genes or proteins selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3. The cell may be one that normally or naturally contains or produces one or more genes or proteins selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3 in the native cells. Or the cell may comprise an exogenous nucleic acid encoding at least one selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3. The term "exogenous nucleic acid" may refer to a nucleic acid that is normally or naturally not present in or produced by an original cell and may be introduced into a cell (e.g., by electroporation, transformation, transfection, And introducing other nucleic acids into the cells).

The Clec4d, Ctla2b, Ctla4, or Csf3 gene may be derived from different organisms, but may include a gene encoding a protein that exhibits the same or similar activity as the protein they encode. Likewise, the Clec4d, Ctla2b or Ctla4, or Csf3 protein may contain a protein having similar activity (for example, an isoenzyme or a homologue) even if the protein name is different.

The cells may be contained in a sample isolated from the subject. The cells may be contained in blood, serum, urine, feces, saliva, tears, cerebrospinal fluid, plasma or other tissues separated from the subject. The cells may be monocytes, vascular endothelial cells or macrophages. The cell may be an immunocyte derived from a subject suspected of having a cardiovascular disease, for example, a cardiovascular disease patient, such as a mononuclear cell, a vascular endothelial cell or a macrophage.

The method may include measuring the expression level of the at least one gene selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3 or the activity level of the protein in the cell to which the test substance is contacted.

The expression level of the gene may be determined by sequencing, RNase protection assay (RPA), nucleic acid microarray, Northern blot, polymerase chain reaction (PCR) Immunohistochemistry, Immunoblot and FACS (Immunohistochemistry), immunohistochemistry (RT), immunoprecipitation, radioimmunoassay (RIA), enzyme immunoassay (ELISA) ≪ RTI ID = 0.0 > and / or < / RTI > At this time, the expression level of the gene may be measured using a primer, a probe, or an antisense sequence thereof that specifically binds to the nucleotide sequence of Clec4d, Ctla2b, Ctla4 or Csf3 or its complementary sequence. The primer can form a base pair with a complementary template with a nucleic acid sequence having a free 3'hydroxyl group and can form a nucleotide sequence that acts as a polymerization initiator in the polymerization reaction for template strand replication it means. The primer may be one used in a nucleic acid amplification reaction. The amplification indicates an increase in the number of copies of the target sequence or its complementary sequence. The nucleic acid amplification reaction can be performed by a method known in the art. Wherein the primer is selected from the group consisting of 10 to 100 nt, 15 to 100 nt, 10 to 80 nt, 10 to 50 nt, 10 to 30 nt, 15 to 80 nt, 15 to 50 nt, 15 to 30 nt, 20 to 100 nt, 80 nt, 20 to 50 nt, or 20 to 30 nt. The probe means a nucleic acid fragment such as RNA or DNA capable of specifically binding to a nucleotide sequence of a target nucleic acid, for example, Clec4d, Ctla2b, Ctla4 or Csf3, and is labeled so that Clec4d, Ctla2b, Ctla4 or Csf3 Presence, amount and amount of expression can be confirmed. The probe may be prepared in the form of an oligonucleotide probe, a single strand DNA probe, a double strand DNA probe, or an RNA probe. For example, hybridization is carried out using a probe complementary to the DNA or RNA of Clec4d, Ctla2b, Ctla4 or Csf3, and the expression level of Clec4d, Ctla2b, Ctla4, or Csf3 gene can be confirmed through the degree of hybridization. Selection of suitable probes and hybridization conditions can be appropriately selected according to techniques known in the art. Wherein the probe is selected from the group consisting of 10 to 100 nt, 15 to 100 nt, 10 to 80 nt, 10 to 50 nt, 10 to 30 nt, 15 to 80 nt, 15 to 50 nt, 15 to 30 nt, 20 to 100 nt, 80 nt, or 30 to 50 nt.

The activity level of the protein may be one or more selected from the group consisting of SDS-polyacrylamide gel electrophoresis (SDS-PAGE), immunoblot, immunofluorescence, enzyme immunoassay, mass spectrometry, and protein chip . Determination of the activity level of the protein may be accomplished by methods known in the art.

The method comprises comparing the level of expression of the measured gene or the activity of the protein with that of the control group; And determining the test substance as a candidate for a therapeutic agent for a cardiovascular disease when the expression level of the measured gene or the activity level of the protein is lower than that of the control group. For example, the method includes the step of determining the test substance as a candidate for a therapeutic agent for cardiovascular disease when the measured expression level of the gene of Clec4d, Ctla2b or Ctla4, and Csf3 or the activity level of the protein is lower than that of the control group .

The control group may be a cell not in contact with the test substance, a cell contacted with a substance from which the active ingredient is excluded from the test substance, or a cell contacted with a solvent which dissolves the active ingredient in the test substance. The expression level of the gene or the activity level of the protein is lower than that of the control group in comparison with the expression level of the gene in the control group or that the activity level of the gene is lower than the activity level of the protein in the control group . This includes the absence of gene expression or protein activity, or the inability to function as a protein. The expression level or activity level may be at least about 5%, at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 55% , About 70% or more, about 75% or more, about 80% or more, about 85% or more, about 90% or more, about 95% or about 100% or less.

In order to prevent or treat cardiovascular diseases, there is a need for the development of drugs capable of modulating and inhibiting the activity of inflammatory signaling molecules, and / or the secretion of anti-inflammatory cytokines. When the expression of Clec4d, Ctla2b or Ctla4, or Csf3 gene or the activity of protein is inhibited in cells, the activity of inflammatory signal transduction molecules and / or the secretion of anti-inflammatory cytokines can be significantly reduced as compared with the control . The candidate substance for treating a cardiovascular disease may be one that inhibits the expression of at least one gene selected from the group consisting of Clec4d, Ctla2b, Ctla4, and Csf3, or the activity of the protein. Wherein the candidate substance for treating a cardiovascular disease inhibits the expression of at least one gene selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3, or the activity of a protein, thereby inhibiting the activity of the inflammatory signal transduction molecule and / It may be to inhibit secretion.

The test substance is selected from the group consisting of low molecular compounds, antibodies, antisense nucleotides, short interfering RNA (siRNA), short hairpin RNA (shRNA), nucleic acid, protein, , But is not limited thereto.

The method comprising: incubating the cell with palmitate before or during the contacting step and the measuring step; And incubating palmitic acid and incubated cells with minimally-oxidized low density lipoprotein (mMLDL). The method may also comprise incubating the palmitic acid and incubated cells with lipopolysaccharides (LPS) in the step of incubating with minimal oxidized low density lipoprotein.

In the step of incubating with the palmitic acid, the concentration of palmitic acid may be 10 to 1000 μM, 20 to 500 μM, 30 to 300 μM, 40 to 250 μM, 50 to 200 μM, 75 to 150 μM, or 90 to 110 μM. The palmitic acid may be incubated with the cells for 2 to 150 hours, 4 to 70 hours, 10 to 30 hours, 12 to 20 hours, or 14 to 18 hours.

In the step of incubating with the mmLDL, the concentration of mmLDL is in the range of 5 to 500 μg / ml, 10 to 250 μg / ml, 15 to 150 μg / ml, 20 to 120 μg / ml, 25 to 100 μg / 45 to 55 μg / ml. The mmLDL may be incubated with the cells for 5 minutes to 10 hours, 10 minutes to 8 hours, 20 minutes to 7 hours, 20 minutes to 40 minutes, or 3 hours to 7 hours.

In the step of incubating with the LPS, the concentration of LPS is in the range of 1 to 100 ng / ml, 2 to 50 ng / ml, 4 to 25 ng / ml, 5 to 20 ng / ml, 7.5 to 12.5 ng / Lt; / RTI > The LPS may be incubated with the cells for 5 minutes to 10 hours, 10 minutes to 8 hours, 20 minutes to 7 hours, 20 minutes to 40 minutes, or 3 hours to 7 hours.

In order to prevent or treat cardiovascular diseases, the cholesterol environment, the form of low density lipoprotein (LDL) and the form of saturated fatty acid (SFA) are important, and the inflammation-induced There is a need for the development of drugs capable of modulating and inhibiting the activity of signaling molecules and / or the secretion of anti-inflammatory cytokines. Palmitate and mmLDL can promote the activation of inflammatory signaling molecules and / or the secretion of anti-inflammatory cytokines synergistically in immune cells. Palmitate and mmLDL can synergistically increase the expression of Clec4d, Ctla2b or Ctla4, or Csf3 gene in immune cells. LPS can further stimulate the activity of inflammatory signaling molecules and / or the secretion of anti-inflammatory cytokines in immune cells. Therefore, the candidate substance for treating a cardiovascular disease may be one that inhibits the expression of Clec4d, Ctla2b, Ctla4, or Csf3 gene or protein. The candidate agent for treating a cardiovascular disease may be one which inhibits the expression of Clec4d, Ctla2b or Ctla4, or the Csf3 gene, or the activity of a protein, thereby inhibiting the activity of an inflammatory signal transduction molecule and / or the secretion of an anti-inflammatory cytokine.

The method comprises measuring the expression level or protein activity level of at least one gene selected from the group consisting of CXCL2, TNF-α, pERK, pp38, Ccr5, IL-6, IL-1β and Nlrp3 ; Comparing the expression level of the measured gene or the activity level of the protein with the level of the control group; And determining the test substance as a candidate for a therapeutic agent for a cardiovascular disease when the expression level of the measured gene or the activity level of the protein is lower than that of the control group. For example, the method may include measuring the level of expression or protein activity of CXCL2, TNF-a, pERK, pp38, Ccr5, IL-6, IL-l [beta] and Nlrp3 genes in cells to which the test substance is contacted; Comparing the level of expression or protein expression of the measured CXCL2, TNF-alpha, pERK, pp38, Ccr5, IL-6, IL-l [beta] and Nlrp3 genes with the level of the control; When the level of expression of CXCL2, TNF-α, pERK, pp38, Ccr5, IL-6, IL-1β and Nlrp3 gene or the protein is low compared to the level of the control group, the test substance is determined as a candidate for a therapeutic agent for cardiovascular disease And the like.

The CXCL2 protein is a chemokine (CXC motif) ligand 2, a chemokine secreted from monocytes and macrophages, for example, NCBI Accession number NP_002080.1 or SEQ ID NO: 25, or NCBI Accession number NP_033166.1 or SEQ ID NO: 26. The CXCL2 gene may be a gene encoding a CXCL2 protein, for example, having a nucleotide sequence of NCBI Accession number NM_002089.3 or NM_009140.2.

The TNF-alpha protein is a tumor necrosis factor alpha, a cell signal transduction cytokine involved in systemic inflammation, for example, NCBI Accession number NP_000585.2 or SEQ ID NO: 27, or NCBI Accession number NP_038721. 1, NP_001265530.1, SEQ ID NO: 28 or SEQ ID NO: 29. The TNF-a gene may be a gene encoding the TNF-a protein, having the nucleotide sequence of NCBI Accession number NM_000594.3, NM_013693.3 or NM_001278601.1.

The pERK protein is a phosphorylated extracellular signal-regulated kinase (ERK), which phosphorylates transcription factors that regulate the expression of genes involved in apoptosis and apoptosis. The ERK protein is, for example, NCBI Accession SEQ ID NO: 31, SEQ ID NO: 32 or SEQ ID NO: 33, or NCBI Accession number NP_001305031.1, NP_056621.4, NP_081694.1, NP_038899, NP_00127896.1, NP_001278968.1, NP_002739.1, SEQ ID NO: 36, SEQ ID NO: 38, SEQ ID NO: 38 or SEQ ID NO: 39. The ERK gene is a gene coding for the ERK protein, which is a gene coding for the ERK protein. NCKI Accession number NM_002747.3, NM_001292040.1, NM_001292039.1, NM_002748.3, NM_001318102.1, NM_027418.2, NM_015806.5, NM_011950.2, NM_013871.3, Or a nucleotide sequence of NM_011161.5.

The pp38 protein belongs to the mitogen-activated protein kinase (MAPK), which is phosphorylated by the inflammatory cytokine p38, which regulates the expression of genes related to cell stress response, apoptosis and apoptosis. p38 The protein may be, for example, an amino acid of NCBI Accession number NP_620583.1, NP_620582.1, NP_620581.1, NP_001306.1 or SEQ ID NO: 40 or NP_036081.1, NP_001161980.1, NP_001161985.1, NP_001161986.1 or SEQ ID NO: 41 Sequence. ≪ / RTI > The p38 gene is a gene coding for the p38 protein and has a nucleotide sequence of NC_I Accession number NM_001315.2, NM_139014.2, NM_139013.2, NM_139012.2, NM_011951.3, NM_001168508.1, NM_001168513.1, or NM_001168514.1 .

The Ccr5 protein is a CC chemokine receptor type 5, a protein located on the surface of a leukocyte, for example, NCBI Accession NP_000570.1, NP_001093638.1 or SEQ ID NO: 42, or NP_034047.2 or SEQ ID NO: 43 < / RTI > The Ccr5 gene may be a gene encoding a Ccr5 protein, which has a nucleotide sequence of NCBI Accession number NM_001100168.1, NM_000579.3, or NM_009917.5.

The IL-6 protein is Interleukin-6, a proinflammatory cytokine and an anti-inflammatory myokine, for example, NCBI Accession NP_001305024.1, NP_000591.1 or SEQ ID NO: 44, or NP_112445. 1, NP_001300983.1 or SEQ ID NO: 45. The IL-6 gene may be a gene encoding the IL-6 protein, having the nucleotide sequence of NCBI Accession Nos. NM_001318095.1, NM_000600.4, NM_031168.2, or NM_001314054.1.

IL-1 [beta] protein is an interleukin-1 beta, a lymphocyte activating factor, produced from activated macrophages and an important mediator in the immune response. The IL-1? Protein may be, for example, having the amino acid sequence of NCBI Accession NP_000567.1 or SEQ ID NO: 46, or NP_032387.1 or SEQ ID NO: 47. The IL-1? Gene may be a gene encoding the IL-1? Protein, having the nucleotide sequence of NCBI Accession number NM_000576.2 or NM_008361.4.

Nlrp3 is a protein 3 containing NACHT, LRR and PYD domains (NACHT, LRR and PYD domains-containing protein 3: NALP3), which is mainly expressed in macrophages and activated Nlrp3 induces an immune response. The Nlrp3 protein is, for example, one having the amino acid sequence of NCBI Accession NP_001230062.1, NP_001120933.1, NP_001120934.1, NP_001073289.1, NP_004886.3, NP_899632.1 or SEQ ID NO: 48, NP_665826.1 or SEQ ID NO: 49 . The Nlrp3 gene may be a gene encoding Nlrp3 protein having a nucleotide sequence of NCBI Accession Nos. NM_001243133.1, NM_001127461.2, NM_001127462.2, NM_001079821.2, NM_004895.4, NM_183395.2, or NM_145827.3 .

When the expression of CXCL2, TNF-α, pERK, pp38, Ccr5, IL-6, IL-1β or Nlrp3 gene or the activity of a protein is inhibited in a cell or the like, activation of an inflammatory signal transduction molecule and / May be significantly reduced compared to the control group. The agent for treating a cardiovascular disease may be one that inhibits the expression of at least one gene selected from the group consisting of CXCL2, TNF-α, pERK, pp38, Ccr5, IL-6, IL-1β and Nlrp3 or protein. Wherein the final test substance of the cardiovascular disease inhibits the expression of at least one gene selected from the group consisting of CXCL2, TNF-alpha, pERK, pp38, Ccr5, IL-6, IL- Lt; RTI ID = 0.0 > and / or < / RTI > secretion of anti-inflammatory cytokines.

The cardiovascular disease may be a disease in which blood vessels are narrowed due to abnormal cholesterol regulation or proliferation of vascular endothelial cells. The cardiovascular disease may be a disease caused by an activation of an inflammatory reaction with immune cells due to abnormal cholesterol control in hypertrophy, obesity, and accompanying oxidative stress. The cardiovascular disease may be one or more selected from the group consisting of atherosclerosis, hypertension, myocardial infarction and angina.

Another aspect is a method for detecting a test substance comprising contacting at least one protein selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3 with a test substance; Measuring the activity level of the protein contacted with the test substance; Comparing the activity level of the measured protein with the activity level of the control group; And determining the candidate substance as a candidate for a therapeutic agent for a cardiovascular disease when the activity level of the protein measured is lower than that of the control group.

The method comprising incubating the protein with palmitic acid before or during the contacting step and the measuring step; And incubating the palmitic acid and the incubated protein with minimally-oxidized low density lipoprotein (mMLDL). The method may also include incubating the palmitic acid and the incubated protein with lipopolysaccharides (LPS) in the step of incubating with minimal oxidized low density lipoprotein.

The at least one protein selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3 is selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3, or a protein expressed normally or naturally Type protein. Or a protein expressed from a cell in vitro using an expression vector comprising a nucleic acid molecule encoding at least one protein selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3. The expression vector may be a baculovirus expression vector, a mammalian expression vector, or a bacterial expression vector. The cells may be insect cells, mammalian cells, or bacterial cells, but are not limited thereto. The expression vector refers to a gene construct comprising an essential regulatory element operably linked to the expression of the gene insert capable of expressing the desired protein in a suitable host cell. The expression vector can be used for the purpose of delivering Clec4d, Ctla2b or Ctla4, or Csf3 protein to a cell. The expression vector may be prepared to include expression regulatory elements such as promoter, operator, initiation codon, termination codon, polyadenylation signal, enhancer. Expression vectors can be self-replicating or integrated into host DNA. The vector may include, but is not limited to, a plasmid vector, a cosmid vector, an episome vector, a viral vector, and the like.

Said method comprising the steps of: contacting at least one protein selected from the group consisting of CXCL2, TNF-alpha, pERK, pp38, Ccr5, IL-6, IL-1 beta and Nlrp3 with a test substance; Measuring the activity level of the protein contacted with the test substance; Comparing the activity level of the measured protein with the activity level of the control group; And determining the test substance as a candidate for a therapeutic agent for cardiovascular disease when the measured activity level of the protein is lower than that of the control group.

The control group may be a protein which is in contact with a protein not in contact with a test substance, a protein in contact with a substance in which the active ingredient is excluded from the test substance, or a solvent which dissolves the active ingredient in the test substance. The activity level of the protein is lower than that of the control group, which may be lower than the activity of the protein in the control group. This includes the absence of protein activity. The activity level may be at least about 5%, at least about 10%, at least about 20%, at least about 30%, at least about 40%, at least about 50%, at least about 55%, at least about 60% About 70% or more, about 75% or more, about 80% or more, about 85% or more, about 90% or more, about 95% or more or about 100% or less.

Incubating the protein with palmitic acid, minimal oxidized low density lipoprotein and / or lipopolysaccharide may be performed by incubating the cells with palmitic acid, minimal oxidized low density lipoprotein and / or lipopolysaccharide, Same as. As described above for Clec4d, Ctla2b, Ctla4, Csf3, CXCL2, TNF- ?, pERK, pp38, Ccr5, IL-6, IL-1 ?, Nlrp3 and cardiovascular diseases.

One aspect provides a pharmaceutical composition for the treatment of cardiovascular diseases comprising, as an active ingredient, an expression of at least one gene selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3 or an inhibitor of protein activity.

The expression inhibitor of one or more genes selected from the group consisting of Clec4d, Ctla2b or Ctla4 and Csf3 may be an antisense nucleotide complementary to Clec4d, Ctla2b or Csf3 mRNA, a small hairpin RNA (shRNA) A small interfering RNA (siRNA), and a ribozyme. The activity inhibitor of at least one protein selected from the group consisting of Clec4d, Ctla2b, Ctla2b, Ctla4, and Csf3 may be a compound that specifically binds to Clec4d, Ctla2b or Ctla4 or Csf3 protein, a peptide, a peptide mimetic, , An aptamer, and an antibody. Suitable inhibitors of gene expression inhibitors and / or proteins may be appropriately selected according to techniques known in the art.

The siRNA is composed of a sense sequence of 15 to 30 mers selected in the nucleotide sequence of Clec4d, Ctla2b or Ctla4, or a gene encoding the Csf3 protein, and an antisense sequence complementarily binding to the sense sequence Lt; / RTI > The antisense sequence may be one that binds (hybridizes) to a complementary base sequence of DNA, immature-mRNA or mature mRNA to inhibit the flow of genetic information as a protein in DNA, as defined in the Watson-click base pair. Because antisense sequences are the long chains of monomeric units they can be synthesized against the target RNA sequence.

The peptide mimetics inhibit the binding domain of Clec4d, Ctla2b or Ctla4, or Csf3 protein, or inhibit the activity of Clec4d, Ctla2b or Ctla4, or Csf3 protein. Peptide mimetics may be peptides or non-peptides and may be derived from amino acids linked by non-peptide bonds, such as psi bonds (Benkirane, N., et al. J. Biol. Chem., 271: 33218-33224, Lt; / RTI > Also included are cyclic mimetics comprising a conformationally constrained peptide, cyclic mimetics, at least one exocyclic domain, a binding moiety (binding amino acid) It may be. The peptide mimetics are structurally similar to the secondary structural features of the Clec4d, Ctla2b or Ctla4, or Csf3 proteins and are characterized by antibodies (Park, BW et al. Nat Biotechnol 18, 194-198, 2000) or water soluble receptors (Takasaki, al. Nat Biotechnol 15, 1266-1270, 1997), and may be a novel small molecule capable of acting as an effect equivalent to a natural antagonist.

The aptamer is a single-chain DNA or RNA molecule. It can be produced by an evolutionary method using an oligonucleotide library called a systematic evolution of ligands by exponential enrichment (SELEX). The aptamer has high affinity and selectivity to specific chemical molecules or biological molecules (C. Tuerand L. Gold, Science 249, 505-510, 2005; AD Ellington and JW Szostak, Nature 346, 818-822, 1990; M. Famulok, et. DS Wilson and Szostak, Annu Rev. Biochem. 68, 611-647, 1999). Aptamers can specifically bind to a target and modulate the activity of the target, for example, by blocking the ability of the target to function through binding.

The antibody may be one that specifically and directly binds to Clec4d, Ctla2b or Ctla4, or Csf3 protein to effectively inhibit the activity of Clec4d, Ctla2b or Csf3. The antibody specifically binding to Clec4d, Ctla2b or Csf3 may be a polyclonal antibody or a monoclonal antibody. The antibody specifically binding to Clec4d, Ctla2b or Csf3 may be prepared by a known method known to those skilled in the art, and commercially available Clec4d, Ctla2b or Ctla4 or Csf3 antibody can be purchased and used. The antibody can be prepared by injecting the immunocompetent Clec4d, Ctla2b or Ctla4, or Csf3 protein into an external host according to conventional methods known to those skilled in the art. External hosts include mammals such as mice, rats, sheep, and rabbits. The immunogen may be injected intraperitoneally, intraperitoneally or subcutaneously, and blood collected from an external host may be collected periodically to collect sera showing specificity to the formed titer and the antigen to isolate the antibody.

The pharmaceutical composition may contain the active ingredient in an amount of 0.0001 to 50% by weight based on the total weight of the composition. The pharmaceutical composition may be one comprising at least one pharmaceutically acceptable carrier in addition to the above-described effective ingredient for administration. The pharmaceutically acceptable carrier may be a mixture of one or more of saline, sterilized water, Ringer's solution, buffered saline, dextrose solution, maltodextrin solution, glycerol, ethanol, liposome, Other conventional additives such as buffers, bacteriostats and the like may be added. In addition, it can be formulated into injection formulations, pills, capsules, granules or tablets such as aqueous solutions, suspensions, emulsions and the like by additionally adding diluents, dispersants, surfactants, binders and lubricants, Specific antibody or other ligand can be used in combination with the carrier. Can be formulated according to each disease or component, using methods well known in the art or as disclosed in Remington ' s Pharmaceutical Science, Mack Publishing Company, Easton PA.

The pharmaceutical compositions may be formulated for oral, topical, parenteral, intranasal, intravenous, intramuscular, subcutaneous, intravaginal, transdermal, and the like administration. The pharmaceutical composition may be administered orally or parenterally at the time of clinical administration and may be administered orally or parenterally in the form of intraperitoneal injection, rectal injection, subcutaneous injection, intravenous injection, intramuscular injection, intrauterine injection, May be administered by intrathecal injection. The administration subject may be a mammal such as a human, a pig, a cow, a horse, a dog, a cat, or an experimental animal such as a mouse, a rat, a rabbit, a guinea pig, or a hamster. The dosage of the composition used can be controlled by various parameters, in particular by gene, vector, mode of administration, type of cardiovascular disease or duration of treatment. In addition, the range can be adjusted depending on the body weight, age, sex, health condition, diet, excretion rate, and severity of disease. The daily dose may be administered in a dose of about 0.0001 to 100 mg / kg, or 0.001 to 10 mg / kg, once to several times a day.

As described above for Clec4d, Ctla2b, Ctla4, Csf3, CXCL2, TNF- ?, pERK, pp38, Ccr5, IL-6, IL-1 ?, Nlrp3 and cardiovascular diseases.

Clec4d, Ctla2b or Ctla4, and Csf3 can be usefully screened for a therapeutic agent for cardiovascular disease by measuring the expression of the gene or the activity of the protein using the target as a therapeutic target for cardiovascular disease. In addition, expression of at least one gene selected from the group consisting of Clec4d, Ctla2b or Ctla4, and Csf3 or an inhibitor of protein activity may be useful as a pharmaceutical composition for treating cardiovascular diseases.

FIG. 1A shows the results of measuring the secretion amount of CXCL2 and TNF-a in a control group and a test group to which palmitic acid, mmLDL, LPS, or a combination thereof was administered. FIG. 1B shows the results of measurement of the expression levels of pERK, ERK, pp38 and p38 in a control group and a test group to which a combination of palmitic acid, LPS, and mmLDL was administered.
Figure 2 shows the number of differentially expressed genes in RNA sequence analysis as a van diagram.
Figure 3a is a heat map result showing the change in gene in the TLR-4 dependent pathway when palmitic acid, mmLDL, or a combination thereof is administered. Figure 3b shows the results of measuring RNA sequencing and expression levels of genes in the TLR-4 dependent pathway when palmitic acid, mmLDL, or a combination thereof is administered.
Figure 4a is a heat map showing the change in gene in the TLR-4 independent pathway when palmitic acid, mmLDL, or a combination thereof is administered. Figure 4b shows the results of measuring RNA sequencing and expression levels of genes in the TLR-4 independent pathway when palmitic acid, mmLDL, or a combination thereof is administered.
Figure 5A shows the results of RNA sequencing and expression levels of Nlrp3 when palmitic acid, mmLDL, or a combination thereof is administered. FIG. 5B shows the results of measurement of the expression level of Il-1 beta in a control group and a test group to which TLR4 inhibitor was administered and a combination of palmitic acid, LPS and mmLDL was administered.

Hereinafter, the present invention will be described in more detail with reference to examples. However, these examples are for illustrative purposes only, and the scope of the present invention is not limited to these examples.

Example  One: Atherosclerosis  Analysis of gene expression changes in the model

1. Palmitic acid and minimal oxidized low density Lipoprotein  (minimally-oxidized low density lipoprotein: mmLDL ) And J774 cell culture

The macrophage-like cell line J774A.1 (hereinafter referred to as J774 cell), derived from murine, was obtained from Yury I Miller (Department of Medicine, University of California San Diego, La Jolla, CA, USA). J774 cells were cultured in Dulbecco's modified Eagle's medium (DMEM) (Grand Island, NY, USA) containing 1% penicillin-streptomycin and 10% fetal bovine serum (FBS) ≪ / RTI >

Low density lipoproteins (density, 1.019 to 1.063 g / ml) were obtained by sequential centrifugation of plasma from healthy donors. 50 [mu] g / ml of LDL was incubated in murine and fibroblasts expressing human 15-lipoxygenase and serum-free DMEM for about 18 hours to obtain mmLDL. Lipopolysaccharides (LPS) and palmitate were used commercially (Sigma-Aldrich, St. Louis, Mo., USA). The palmitic acid was dissolved in 70% ethanol containing 0.1 M NaOH at a concentration of 30 mM to prepare a stock solution.

2. Palmitic acid mmLDL  Secretion of anti-inflammatory cytokines and MAPK  Analysis of effects on expression of pathway factor

(1) Palmitic acid and to mmLDL  Of the secretion level of anti-inflammatory cytokines

The effect of palmitic acid and mmLDL on cytokine secretion was measured by Enzyme-Linked Immunosorbent Assay (ELISA). J774 cells were seeded in a 60 mm culture dish at a concentration of 10 ⁶ cells / well and cultured at 37 ° C overnight. J774 cells were treated with 100 μM palmitic acid and cultured for 16 hours, followed by 10 ng / ml of LPS and / or 50 μg / ml of mMLDL and incubated for 5 hours. Specifically, in the first test group, palmitic acid was administered to J774 cells. In the third test group, palmitoic acid and mmLDL were administered to J774 cells. In the second test group, mmLDL was administered to J774 cells. In the fifth test group, palmitic acid and LPS were administered to J774 cells. In the fourth test group, LPS was administered to J774 cells. In the sixth test group, LPS and mmLDL were administered. In the seventh test group, palmitoic acid, LPS and mmLDL were administered to J774 cells. For the control group, J774 cells were administered with ethanol and / or dimethyl sulfoxide (DMSO). Subsequently, a supernatant of the medium was obtained in each test group. The amount of CXCL2 and TNF-? Secreted in the supernatant was measured using an ELISA kit (R & D Systems, Minneapolis, MN, USA). The above procedure was performed according to the manufacturer's instructions. 96 well plates were coded with capture antibody at a concentration of 1 mg / well. The antibody-coated plate was washed twice with phosphate-buffered saline (PBS) containing 0.05% Tween-20. Next, biotin-conjugated secondary antibody was added to each well. The plate was read at an absorbance of 450 nm according to the manufacturer's instructions.

FIG. 1A shows the results of measuring the secretion amount of CXCL2 and TNF-a in a control group and a test group to which palmitic acid, mmLDL, LPS, or a combination thereof was administered. In the third test group treated with palmitic acid and mmLDL, the secretion of each of CXCL2 and TNF-α was significantly increased compared with the control group. Similarly, the seventh test group treated with palmitic acid and mmLDL significantly increased the secretion of CXCL2 and TNF-α compared to the fourth test group.

(2) to mmLDL  by MAPK  Measurement of expression level of pathway factor

The effect of palmitic acid and mmLDL on the phosphorylation of ERK and p38, inflammatory signaling molecules, was measured by immunoblot. J774 cells were seeded in a 60 mm culture dish at a concentration of 10 ⁶ cells / well and cultured at 37 ° C overnight. J774 cells were treated with 100 μM palmitic acid and cultured for 16 hours. LPS and / or 50 μg / ml of 10 ng / ml of mMLDL were added and cultured for 30 minutes. Except when administered, the test group and the control group received the same substances as (2). Cells were then harvested and centrifuged at 14,000 rpm for 45 minutes at 4 < 0 > C to obtain cell pellets. The obtained cells were dissolved in a cell lysis buffer. The cell lysis buffer contained 1 M HEPES (pH 7.5), 5 M NaCl, 0.5 M ethylenediaminetetraacetic acid (EDTA), 1% Triton X-100, and a protease inhibitor cocktail (Roche, Indianapolis, IN, USA). Proteins contained in the cell lysate were quantified, and 20 μg of the protein was separated by 12.5% polyacrylamide gel electrophoresis (SDS-PAGE) for each test group and the control group. The separated proteins were transferred to a polyvinylidene fluoride (PVDF) membrane (EMD Millipore, Billerica, MA, USA). The PVDF membrane was reacted with the primary antibody overnight at 4 ° C. On the next day, the PVDF membrane was washed with TBST solution containing Tween, Tris, and NaCl, and reacted with HRP-conjugated secondary antibody at room temperature. Protein bands were visualized using an enhanced chemiluminescence (ECL) kit system (EMD Millipore).

FIG. 1B shows the results of measurement of expression levels of pERK, ERK, pp38 and p38 in the control group and the test group to which a combination of palmitic acid, LPS, and mmLDL was administered. In the third test group treated with palmitic acid and mMLDL, the expression of pERK and pp38 was increased in a concentration dependent manner compared to the control group. Similarly, in the seventh test group treated with palmitic acid and mMLDL, the expression of pERK and pp38 was increased in a concentration-dependent manner compared to the fourth test group.

Thus, it was confirmed that the activity and expression of inflammatory signal transduction molecules and secretory cytokines in macrophages were increased.

3. Palmitic acid to mmLDL  Search for genes and pathways controlled by

Differentially Expressed Genes Using RNA Sequencing and Using Kyoto Encyclopedia of Genes and Genomes (KEGG) Software to Detect Genes and Pathways Regulated by Palmitate and mmLDL Differentially Expressed Genes: DEG) were analyzed. J774 cells were treated with 100 μM palmitic acid and cultured for 16 hours. LPS and / or 50 μg / ml of 10 ng / ml of mMLDL were added and cultured for 4 hours. An RNA sequencing library was constructed using the Illumina Truseq Stranded Total RNA sample preparation kit (Illumina, San Diego, Calif., USA) according to the manufacturer's instructions. The rRNA was depleted in 1 μg of RNA using a Ribo-zero rRNA removal kit (human / mouse / rat) (Illumina). RNA with rRNA removed was fragmented using divalent cations. First strand cDNA was synthesized from RNA fragments using reverse transcriptase and random primers. DNA polymerase I and RNase H were used to synthesize cDNA second strand from cDNA first strand. Thereafter, adenine (A) was bound to the end of the cDNA strand, the adapter was ligation, and amplified by PCR to prepare a cDNA library. The amplified library quality was measured by capillary electrophoresis (Agilent, Santa Clara, CA, USA). Q-PCR was performed using SYBR Green Master Mix (Applied Biosystems, Waltham, Mass., USA) with the library as a template. Then, an equal amount of label was bound to the library. The labeled binding library was loaded into the Illumina NextSeq 500 system and the sequence of the RNA from the library was analyzed according to the manufacturer's instructions for 2x75 sequencing. DEG pathway analysis was performed using KEGG software to identify signal transduction pathways and related genes affected by palmitic acid and mmLDL.

Figure 2 shows the number of differentially expressed genes in RNA sequence analysis as a van diagram. Referring to FIG. 2, in the case of administration of palmitic acid, expression of 8 genes, in the case of administration of mmLDL, of 314 genes, and in the case of administration of palmitic acid and mmLDL, expression of 254 genes were respectively increased.

Table 1 shows the genes whose expression was significantly changed when palmitic acid and mmLDL were administered and the pathway to which these genes belong. When palmitic acid and mmLDL were administered, the genes associated with the terpenoid skeletal synthesis pathway were decreased. In Table 1, genes in pathways with high significance were further selected.

Route gene P Cytokine-cytokine receptor interaction. CSF3 , CSF2 , IL6, CCL2 , PDGFB , CCR1 , CXCL2 , IL4RA , CCL7 , IL17RA , CXCL10 , OSM, INHBA , TNFRSF9 , TNFRSF1B , IL23A , CCR5 , IL1B , IL1A 4.3E-07 Hematopoietic cell line
(Hematopoietic cell lineage) CSF3 , CSF2 , IL6, H2-EB1, IL4RA , H2-AA, IL1B , ITGA4 , IL1A , ITGAM 2.2E-05 Terpenoid skeletal synthesis
(Terpenoid backbone biosynthesis) HMGCR , FDPS , HMGCS1 , ACAT2 , IDI1 9.9E-05 Graft-versus-host disease
(Graft-versus-host disease) IL6, CD80, H2-EB1, H2-AA, IL1B , IL1A 4.3E-03 NOD-like receptor signal pathway
(NOD-like receptor signaling pathway) IL6, CCL2 , CXCL2 , IL1B , NLRP3 , CCL7 5.7E-03 Cell adhesion molecule
(CAMs) ICAM1 , ITGAL , SIGLEC1 , CD80, ITGAV , H2-EB1, H2-AA, ITGA4 , ITGAM 7.7E-03 Axon guidance SEMA5A , EPHA4 , PLXNC1 , RND1 , KRAS , SEMA6D , SEMA4C , EPHA2 1.1E-02 Immune network for IgA production (Intestinal immune network for IgA production) IL6, CD80, H2-EB1, H2-AA, ITGA4 1.8E-02 Non-large cardiomyopathy
(Hypertrophic cardiomyopathy: HCM) ACTG1 , TNNT2 , IL6, ITGAV , LMNA , ITGA4 2.0E-02 Chemokine signal pathway
(Chemokine signaling pathway) KRAS , CCL2 , CCR5 , FGR , CCR1 , CXCL2 , PXN , CCL7 , CXCL10 2.0E-02 Type I diabetes (Type I diabetes mellitus) CD80, H2-EB1, H2-AA, IL1B , IL1A 2.9E-02 Viral myocarditis ACTG1 , ICAM1 , ITGAL , CD80, H2-EB1, H2-AA 3.0E-02 Toll-like receptor signaling pathway
(Toll-like receptor signaling pathway) FOS , IL6, CD80, IL1B , TLR8 , CXCL10 3.7E-02 Arrhythmogenic right ventricular cardiomyopathy (ARVC) ACTG1 , ITGAV , LMNA , GJA1 , ITGA4 5.1E-02 MAPK signal path DUSP5 , FOS , KRAS , PDGFB , RASGRP3 , NR4A1 , IL1B , GADD45A , IL1A , DUSP6 5.8E-02

4. Palmitic acid to mmLDL  Controlled by TLR4  Gene analysis of dependent pathways

(1) Palmitic acid and to mmLDL  by TLR4  Expression Analysis of Gene in Dependent Pathway 1

The effect of palmitate and mmLDL on the genes in the TLR4 pathway is shown in the heat map. FIG. 3A is a heat map result (≥ Log 2 fold change) showing the change in gene in the TLR-4 dependent pathway when palmitic acid, mmLDL or a combination thereof is administered. Figure 3b shows the results of measuring RNA sequencing and expression levels of genes in the TLR-4 dependent pathway when palmitic acid, mmLDL, or a combination thereof is administered. 3A and 3B, the expression of Ccr5, Il-6, Csf3 and Il-1β was significantly increased when palmitic acid and mmLDL were administered.

(2) to mmLDL  by TLR4  Expression Analysis of Gene in Dependent Pathway 2

The effect of palmitic acid and mmLDL on the expression of Ccr5, Il-6, Csf3 and Il-1β was measured by real-time polymerase chain reaction (RT-PCR). J774 cells were seeded in a 60 mm culture dish at a concentration of 10 ⁶ cells / well and cultured at 37 ° C overnight. J774 cells were treated with 100 μM palmitic acid and cultured for 16 hours. LPS and / or 50 μg / ml of 10 ng / ml of mMLDL were added and cultured for 4 hours. Specifically, in the first test group, LPS was administered to J774 cells. In the second test group, palmitoic acid and LPS were administered to J774 cells. In the third test group, mmLDL and LPS were administered to J774 cells. In the fourth test group, palmitoic acid, mmLDL and LPS were administered to J774 cells. For the control group, J774 cells were administered with ethanol and / or DMSO. Cells were harvested and RNA was extracted according to the manufacturer's instructions using Ribospin RNA Extraction Kit (GeneAll, Seoul, Korea). CDNA was synthesized from 1 μg of RNA using the QuantiTect Reverse Kit (QIAGEN, Venlo, Netherlands). Using the synthesized cDNA as a template, primers specific for each gene (Table 2 below) and a SYBR-Green dye system and a LightCycler 480 real-time PCR instrument (Roche Applied Science, Benzberg, Germany) -PCR was performed. The analyzed gene expression levels were normalized to beta-actin.

gene Forward primer (5 '-> 3') The reverse primer (5 '-> 3') Ccr5 GTTTGCCTCTCTCCCAGAAATA
(SEQ ID NO: 50) CTGAGTAGCAGATGACCATGAC
(SEQ ID NO: 51) Il-6 TGGATGCTACCAAACTGGATATAA
(SEQ ID NO: 52) AGGACTCTGGCTTTGTCTTTC
(SEQ ID NO: 53) Csf3 CAGAAAGCCCTTTCCAGATAGT
(SEQ ID NO: 54) AGCCTTCTCTCTCTCTGCTCTAA
(SEQ ID NO: 55) Il-1? TCATTGTGGCTGTGGAGAAG
(SEQ ID NO: 56) GCCTGTAGTGCAGTTGTCTAA
(SEQ ID NO: 57)

FIG. 3B shows the results of measuring the expression levels of Ccr5, Il-6, Csf3 and Il-1β in a control group and a test group to which a combination of palmitic acid, LPS, and mmLDL was administered. 3b, Ccr5, Il-6, Csf3, and Il-l [beta] showed significantly higher levels of expression when palmitic acid and mmLDL were treated than when palmitic acid and mmLDL alone were treated Respectively.

5. Palmitic acid to mmLDL  Controlled by TLR4  Gene analysis of an independent pathway

(1) Palmitic acid and to mmLDL  by TLR4  Expression analysis of genes in an independent pathway 1

The effect of palmitic acid and mmLDL on TLR4-independent pathway genes is shown in a heat map. 4A is a heat map result (≥ Log 2 fold change) showing the change in gene in the TLR-4 independent pathway when palmitic acid, mmLDL or a combination thereof is administered. Figure 4b shows the results of RNA sequencing and expression levels of genes in the TLR-4 independent pathway when palmitic acid, mmLDL, or a combination thereof is administered. 4A and 4B, when palmitic acid and mmLDL were administered, the expression levels of Clec4d and Ctla2 were remarkably increased.

(2) to mmLDL  by TLR4  Expression analysis of genes in an independent pathway 2

The effect of palmitic acid and mmLDL on the expression of Clec4d and Ctla2b was determined by RT-PCR. J774 cells were seeded in a 60 mm culture dish at a concentration of 10 ⁶ cells / well and cultured at 37 ° C overnight. J774 cells were treated with 100 μM palmitic acid and cultured for 16 hours. LPS and / or 50 μg / ml of 10 ng / ml of mMLDL were added and cultured for 4 hours. RT-PCR was carried out in the same manner as in (2) of 4 except that Clec4d and Ctla2b-specific primers (Table 3) were used.

gene Forward primer (5 '-> 3') The reverse primer (5 '-> 3') Clec4d TGGAGAGCCTTCCAGTCTAA
(SEQ ID NO: 58) ATCAGCAAGTCCCAGGAAATAA
(SEQ ID NO: 59) Ctla-2b GACTCATGTGGGAGGAGAATAAG
(SEQ ID NO: 60) GGCAAATCAGGAGCCATTTC
(SEQ ID NO: 61)

4B shows the results of measuring the expression levels of Clec4d and Ctla2b in a control group and a test group to which a combination of palmitic acid, LPS, and mmLDL was administered. Referring to FIG. 4B, when both palmitic acid and mmLDL were treated, Clec4d and Ctla2b significantly increased expression levels compared to palmitic acid and mmLDL alone.

6. TLR4  The inhibitor is palmitic acid to mmLDL  On the expression of the gene

(1) Palmitic acid and to mmLDL  Expression of inflammatory genes

The effect of palmitic acid and mmLDL on the expression of genes in the NLRP3-dependent pathway was measured by RNA sequencing and RT-PCR. J774 cells were seeded in a 60 mm culture dish at a concentration of 10 ⁶ cells / well and cultured at 37 ° C overnight. J774 cells were treated with 100 μM palmitic acid and cultured for 16 hours. LPS and / or 50 μg / ml of 10 ng / ml of mMLDL were added and cultured for 4 hours. RT-PCR was performed in the same manner as in (4), except that Nlrp3-specific primers (Table 4) were used.

gene Forward primer (5 '-> 3') The reverse primer (5 '-> 3') Nlrp3 CCGTCTACGTCTTCTTCCTTTC
(SEQ ID NO: 62) CGCAGATCACACTCCTCAAATA
(SEQ ID NO: 63)

Figure 5A shows the results of RNA sequencing and expression levels of Nlrp3 when palmitic acid, mmLDL, or a combination thereof is administered. Referring to FIG. 5A, when palmitic acid and mmLDL were administered, the level of Nlrp3 expression was significantly increased compared to when palmitic acid and mmLDL were treated alone.

(2) TLR4  Analysis of expression of inflammatory genes by inhibitors

(2.1) TLR4  Analysis of expression of inflammatory genes by inhibitors

We confirmed that the increased secretion and expression of Il-1β by palmitic acid and mmLDL was due to the TLR4-dependent pathway. The TLR4 inhibitor TAK-242 was commercially available (Invivogen, San Diego, CA, USA) and dissolved in DMSO at 10 mM to prepare a stock solution.

J774 cells were seeded in a 60 mm culture dish at a concentration of 10 ⁶ cells / well and cultured at 37 ° C overnight. J774 cells were treated with 1 μM of TLR4 inhibitor and incubated for 1 hour. Subsequently, 100 μM palmitic acid was added and cultured for 16 hours, followed by 10 ng / ml of LPS and / or 50 μg / ml of mMLDL and cultured for 4 hours. RT-PCR was performed in the same manner as in (4), except that a primer specific for Il-1? Was used.

FIG. 5B shows the results of measuring the expression levels of IL-1 beta in a control group and a test group to which TLR4 inhibitor is administered and palmitic acid, LPS, mmLDL or a combination thereof is administered. Referring to FIG. 5B, when TAK-242 was administered, the expression level of Il-1 beta was significantly decreased by palmitic acid and mmLDL. Thus, the expression of the proinflammatory factor, Il-1 [beta], is dependent on the TLR4 pathway.

(2.2) TLR4  Analysis of secretion level of inflammatory factors by inhibitor

In (2) (2.1) of 6, the supernatant of the medium was obtained and the secretion level of Il-1? Was measured by ELISA. The ELISA was carried out in the same manner as in (2) (1).

FIG. 5B shows the results of measurement of secretion levels of Il-1 beta in a control group and a test group to which a TLR4 inhibitor is administered and palmitic acid, LPS, mmLDL, or a combination thereof is administered. Referring to FIG. 5B, after administration of TAK-242, the level of secretion of Il-1 beta by palmitic acid and mmLDL was somewhat reduced.

As a result, novel therapeutic targets for the treatment of cardiovascular disease were identified by identifying the key factors, Clec4d, Ctla2b or Ctla4, and Csf3, in the pathway leading to inflammatory response by palmitate and mmLDL. In addition, it is understood that an agent for inhibiting the expression or activity of Clec4d, Ctla2b or Ctla4, or Csf3 can be usefully used as a therapeutic agent for cardiovascular diseases.

7. Statistical analysis

Endotoxin levels were <0.05 EU / ㎖ in all experiments. All the above experiments were repeated three times. All data were expressed as mean ± SD. Variables and Tukey multiple comparisons were used to compare consecutive variables between groups. p <0.05 was considered statistically significant. GraphPad Prism 5.0 was used for all data analysis (GraphPad Software Inc., San Diego, Calif., USA).

<110> Industry-Academic Cooperation Foundation, Yonsei University <120> Therapeutic target for cardiovascular disease and method for          screening therapeutic agent for cardiovascular disease using the          same <130> PN117126 <160> 63 <170> Kopatentin 2.0 <210> 1 <211> 215 <212> PRT <213> Homo sapiens <400> 1 Met Gly Leu Glu Lys Pro Gln Ser Lys Leu Glu Gly Gly Met His Pro   1 5 10 15 Gln Leu Ile Pro Ser Val Ile Ala Val Val Phe Ile Leu Leu Leu Ser              20 25 30 Val Cys Phe Ile Ala Ser Cys Leu Val Thr His His Asn Phe Ser Arg          35 40 45 Cys Lys Arg Gly Thr Gly Val His Lys Leu Glu His His Ala Lys Leu      50 55 60 Lys Cys Ile Lys Glu Lys Ser Glu Leu Lys Ser Ala Glu Gly Ser Thr  65 70 75 80 Trp Asn Cys Cys Pro Ile Asp Trp Arg Ala Phe Gln Ser Asn Cys Tyr                  85 90 95 Phe Pro Leu Thr Asp Asn Lys Thr Trp Ala Glu Ser Glu Arg Asn Cys             100 105 110 Ser Gly Met Gly Ala His Leu Met Thr Ile Ser Thr Glu Ala Glu Gln         115 120 125 Asn Phe Ile Ile Gln Phe Leu Asp Arg Arg Leu Ser Tyr Phe Leu Gly     130 135 140 Leu Arg Asp Glu Asn Ala Lys Gly Gln Trp Arg Trp Val Asp Gln Thr 145 150 155 160 Pro Phe Asn Pro Arg Arg Val Phe Trp His Lys Asn Glu Pro Asp Asn                 165 170 175 Ser Gln Gly Glu Asn Cys Val Val Leu Val Tyr Asn Gln Asp Lys Trp             180 185 190 Ala Trp Asn Asp Val Pro Cys Asn Phe Glu Ala Ser Arg Ile Cys Lys         195 200 205 Ile Pro Gly Thr Thr Leu Asn     210 215 <210> 2 <211> 1973 <212> DNA <213> Homo sapiens <400> 2 ctttgaaaaa gacttctttt gagctaactt tcttatactg gtacctttct aatctcacta 60 caatatgtaa cattggtgtt cgatctcaag tatttctgaa tatattcccc tatccacaga 120 aatatactct gggggaaaaa aaatagaaca aattcttgcc gtcctgacca ttgaacaaga 180 gactaattag acaatggggc tagaaaaacc tcaaagtaaa ctggaaggag gcatgcatcc 240 ccagctgata ccttcggtta ttgctgtagt tttcatctta cttctcagtg tctgttttat 300 tgcaagttgt ttggtgactc atcacaactt ttcacgctgt aagagaggca caggagtgca 360 caagttagag caccatgcaa agctcaaatg catcaaagag aaatcagaac tgaaaagtgc 420 tgaagggagc acctggaact gttgtcctat tgactggaga gccttccagt ccaactgcta 480 ttttcctctt actgacaaca agacgtgggc tgagagtgaa aggaactgtt cagggatggg 540 ggcccatctg atgaccatca gcacggaagc tgagcagaac tttattattc agtttctgga 600 tagacggctt tcctatttcc ttggacttag agatgagaat gccaaaggtc agtggcgttg 660 ggtggaccag acgccattta acccacgcag agtattctgg cataagaatg aacccgacaa 720 ctctcaggga gaaaactgtg ttgttcttgt ttataaccaa gataaatggg cctggaatga 780 tgttccttgt aactttgaag caagtaggat ttgtaaaata cctggaacaa cattgaacta 840 gaaactcaga aagtggtcct tgtgatggaa agagaaaaga aaaaccaatt agaataaggc 900 agaatgtacg tgcgtcattg gaacacagaa aacatgctgg ttcatacagc gtttttagtc 960 ataatggtct tttttatttt gtttgattca ttcgagacaa catgtgtgta tgtgtgtgtg 1020 tgtgtgtgta gataatgtgg tttttgtatg gtgtttgatg gaaggaataa tctttctttg 1080 ctttcttagt agtatttcaa ggtgtttact tttcaattgg tgtgcactga atgcatgtat 1140 ggaagaatag cgtgaataat gcaatctctt tgtcattttt ccccttctca gactcttagc 1200 tcttaaaatt caaagatggg atattctaac tggtagtggt gcatcatttt taacccaaat 1260 attgcaagca ctttaaagat ttgaaaccac atttttattg tttgatgttt cattttcaga 1320 ctttttaatg tcagtcatta caattacatt gcatgaggaa aatttttcca gaacaacagt 1380 gtggaatagt tctgaattat gctgttctac agatagaaaa aaagtccaaa tgcctttaaa 1440 aatttacttc ttactccacc caacacgttt ttgcaaagca agaagtcttt gtaagacacc 1500 ttaaacaaag tccttcaatt ctacagcaga ggaaataaaa tcccccagaa gccaaagggc 1560 tcaccttcac attgttagtt catgacagac ccaggtgtgc ttcattagag ataacataca 1620 ttccctttgg tatcacagga agttactggg gattactcga cctcattact tagctaacga 1680 ctggataaaa tttcttaatt gtttgaagta acattgtatt cgtgtttgca ttattaattt 1740 gaatagaaaa taatcacatt ttcaacccat ttatacaaat tgttaatgtt tctttagagc 1800 tgtataacta tagtttgaac tagcaaggaa gttattgttt tgacaaccag aaattatgct 1860 tttctggtgc atgaaacatt aattgcaaag ggcagtcaca tccaacttta ataaaatatg 1920 gtggtctttc ttaaaaaaaa aaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaa 1973 <210> 3 <211> 219 <212> PRT <213> Mus musculus <400> 3 Met Trp Leu Glu Glu Ser Gln Met Lys Ser Lys Gly Thr Arg His Pro   1 5 10 15 Gln Leu Ile Pro Cys Val Phe Ala Val Val Ser Ile Ser Phe Leu Ser              20 25 30 Ala Cys Phe Ile Ser Thr Cys Leu Val Thr His His Tyr Phe Leu Arg          35 40 45 Trp Thr Arg Gly Ser Val Val Lys Leu Ser Asp Tyr His Thr Arg Val      50 55 60 Thr Cys Ile Arg Glu Glu Pro Gln Pro Gly Ala Thr Gly Gly Thr Trp  65 70 75 80 Thr Cys Cys Pro Val Ser Trp Arg Ala Phe Gln Ser Asn Cys Tyr Phe                  85 90 95 Pro Leu Asn Asp Asn Gln Thr Trp His Glu Ser Glu Arg Asn Cys Ser             100 105 110 Gly Met Ser Ser His Leu Val Thr Ile Asn Thr Glu Ala Glu Gln Asn         115 120 125 Phe Val Thr Gln Leu Leu Asp Lys Arg Phe Ser Tyr Phe Leu Gly Leu     130 135 140 Ala Asp Glu Asn Val Glu Gly Gln Trp Gln Trp Val Asp Lys Thr Pro 145 150 155 160 Phe Asn Pro His Thr Val Phe Trp Glu Lys Gly Glu Ser Asn Asp Phe                 165 170 175 Met Glu Glu Asp Cys Val Val Leu Val His Val His Glu Lys Trp Val             180 185 190 Trp Asn Phe Pro Cys His Phe Glu Val Arg Arg Ile Cys Lys Leu         195 200 205 Pro Gly Ile Thr Phe Asn Trp Lys Pro Ser Lys     210 215 <210> 4 <211> 1336 <212> DNA <213> Mus musculus <400> 4 tttccttttc ctctttgctc cctgtgttct agtacattag aggaaactct gtttttaaga 60 tttattcttt ttcacttggc tctcttgtac tgaacactta ctgatcttac cagaataaga 120 cagatgtatt gatgtccaat cttaaatgtt tctgaagaga tacttgtgat ggaaatatac 180 actctgggca agaagtaaaa agcagactgg aatctcttga cacacataat gtggctggaa 240 gaatcccaaa tgaaatcaaa aggtacccga catccccaac tgatcccttg cgtcttcgct 300 gttgtttcca tctcgttcct tagtgcttgc tttatttcaa cttgtttggt gactcatcat 360 tactttttac gctggacgag aggaagtgtg gtgaaactgt cagactacca cacgagagta 420 acgtgcatcc gagaggagcc acagcctgga gctacaggag gtacttggac ctgctgtcct 480 gttagctgga gagccttcca gtctaactgt tactttcctc ttaatgacaa ccagacctgg 540 catgagagcg agaggaactg ctcagggatg agcagtcatc tggtgaccat caacaccgaa 600 gcagaacaga attttgtgac ccagcttttg gataaacggt tttcttattt cctgggactt 660 gctgatgaga atgtggaagg ccaatggcag tgggtggaca agacgccatt taacccacac 720 acggtattct gggaaaaggg ggaatccaat gactttatgg aagaagactg tgttgtcctt 780 gttcatgtcc atgaaaaatg ggtctggaat gactttcctt gtcactttga ggtgagaagg 840 atttgtaaat tacctggaat aacattcaat tggaagccct cgaagtgatc cttgatgagg 900 atgaagaaga tgaacctatc agacccaggg aaagtgtcct tgcatcatag aaataaagaa 960 cacactgggt catagaccac tgttaaccat gaggatgttt ttcattattt ttgtttaaat 1020 ttaattcaat atacatagca taatttgtgc tctctcactc tctctgtctc tctgtgtgtg 1080 ggggtacaca ctggcatgca cacacatgct aatgcacaca tatgtttaag atagtgaggt 1140 tttgtgtcct gttttgatag agaaaatgat catcatccta tgggattcag tttttaatca 1200 ttgtttcctt gatagtattt taagaattta cttttcagtt ggtatgtttt tgttatatat 1260 ttaatgaatg gattagataa tataatcttt ttgtcatttt tctcctgcct cttaaaatta 1320 aaacttacag cagaca 1336 <210> 5 <211> 218 <212> PRT <213> Mus musculus <400> 5 Met Trp Leu Glu Glu Ser Gln Met Lys Ser Lys Gly Thr Arg His Pro   1 5 10 15 Gln Leu Ile Pro Cys Val Phe Ala Val Val Ser Ile Ser Phe Leu Ser              20 25 30 Ala Cys Phe Ile Ser Thr Cys Leu Val Thr His His Tyr Phe Leu Arg          35 40 45 Trp Thr Arg Gly Ser Val Val Lys Leu Ser Asp Tyr His Thr Arg Val      50 55 60 Thr Cys Ile Arg Glu Glu Pro Gln Pro Gly Ala Thr Gly Thr Trp Thr  65 70 75 80 Cys Cys Pro Val Ser Trp Arg Ala Phe Gln Ser Asn Cys Tyr Phe Pro                  85 90 95 Leu Asn Asp Asn Gln Thr Trp His Glu Ser Glu Arg Asn Cys Ser Gly             100 105 110 Met Ser Ser His Leu Val Thr Ile Asn Thr Glu Ala Glu Gln Asn Phe         115 120 125 Val Thr Gln Leu Leu Asp Lys Arg Phe Ser Tyr Phe Leu Gly Leu Ala     130 135 140 Asp Glu Asn Val Glu Gly Gln Trp Gln Trp Val Asp Lys Thr Pro Phe 145 150 155 160 Asn Pro His Thr Val Phe Trp Glu Lys Gly Glu Ser Asn Asp Phe Met                 165 170 175 Glu Asp Cys Val Val Leu Val His Val Glu Lys Trp Val Trp             180 185 190 Asn Asp Phe Pro Cys His Phe Glu Val Arg Arg Ile Cys Lys Leu Pro         195 200 205 Gly Ile Thr Phe Asn Trp Lys Pro Ser Lys     210 215 <210> 6 <211> 1333 <212> DNA <213> Mus musculus <400> 6 tttccttttc ctctttgctc cctgtgttct agtacattag aggaaactct gtttttaaga 60 tttattcttt ttcacttggc tctcttgtac tgaacactta ctgatcttac cagaataaga 120 cagatgtatt gatgtccaat cttaaatgtt tctgaagaga tacttgtgat ggaaatatac 180 actctgggca agaagtaaaa agcagactgg aatctcttga cacacataat gtggctggaa 240 gaatcccaaa tgaaatcaaa aggtacccga catccccaac tgatcccttg cgtcttcgct 300 gttgtttcca tctcgttcct tagtgcttgc tttatttcaa cttgtttggt gactcatcat 360 tactttttac gctggacgag aggaagtgtg gtgaaactgt cagactacca cacgagagta 420 acgtgcatcc gagaggagcc acagcctgga gctacaggta cttggacctg ctgtcctgtt 480 agctggagag ccttccagtc taactgttac tttcctctta atgacaacca gacctggcat 540 gagagcgaga ggaactgctc agggatgagc agtcatctgg tgaccatcaa caccgaagca 600 gaacagaatt ttgtgaccca gcttttggat aaacggtttt cttatttcct gggacttgct 660 gatgagaatg tggaaggcca atggcagtgg gtggacaaga cgccatttaa cccacacacg 720 gtattctggg aaaaggggga atccaatgac tttatggaag aagactgtgt tgtccttgtt 780 catgtccatg aaaaatgggt ctggaatgac tttccttgtc actttgaggt gagaaggatt 840 tgtaaattac ctggaataac attcaattgg aagccctcga agtgatcctt gatgaggatg 900 aagaagatga acctatcaga cccagggaaa gtgtccttgc atcatagaaa taaagaacac 960 actgggtcat agaccactgt taaccatgag gatgtttttc attatttttg tttaaattta 1020 attcaatata catagcataa tttgtgctct ctcactctct ctgtctctct gtgtgtgggg 1080 gtacacactg gcatgcacac acatgctaat gcacacatat gtttaagata gtgaggtttt 1140 gtgtcctgtt ttgatagaga aaatgatcat catcctatgg gattcagttt ttaatcattg 1200 tttccttgat agtattttaa gaatttactt ttcagttggt atgtttttgt tatatattta 1260 atgaatggat tagataatat aatctttttg tcatttttct cctgcctctt aaaattaaaa 1320 cttacagcag aca 1333 <210> 7 <211> 113 <212> PRT <213> Mus musculus <400> 7 Met Met Ser Ala Ala Pro Ser Pro Asp Pro Ser Leu Asp Asn Glu Trp   1 5 10 15 Lys Glu Trp Lys Thr Thr Phe Ala Lys Ala Tyr Ser Leu Asp Glu Glu              20 25 30 Arg His Arg Arg Leu Met Trp Glu Glu Asn Lys Lys Lys Ile Glu Ala          35 40 45 His Asn Asp Tyr Glu Arg Gly Lys Thr Ser Phe Tyr Met Gly Leu      50 55 60 Asn Gln Phe Ser Asp Leu Thr Pro Glu Glu Phe Arg Thr Asn Cys Cys  65 70 75 80 Gly Ser Ser Met Cys Arg Gly Glu Met Ala Pro Asp Leu Pro Glu Tyr                  85 90 95 Glu Asp Leu Gly Lys Asn Ser Tyr Leu Thr Pro Gly Arg Ala Gln Pro             100 105 110 Glu     <210> 8 <211> 856 <212> DNA <213> Mus musculus <400> 8 gccacaggct gctctcctca tgcaccacta gcctcctctg tcagttgctc ttcagaggcc 60 ctggacaaca aagttctggt ttctatctgt gaacagaagc tgcagcactt cagtgctgtc 120 ttcctgctca tcctctgctt gggaatgatg tcagctgctc catcccctga tccaagtttg 180 gataatgagt ggaaagaatg gaagacaaca tttgcaaaag cctacagtct ggatgaagaa 240 agacacagaa gactcatgtg ggaggagaat aagaagaaaa ttgaggcaca caatgcagac 300 tatgagcggg gcaagaccag cttctacatg ggcctgaatc aatttagtga cttgactcca 360 gaagaattca ggacaaattg ctgtggaagc tcaatgtgta gaggagaaat ggctcctgat 420 ttgcctgaat atgaagattt gggaaagaac agctatctga cacctggaag ggctcagcca 480 gagtaacagt tgtggcttga ctggtaacaa tatcatagtc taggagtgaa gcaagcactc 540 tgcattgtct tgggagtctt ctgtctcctg gtatgggagg aatgatgctg tagaggctca 600 acagcaggaa gcagcccatc acagtgtcca gcatgtgctg tgtgatatgc aaaactgaac 660 tcatataaga tcaacactgt ggtgtgaact ctgaggcaca gtcatactga gaaatcattg 720 ccaattttta gctctgaagt tccttaaatg attctcacaa ggatctttag ttttcacttt 780 aataagatga atatatgcca tgatgtcttc aaattaaagc tagatattaa attagtactc 840 tgcttcaaaa aaaaaa 856 <210> 9 <211> 112 <212> PRT <213> Mus musculus <400> 9 Met Met Ser Ala Ala Pro Ser Pro Asp Pro Ser Leu Asp Asn Glu Trp   1 5 10 15 Lys Glu Trp Lys Thr Thr Phe Ala Lys Ala Tyr Ser Leu Asp Glu Glu              20 25 30 Arg His Arg Arg Leu Met Trp Glu Glu Asn Lys Lys Lys Ile Glu Ala          35 40 45 His Asn Asp Tyr Glu Arg Gly Lys Thr Ser Phe Tyr Met Gly Leu      50 55 60 Asn Gln Phe Ser Asp Leu Thr Pro Glu Glu Phe Arg Thr Asn Cys Cys  65 70 75 80 Gly Ser Ser Met Cys Arg Gly Glu Met Ala Pro Asp Leu Pro Glu Tyr                  85 90 95 Glu Asp Leu Gly Lys Asn Ser Tyr Leu Thr Pro Gly Arg Ala Gln Val             100 105 110 <210> 10 <211> 1157 <212> DNA <213> Mus musculus <400> 10 gccacaggct gctctcctca tgcaccacta gcctcctctg tcagttgctc ttcagaggcc 60 ctggacaaca aagttctggt ttctatctgt gaacagaagc tgcagcactt cagtgctgtc 120 ttcctgctca tcctctgctt gggaatgatg tcagctgctc catcccctga tccaagtttg 180 gataatgagt ggaaagaatg gaagacaaca tttgcaaaag cctacagtct ggatgaagaa 240 agacacagaa gactcatgtg ggaggagaat aagaagaaaa ttgaggcaca caatgcagac 300 tatgagcggg gcaagaccag cttctacatg ggcctgaatc aatttagtga cttgactcca 360 gaagaattca ggacaaattg ctgtggaagc tcaatgtgta gaggagaaat ggctcctgat 420 ttgcctgaat atgaagattt gggaaagaac agctatctga cacctggaag ggctcaggta 480 taacagagtc acactcccac cctcagctgc ccacagaaag ctgagaagga actgaagtcc 540 tcacttgctt caggtgtgaa acaacatgca gttcactgtt tgactaagtg ttcagctctt 600 tgaaatgtgg ggagagtttt tactgtgttg tggctgacaa ccttttcttc ctgattgata 660 tactgtattt tggcattgaa cattgtgaac atgaaggcaa tatatgtcag agtagtattg 720 tgacaaagcc actgtactgc agccattcta attaatgtga tgtgtgcttt tattccagcc 780 agagtaacag ttgtggcttg actggtaaca atatcatagt ctaggagtga agcaagcact 840 ctgcattgtc ttgggagtct tctgtctcct ggtatgggag gaatgatgct gtagaggctc 900 aacagcagga agcagcccat cacagtgtcc agcatgtgct gtgtgatatg caaaactgaa 960 ctcatataag atcaacactg tggtgtgaac tctgaggcac agtcatactg agaaatcatt 1020 gccaattttt agctctgaag ttccttaaat gattctcaca aggatcttta gttttcactt 1080 taataagatg aatatatgcc atgatgtctt caaattaaag ctagatatta aattagtact 1140 ctgcttcaaa aaaaaaa 1157 <210> 11 <211> 223 <212> PRT <213> Homo sapiens <400> 11 Met Ala Cys Leu Gly Phe Gln Arg His Lys Ala Gln Leu Asn Leu Ala   1 5 10 15 Thr Arg Thr Trp Pro Cys Thr Leu Leu Phe Phe Leu Leu Phe Ile Pro              20 25 30 Val Phe Cys Lys Ala Met His Val Ala Gln Pro Ala Val Val Leu Ala          35 40 45 Ser Ser Arg Gly Ile Ala Ser Phe Val Cys Glu Tyr Ala Ser Pro Gly      50 55 60 Lys Ala Thr Glu Val Arg Val Thr Val Leu Arg Gln Ala Asp Ser Gln  65 70 75 80 Val Thr Glu Val Cys Ala Ala Thr Tyr Met Met Gly Asn Glu Leu Thr                  85 90 95 Phe Leu Asp Asp Ser Ile Cys Thr Gly Thr Ser Ser Gly Asn Gln Val             100 105 110 Asn Leu Thr Ile Gln Gly Leu Arg Ala Met Asp Thr Gly Leu Tyr Ile         115 120 125 Cys Lys Val Glu Leu Met Tyr Pro Pro Pro Tyr Tyr Leu Gly Ile Gly     130 135 140 Asn Gly Thr Gln Ile Tyr Val Ile Asp Pro Glu Pro Cys Pro Asp Ser 145 150 155 160 Asp Phe Leu Leu Trp Ile Leu Ala Ala Val Ser Ser Gly Leu Phe Phe                 165 170 175 Tyr Ser Phe Leu Leu Thr Ala Val Ser Leu Ser Lys Met Leu Lys Lys             180 185 190 Arg Ser Pro Leu Thr Thr Gly Val Tyr Val Lys Met Pro Pro Thr Glu         195 200 205 Pro Glu Cys Glu Lys Gln Phe Gln Pro Tyr Phe Ile Pro Ile Asn     210 215 220 <210> 12 <211> 2033 <212> DNA <213> Homo sapiens <400> 12 cttctgtgtg tgcacatgtg taatacatat ctgggatcaa agctatctat ataaagtcct 60 tgattctgtg tgggttcaaa cacatttcaa agcttcagga tcctgaaagg ttttgctcta 120 cttcctgaag acctgaacac cgctcccata aagccatggc ttgccttgga tttcagcggc 180 acaaggctca gctgaacctg gctaccagga cctggccctg cactctcctg ttttttcttc 240 tcttcatccc tgtcttctgc aaagcaatgc acgtggccca gcctgctgtg gtactggcca 300 gcagccgagg catcgccagc tttgtgtgtg agtatgcatc tccaggcaaa gccactgagg 360 tccgggtgac agtgcttcgg caggctgaca gccaggtgac tgaagtctgt gcggcaacct 420 acatgatggg gaatgagttg accttcctag atgattccat ctgcacgggc acctccagtg 480 gaaatcaagt gaacctcact atccaaggac tgagggccat ggacacggga ctctacatct 540 gcaaggtgga gctcatgtac ccaccgccat actacctggg cataggcaac ggaacccaga 600 tttatgtaat tgatccagaa ccgtgcccag attctgactt cctcctctgg atccttgcag 660 cagttagttc ggggttgttt ttttatagct ttctcctcac agctgtttct ttgagcaaaa 720 tgctaaagaa aagaagccct cttacaacag gggtctatgt gaaaatgccc ccaacagagc 780 cagaatgtga aaagcaattt cagccttatt ttattcccat caattgagaa accattatga 840 agaagagagt ccatatttca atttccaaga gctgaggcaa ttctaacttt tttgctatcc 900 agctattttt atttgtttgt gcatttgggg ggaattcatc tctctttaat ataaagttgg 960 atgcggaacc caaattacgt gtactacaat ttaaagcaaa ggagtagaaa gacagagctg 1020 ggatgtttct gtcacatcag ctccactttc agtgaaagca tcacttggga ttaatatggg 1080 gatgcagcat tatgatgtgg gtcaaggaat taagttaggg aatggcacag cccaaagaag 1140 gaaaaggcag ggagcgaggg agaagactat attgtacaca ccttatattt acgtatgaga 1200 cgtttatagc cgaaatgatc ttttcaagtt aaattttatg ccttttattt cttaaacaaa 1260 tgtatgatta catcaaggct tcaaaaatac tcacatggct atgttttagc cagtgatgct 1320 aaaggttgta ttgcatatat acatatatat atatatatat atatatatat atatatatat 1380 atatatatat atatatattt taatttgata gtattgtgca tagagccacg tatgtttttg 1440 tgtatttgtt aatggtttga atataaacac tatatggcag tgtctttcca ccttgggtcc 1500 cagggaagtt ttgtggagga gctcaggaca ctaatacacc aggtagaaca caaggtcatt 1560 tgctaactag cttggaaact ggatgaggtc atagcagtgc ttgattgcgt ggaattgtgc 1620 tgagttggtg ttgacatgtg ctttggggct tttacaccag ttcctttcaa tggtttgcaa 1680 ggaagccaca gctggtggta tctgagttga cttgacagaa cactgtcttg aagacaatgg 1740 cttactccag gagacccaca ggtatgacct tctaggaagc tccagttcga tgggcccaat 1800 tcttacaaac atgtggttaa tgccatggac agaagaaggc agcaggtggc agaatggggt 1860 gcatgaaggt ttctgaaaat taacactgct tgtgttttta actcaatatt ttccatgaaa 1920 atgcaacaac atgtataata tttttaatta aataaaaatc tgtggtggtc gttttaaaaa 1980 aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaaaaa 2033 <210> 13 <211> 174 <212> PRT <213> Homo sapiens <400> 13 Met Ala Cys Leu Gly Phe Gln Arg His Lys Ala Gln Leu Asn Leu Ala   1 5 10 15 Thr Arg Thr Trp Pro Cys Thr Leu Leu Phe Phe Leu Leu Phe Ile Pro              20 25 30 Val Phe Cys Lys Ala Met His Val Ala Gln Pro Ala Val Val Leu Ala          35 40 45 Ser Ser Arg Gly Ile Ala Ser Phe Val Cys Glu Tyr Ala Ser Pro Gly      50 55 60 Lys Ala Thr Glu Val Arg Val Thr Val Leu Arg Gln Ala Asp Ser Gln  65 70 75 80 Val Thr Glu Val Cys Ala Ala Thr Tyr Met Met Gly Asn Glu Leu Thr                  85 90 95 Phe Leu Asp Asp Ser Ile Cys Thr Gly Thr Ser Ser Gly Asn Gln Val             100 105 110 Asn Leu Thr Ile Gln Gly Leu Arg Ala Met Asp Thr Gly Leu Tyr Ile         115 120 125 Cys Lys Val Glu Leu Met Tyr Pro Pro Pro Tyr Tyr Leu Gly Ile Gly     130 135 140 Asn Gly Thr Gln Ile Tyr Val Ile Ala Lys Glu Lys Lys Pro Ser Tyr 145 150 155 160 Asn Arg Gly Leu Cys Glu Asn Ala Pro Asn Arg Ala Arg Met                 165 170 <210> 14 <211> 1923 <212> DNA <213> Homo sapiens <400> 14 cttctgtgtg tgcacatgtg taatacatat ctgggatcaa agctatctat ataaagtcct 60 tgattctgtg tgggttcaaa cacatttcaa agcttcagga tcctgaaagg ttttgctcta 120 cttcctgaag acctgaacac cgctcccata aagccatggc ttgccttgga tttcagcggc 180 acaaggctca gctgaacctg gctaccagga cctggccctg cactctcctg ttttttcttc 240 tcttcatccc tgtcttctgc aaagcaatgc acgtggccca gcctgctgtg gtactggcca 300 gcagccgagg catcgccagc tttgtgtgtg agtatgcatc tccaggcaaa gccactgagg 360 tccgggtgac agtgcttcgg caggctgaca gccaggtgac tgaagtctgt gcggcaacct 420 acatgatggg gaatgagttg accttcctag atgattccat ctgcacgggc acctccagtg 480 gaaatcaagt gaacctcact atccaaggac tgagggccat ggacacggga ctctacatct 540 gcaaggtgga gctcatgtac ccaccgccat actacctggg cataggcaac ggaacccaga 600 tttatgtaat tgctaaagaa aagaagccct cttacaacag gggtctatgt gaaaatgccc 660 ccaacagagc cagaatgtga aaagcaattt cagccttatt ttattcccat caattgagaa 720 accattatga agaagagag ccatatttca atttccaaga gctgaggcaa ttctaacttt 780 tttgctatcc agctattttt atttgtttgt gcatttgggg ggaattcatc tctctttaat 840 ataaagttgg atgcggaacc caaattacgt gtactacaat ttaaagcaaa ggagtagaaa 900 gacagagctg ggatgtttct gtcacatcag ctccactttc agtgaaagca tcacttggga 960 ttaatatggg gatgcagcat tatgatgtgg gtcaaggaat taagttaggg aatggcacag 1020 cccaaagaag gaaaaggcag ggagcgaggg agaagactat attgtacaca ccttatattt 1080 acgtatgaga cgtttatagc cgaaatgatc ttttcaagtt aaattttatg ccttttattt 1140 cttaaacaaa tgtatgatta catcaaggct tcaaaaatac tcacatggct atgttttagc 1200 cagtgatgct aaaggttgta ttgcatatat acatatatat atatatatat atatatatat 1260 atatatatat atatatatat atatatattt taatttgata gtattgtgca tagagccacg 1320 tatgtttttg tgtatttgtt aatggtttga atataaacac tatatggcag tgtctttcca 1380 ccttgggtcc cagggaagtt ttgtggagga gctcaggaca ctaatacacc aggtagaaca 1440 caaggtcatt tgctaactag cttggaaact ggatgaggtc atagcagtgc ttgattgcgt 1500 ggaattgtgc tgagttggtg ttgacatgtg ctttggggct tttacaccag ttcctttcaa 1560 tggtttgcaa ggaagccaca gctggtggta tctgagttga cttgacagaa cactgtcttg 1620 aagacaatgg cttactccag gagacccaca ggtatgacct tctaggaagc tccagttcga 1680 tgggcccaat tcttacaaac atgtggttaa tgccatggac agaagaaggc agcaggtggc 1740 agaatggggt gcatgaaggt ttctgaaaat taacactgct tgtgttttta actcaatatt 1800 ttccatgaaa atgcaacaac atgtataata tttttaatta aataaaaatc tgtggtggtc 1860 gttttaaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaaaaaaaaaaaaaaaaaaaaaa 1920 aaa 1923 <210> 15 <211> 207 <212> PRT <213> Homo sapiens <400> 15 Met Ala Gly Pro Ala Thr Gln Ser Pro Met Lys Leu Met Ala Leu Gln   1 5 10 15 Leu Leu Leu Trp His Ser Ala Leu Trp Thr Val Gln Glu Ala Thr Pro              20 25 30 Leu Gly Pro Ala Ser Ser Leu Pro Gln Ser Phe Leu Leu Lys Cys Leu          35 40 45 Glu Gln Val Arg Lys Ile Gln Gly Asp Gly Ala Ala Leu Gln Glu Lys      50 55 60 Leu Val Ser Glu Cys Ala Thr Tyr Lys Leu Cys His Pro Glu Glu Leu  65 70 75 80 Val Leu Leu Gly His Ser Leu Gly Ile Pro Trp Ala Pro Leu Ser Ser                  85 90 95 Cys Pro Ser Gln Ala Leu Gln Leu Ala Gly Cys Leu Ser Gln Leu His             100 105 110 Ser Gly Leu Phe Leu Tyr Gln Gly Leu Leu Gln Ala Leu Glu Gly Ile         115 120 125 Ser Pro Glu Leu Gly Pro Thr Leu Asp Thr Leu Gln Leu Asp Val Ala     130 135 140 Asp Phe Ala Thr Thr Ile Trp Gln Gln Met Glu Glu Leu Gly Met Ala 145 150 155 160 Pro Ala Leu Gln Pro Thr Gln Gly Ala Met Pro Ala Phe Ala Ser Ala                 165 170 175 Phe Gln Arg Arg Ala Gly Gly Val Leu Val Ala Ser His Leu Gln Ser             180 185 190 Phe Leu Glu Val Ser Tyr Arg Val Leu Arg His Leu Ala Gln Pro         195 200 205 <210> 16 <211> 1602 <212> DNA <213> Homo sapiens <400> 16 agtcgtggcc ccaggtaatt tcctcccagg cctccatggg gttatgtata aaggcccccc 60 tagagctggg ccccaaaaca gcccggagcc tgcagcccag ccccacccag acccatggct 120 ggacctgcca cccagagccc catgaagctg atggccctgc agctgctgct gtggcacagt 180 gcactctgga cagtgcagga agccaccccc ctgggccctg ccagctccct gccccagagc 240 ttcctgctca agtgcttaga gcaagtgagg aagatccagg gcgatggcgc agcgctccag 300 gagaagctgg tgagtgagtg tgccacctac aagctgtgcc accccgagga gctggtgctg 360 ctcggacact ctctgggcat cccctgggct cccctgagca gctgccccag ccaggccctg 420 cagctggcag gctgcttgag ccaactccat agcggccttt tcctctacca ggggctcctg 480 caggccctgg aagggatctc ccccgagttg ggtcccacct tggacacact gcagctggac 540 gtcgccgact ttgccaccac catctggcag cagatggaag aactgggaat ggcccctgcc 600 ctgcagccca cccagggtgc catgccggcc ttcgcctctg ctttccagcg ccgggcagga 660 ggggtcctgg ttgcctccca tctgcagagc ttcctggagg tgtcgtaccg cgttctacgc 720 caccttgccc agccctgagc caagccctcc ccatcccatg tatttatctc tatttaatat 780 ttatgtctat ttaagcctca tatttaaaga cagggaagag cagaacggag ccccaggcct 840 ctgtgtcctt ccctgcattt ctgagtttca ttctcctgcc tgtagcagtg agaaaaagct 900 cctgtcctcc catcccctgg actgggaggt agataggtaa ataccaagta tttattacta 960 tgactgctcc ccagccctgg ctctgcaatg ggcactggga tgagccgctg tgagcccctg 1020 gtcctgaggg tccccacctg ggacccttga gagtatcagg tctcccacgt gggagacaag 1080 aaatccctgt ttaatattta aacagcagtg ttccccatct gggtccttgc acccctcact 1140 ctggcctcag ccgactgcac agcggcccct gcatcccctt ggctgtgagg cccctggaca 1200 agcagaggtg gccagagctg ggaggcatgg ccctggggtc ccacgaattt gctggggaat 1260 ctcgtttttc ttcttaagac ttttgggaca tggtttgact cccgaacatc accgacgcgt 1320 ctcctgtttt tctgggtggc ctcgggacac ctgccctgcc cccacgaggg tcaggactgt 1380 gactcttttt agggccaggc aggtgcctgg acatttgcct tgctggacgg ggactgggga 1440 tgtgggaggg agcagacagg aggaatcatg tcaggcctgt gtgtgaaagg aagctccact 1500 gtcaccctcc acctcttcac cccccactca ccagtgtccc ctccactgtc acattgtaac 1560 tgaacttcag gataataaag tgtttgcctc caaaaaaaaa aa 1602 <210> 17 <211> 204 <212> PRT <213> Homo sapiens <400> 17 Met Ala Gly Pro Ala Thr Gln Ser Pro Met Lys Leu Met Ala Leu Gln   1 5 10 15 Leu Leu Leu Trp His Ser Ala Leu Trp Thr Val Gln Glu Ala Thr Pro              20 25 30 Leu Gly Pro Ala Ser Ser Leu Pro Gln Ser Phe Leu Leu Lys Cys Leu          35 40 45 Glu Gln Val Arg Lys Ile Gln Gly Asp Gly Ala Ala Leu Gln Glu Lys      50 55 60 Leu Cys Ala Thr Tyr Lys Leu Cys His Pro Glu Glu Leu Val Leu Leu  65 70 75 80 Gly His Ser Leu Gly Ile Pro Trp Ala Pro Leu Ser Ser Cys Pro Ser                  85 90 95 Gln Ala Leu Gln Leu Ala Gly Cys Leu Ser Gln Leu His Ser Gly Leu             100 105 110 Phe Leu Tyr Gln Gly Leu Leu Gln Ala Leu Glu Gly Ile Ser Pro Glu         115 120 125 Leu Gly Pro Thr Leu Asp Thr Leu Gln Leu Asp Val Ala Asp Phe Ala     130 135 140 Thr Ile Trp Gln Gln Met Glu Glu Leu Gly Met Ala Pro Ala Leu 145 150 155 160 Gln Pro Thr Gln Gly Ala Met Pro Ala Phe Ala Ser Ala Phe Gln Arg                 165 170 175 Arg Ala Gly Gly Val Leu Val Ala Ser His Leu Gln Ser Phe Leu Glu             180 185 190 Val Ser Tyr Arg Val Leu Arg His Leu Ala Gln Pro         195 200 <210> 18 <211> 1593 <212> DNA <213> Homo sapiens <400> 18 agtcgtggcc ccaggtaatt tcctcccagg cctccatggg gttatgtata aaggcccccc 60 tagagctggg ccccaaaaca gcccggagcc tgcagcccag ccccacccag acccatggct 120 ggacctgcca cccagagccc catgaagctg atggccctgc agctgctgct gtggcacagt 180 gcactctgga cagtgcagga agccaccccc ctgggccctg ccagctccct gccccagagc 240 ttcctgctca agtgcttaga gcaagtgagg aagatccagg gcgatggcgc agcgctccag 300 gagaagctgt gtgccaccta caagctgtgc caccccgagg agctggtgct gctcggacac 360 tctctgggca tcccctgggc tcccctgagc agctgcccca gccaggccct gcagctggca 420 ggctgcttga gccaactcca tagcggcctt ttcctctacc aggggctcct gcaggccctg 480 gaagggatct cccccgagtt gggtcccacc ttggacacac tgcagctgga cgtcgccgac 540 tttgccacca ccatctggca gcagatggaa gaactgggaa tggcccctgc cctgcagccc 600 acccagggtg ccatgccggc cttcgcctct gctttccagc gccgggcagg aggggtcctg 660 gttgcctccc atctgcagag cttcctggag gtgtcgtacc gcgttctacg ccaccttgcc 720 cagccctgag ccaagccctc cccatcccat gtatttatct ctatttaata tttatgtcta 780 tttaagcctc atatttaaag acagggaaga gcagaacgga gccccaggcc tctgtgtcct 840 tccctgcatt tctgagtttc attctcctgc ctgtagcagt gagaaaaagc tcctgtcctc 900 ccatcccctg gactgggagg tagataggta aataccaagt atttattact atgactgctc 960 cccagccctg gctctgcaat gggcactggg atgagccgct gtgagcccct ggtcctgagg 1020 gtccccacct gggacccttg agagtatcag gtctcccacg tgggagacaa gaaatccctg 1080 tttaatattt aaacagcagt gttccccatc tgggtccttg cacccctcac tctggcctca 1140 gccgactgca cagcggcccc tgcatcccct tggctgtgag gcccctggac aagcagaggt 1200 ggccagagct gggaggcatg gccctggggt cccacgaatt tgctggggaa tctcgttttt 1260 cttcttaaga cttttgggac atggtttgac tcccgaacat caccgacgcg tctcctgttt 1320 ttctgggtgg cctcgggaca cctgccctgc ccccacgagg gtcaggactg tgactctttt 1380 tagggccagg caggtgcctg gacatttgcc ttgctggacg gggactgggg atgtgggagg 1440 gagcagacag gaggaatcat gtcaggcctg tgtgtgaaag gaagctccac tgtcaccctc 1500 cacctcttca ccccccactc accagtgtcc cctccactgt cacattgtaa ctgaacttca 1560 ggataataaa gtgtttgcct ccaaaaaaaa aaa 1593 <210> 19 <211> 171 <212> PRT <213> Homo sapiens <400> 19 Met Ala Gly Pro Ala Thr Gln Ser Pro Met Lys Leu Met Ala Leu Gln   1 5 10 15 Leu Leu Leu Trp His Ser Ala Leu Trp Thr Val Gln Glu Ala Thr Pro              20 25 30 Leu Gly Pro Ala Ser Ser Leu Pro Gln Ser Phe Leu Leu Lys Cys Leu          35 40 45 Glu Gln Val Arg Lys Ile Gln Gly Asp Gly Ala Ala Leu Gln Glu Lys      50 55 60 Leu Val Ser Glu Ala Gly Cys Leu Ser Gln Leu His Ser Gly Leu Phe  65 70 75 80 Leu Tyr Gln Gly Leu Leu Gln Ala Leu Glu Gly Ile Ser Pro Glu Leu                  85 90 95 Gly Pro Thr Leu Asp Thr Leu Gln Leu Asp Val Ala Asp Phe Ala Thr             100 105 110 Thr Ile Trp Gln Gln Met Glu Glu Leu Gly Met Ala Pro Ala Leu Gln         115 120 125 Pro Thr Gln Gly Ala Met Pro Ala Phe Ala Ser Ala Phe Gln Arg Arg     130 135 140 Ala Gly Gly Val Leu Val Ala Ser His Leu Gln Ser Phe Leu Glu Val 145 150 155 160 Ser Tyr Arg Val Leu Arg His Leu Ala Gln Pro                 165 170 <210> 20 <211> 1494 <212> DNA <213> Homo sapiens <400> 20 agtcgtggcc ccaggtaatt tcctcccagg cctccatggg gttatgtata aaggcccccc 60 tagagctggg ccccaaaaca gcccggagcc tgcagcccag ccccacccag acccatggct 120 ggacctgcca cccagagccc catgaagctg atggccctgc agctgctgct gtggcacagt 180 gcactctgga cagtgcagga agccaccccc ctgggccctg ccagctccct gccccagagc 240 ttcctgctca agtgcttaga gcaagtgagg aagatccagg gcgatggcgc agcgctccag 300 gagaagctgg tgagtgaggc aggctgcttg agccaactcc atagcggcct tttcctctac 360 caggggctcc tgcaggccct ggaagggatc tcccccgagt tgggtcccac cttggacaca 420 ctgcagctgg acgtcgccga ctttgccacc accatctggc agcagatgga agaactggga 480 atggcccctg ccctgcagcc cacccagggt gccatgccgg ccttcgcctc tgctttccag 540 cgccgggcag gaggggtcct ggttgcctcc catctgcaga gcttcctgga ggtgtcgtac 600 cgcgttctac gccaccttgc ccagccctga gccaagccct ccccatccca tgtatttatc 660 tctatttaat atttatgtct atttaagcct catatttaaa gacagggaag agcagaacgg 720 agccccaggc ctctgtgtcc ttccctgcat ttctgagttt cattctcctg cctgtagcag 780 tgagaaaaag ctcctgtcct cccatcccct ggactgggag gtagataggt aaataccaag 840 tatttattac tatgactgct ccccagccct ggctctgcaa tgggcactgg gatgagccgc 900 tgtgagcccc tggtcctgag ggtccccacc tgggaccctt gagagtatca ggtctcccac 960 gtgggagaca agaaatccct gtttaatatt taaacagcag tgttccccat ctgggtcctt 1020 gcacccctca ctctggcctc agccgactgc acagcggccc ctgcatcccc ttggctgtga 1080 ggcccctgga caagcagagg tggccagagc tgggaggcat ggccctgggg tcccacgaat 1140 ttgctgggga atctcgtttt tcttcttaag acttttggga catggtttga ctcccgaaca 1200 tcaccgacgc gtctcctgtt tttctgggtg gcctcgggac acctgccctg cccccacgag 1260 gt; ggggactggg gatgtgggag ggagcagaca ggaggaatca tgtcaggcct gtgtgtgaaa 1380 ggaagctcca ctgtcaccct ccacctcttc accccccact caccagtgtc ccctccactg 1440 tcacattgta actgaacttc aggataataa agtgtttgcc tccaaaaaaa aaaa 1494 <210> 21 <211> 168 <212> PRT <213> Homo sapiens <400> 21 Met Ala Gly Pro Ala Thr Gln Ser Pro Met Lys Leu Met Ala Leu Gln   1 5 10 15 Leu Leu Leu Trp His Ser Ala Leu Trp Thr Val Gln Glu Ala Thr Pro              20 25 30 Leu Gly Pro Ala Ser Ser Leu Pro Gln Ser Phe Leu Leu Lys Cys Leu          35 40 45 Glu Gln Val Arg Lys Ile Gln Gly Asp Gly Ala Ala Leu Gln Glu Lys      50 55 60 Leu Ala Gly Cys Leu Ser Gln Leu His Ser Gly Leu Phe Leu Tyr Gln  65 70 75 80 Gly Leu Leu Gln Ala Leu Glu Gly Ile Ser Pro Glu Leu Gly Pro Thr                  85 90 95 Leu Asp Thr Leu Gln Leu Asp Val Ala Asp Phe Ala Thr Thr Ile Trp             100 105 110 Gln Gln Met Glu Glu Leu Gly Met Ala Pro Ala Leu Gln Pro Thr Gln         115 120 125 Gly Ala Met Pro Ala Phe Ala Ser Ala Phe Gln Arg Arg Ala Gly Gly     130 135 140 Val Leu Val Ala Ser His Leu Gln Ser Phe Leu Glu Val Ser Tyr Arg 145 150 155 160 Val Leu Arg His Leu Ala Gln Pro                 165 <210> 22 <211> 1485 <212> DNA <213> Homo sapiens <400> 22 agtcgtggcc ccaggtaatt tcctcccagg cctccatggg gttatgtata aaggcccccc 60 tagagctggg ccccaaaaca gcccggagcc tgcagcccag ccccacccag acccatggct 120 ggacctgcca cccagagccc catgaagctg atggccctgc agctgctgct gtggcacagt 180 gcactctgga cagtgcagga agccaccccc ctgggccctg ccagctccct gccccagagc 240 ttcctgctca agtgcttaga gcaagtgagg aagatccagg gcgatggcgc agcgctccag 300 gagaagctgg caggctgctt gagccaactc catagcggcc ttttcctcta ccaggggctc 360 ctgcaggccc tggaagggat ctcccccgag ttgggtccca ccttggacac actgcagctg 420 gacgtcgccg actttgccac caccatctgg cagcagatgg aagaactggg aatggcccct 480 gccctgcagc ccacccaggg tgccatgccg gccttcgcct ctgctttcca gcgccgggca 540 ggaggggtcc tggttgcctc ccatctgcag agcttcctgg aggtgtcgta ccgcgttcta 600 cgccaccttg cccagccctg agccaagccc tccccatccc atgtatttat ctctatttaa 660 tatttatgtc tatttaagcc tcatatttaa agacagggaa gagcagaacg gagccccagg 720 cctctgtgtc cttccctgca tttctgagtt tcattctcct gcctgtagca gtgagaaaaa 780 gctcctgtcc tcccatcccc tggactggga ggtagatagg taaataccaa gtatttatta 840 ctatgactgc tccccagccc tggctctgca atgggcactg ggatgagccg ctgtgagccc 900 ctggtcctga gggtccccac ctgggaccct tgagagtatc aggtctccca cgtgggagac 960 aagaaatccc tgtttaatat ttaaacagca gtgttcccca tctgggtcct tgcacccctc 1020 actctggcct cagccgactg cacagcggcc cctgcatccc cttggctgtg aggcccctgg 1080 acaagcagag gtggccagag ctgggaggca tggccctggg gtcccacgaa tttgctgggg 1140 aatctcgttt ttcttcttaa gacttttggg acatggtttg actcccgaac atcaccgacg 1200 cgtctcctgt ttttctgggt ggcctcggga cacctgccct gcccccacga gggtcaggac 1260 tgtgactctt tttagggcca ggcaggtgcc tggacatttg ccttgctgga cggggactgg 1320 ggatgtggga gggagcagac aggaggaatc atgtcaggcc tgtgtgtgaa aggaagctcc 1380 actgtcaccc tccacctctt caccccccac tcaccagtgt cccctccact gtcacattgt 1440 aactgaactt caggataata aagtgtttgc ctccaaaaaa aaaaa 1485 <210> 23 <211> 208 <212> PRT <213> Mus musculus <400> 23 Met Ala Gln Leu Ser Ala Gln Arg Arg Met Lys Leu Met Ala Leu Gln   1 5 10 15 Leu Leu Leu Trp Gln Ser Ala Leu Trp Ser Gly Arg Glu Ala Val Pro              20 25 30 Leu Val Thr Val Ser Ala Leu Pro Pro Ser Leu Pro Leu Pro Arg Ser          35 40 45 Phe Leu Leu Lys Ser Leu Glu Gln Val Arg Lys Ile Gln Ala Ser Gly      50 55 60 Ser Val Leu Leu Glu Gln Leu Cys Ala Thr Tyr Lys Leu Cys His Pro  65 70 75 80 Glu Glu Leu Val Leu Leu Gly His Ser Leu Gly Ile Pro Lys Ala Ser                  85 90 95 Leu Ser Gly Cys Ser Ser Gln Ala Leu Gln Gln Thr Gln Cys Leu Ser             100 105 110 Gln Leu His Ser Gly Leu Cys Leu Tyr Gln Gly Leu Leu Gln Ala Leu         115 120 125 Ser Gly Ile Ser Pro Ala Leu Ala Pro Thr Leu Asp Leu Leu Gln Leu     130 135 140 Asp Val Ala Asn Phe Ala Thr Ile Trp Gln Gln Met Glu Asn Leu 145 150 155 160 Gly Val Ala Pro Thr Val Gln Pro Thr Gln Ser Ala Met Pro Ala Phe                 165 170 175 Thr Ser Ala Phe Gln Arg Arg Ala Gly Gly Val Leu Ala Ile Ser Tyr             180 185 190 Leu Gln Gly Phe Leu Glu Thr Ala Arg Leu Ala Leu His His Leu Ala         195 200 205 <210> 24 <211> 1363 <212> DNA <213> Mus musculus <400> 24 gtataaaggc cccctggagc tgggccctgg cagagcccag agctgcagcc cagatcaccc 60 agaatccatg gctcaacttt ctgcccagag gcgcatgaag ctaatggccc tgcagctgct 120 gctgtggcaa agtgcactat ggtcaggacg agaggccgtt cccctggtca ctgtcagcgc 180 tctgccacca tccctgcctc tgccccgaag cttcctgctt aagtccctgg agcaagtgag 240 gaagatccag gccagcggct cggtgctgct ggagcagttg tgtgccacct acaagctgtg 300 tcaccccgag gagctggtgt tgctgggcca ctctctggg atcccgaagg cttccctgag 360 tggctgctct agccaggccc tgcagcagac acagtgccta agccagctcc acagtgggct 420 ctgcctctac caaggtctcc tgcaggctct atcgggtatt tcccctgccc tggcccccac 480 cttggacttg cttcagctgg atgttgccaa ctttgccacc accatctggc agcagatgga 540 aaacctaggg gtggccccta ctgtgcagcc cacacagagc gccatgccag ccttcacttc 600 tgccttccag cgccgggcag gaggtgtcct ggccatttcg tacctgcagg gcttcctgga 660 gacggctcgc cttgctctgc accacttggc ctagacctga gcagaaagcc ctttccagat 720 agtttattta tctctattta atatttatgc atatttaagc ctactattta aagacaaaga 780 cgagaaaatg gagctctaag cttctagatc attctctcca cttccgagtt ttgttctcct 840 gcttagagca gagagagaag gctcttgtgt cctcctgtgg aggccaggga aggagatggg 900 taaataccaa gtattgattc ctgctgctgc tccaggcacc cagttctgtg gcagtacccc 960 caaaaaatca gtgagccctg ccgtgctgag gcaccatctc aggggggccc aggcagcatc 1020 tggtctccct tccgggggac aagacatccc tgtttaatat ttaaacagca gtgttcccaa 1080 actgggttct tatatccctt gctctggtca accaggttgc agggtttcct gtcctcacag 1140 gaacgaagtc cctaaagaaa cagtggcagc caggtttagc cccggaattg actggattcc 1200 ttttttaggg ccctgctggc ctggaagttg gagtgggggg cagaggaggc aggaggaagc 1260 ctgggggggg ggttggcatg gagggaggcc ttcccatcca ccctcaccct ccaccccacc 1320 tgtcactata gccaagcttg cggataataa agtgtggtgt tcc 1363 <210> 25 <211> 107 <212> PRT <213> Homo sapiens <400> 25 Met Ala Arg Ala Thr Leu Ser Ala Ala Pro Ser Asn Pro Arg Leu Leu   1 5 10 15 Arg Val Ala Leu Leu Leu Leu Leu              20 25 30 Ala Gly Ala Pro Leu Ala Thr Glu Leu Arg Cys Gln Cys Leu Gln Thr          35 40 45 Leu Gln Gly Ile His Leu Lys Asn Ile Gln Ser Val Lys Val Lys Ser      50 55 60 Pro Gly Pro His Cys Ala Gln Thr Glu Val Ile Ala Thr Leu Lys Asn  65 70 75 80 Gly Gln Lys Ala Cys Leu Asn Pro Ala Ser Pro Met Val Lys Lys Ile                  85 90 95 Ile Glu Lys Met Leu Lys Asn Gly Lys Ser Asn             100 105 <210> 26 <211> 100 <212> PRT <213> Mus musculus <400> 26 Met Ala Pro Pro Thr Cys Arg Leu Leu Ser Ala Leu Val Leu Leu   1 5 10 15 Leu Leu Leu Ala Thr Asn His Gln Ala Thr Gly Ala Val Val Ala Ser              20 25 30 Glu Leu Arg Cys Gln Cys Leu Lys Thr Leu Pro Arg Val Asp Phe Lys          35 40 45 Asn Ile Gln Ser Leu Ser Val Thr Pro Pro Gly Pro His Cys Ala Gln      50 55 60 Thr Glu Val Ile Ala Thr Leu Lys Gly Gly Gln Lys Val Cys Leu Asp  65 70 75 80 Pro Glu Ala Pro Leu Val Gln Lys Ile Ile Gln Lys Ile Leu Asn Lys                  85 90 95 Gly Lys Ala Asn             100 <210> 27 <211> 233 <212> PRT <213> Homo sapiens <400> 27 Met Ser Thr Glu Ser Met Ile Arg Asp Val Glu Leu Ala Glu Glu Ala   1 5 10 15 Leu Pro Lys Lys Thr Gly Gly Pro Gln Gly Ser Arg Arg Cys Leu Phe              20 25 30 Leu Ser Leu Phe Ser Phe Leu Ile Val Ala Gly Ala Thr Thr Leu Phe          35 40 45 Cys Leu Leu His Phe Gly Val Ile Gly Pro Gln Arg Glu Glu Phe Pro      50 55 60 Arg Asp Leu Ser Leu Ile Ser Pro Leu Ala Gln Ala Val Arg Ser Ser  65 70 75 80 Ser Arg Thr Pro Ser Asp Lys Pro Val Ala His Val Val Ala Asn Pro                  85 90 95 Gln Ala Glu Gly Gln Leu Gln Trp Leu Asn Arg Arg Ala Asn Ala Leu             100 105 110 Leu Ala Asn Gly Val Glu Leu Arg Asp Asn Gln Leu Val Val Ser Ser         115 120 125 Glu Gly Leu Tyr Leu Ile Tyr Ser Gln Val Leu Phe Lys Gly Gln Gly     130 135 140 Cys Pro Ser Thr His Val Leu Leu Thr His Thr Ile Ser Arg Ile Ala 145 150 155 160 Val Ser Tyr Gln Thr Lys Val Asn Leu Leu Ser Ala Ile Lys Ser Pro                 165 170 175 Cys Gln Arg Glu Thr Pro Glu Gly Ala Glu Ala Lys Pro Trp Tyr Glu             180 185 190 Pro Ile Tyr Leu Gly Gly Val Phe Gln Leu Glu Lys Gly Asp Arg Leu         195 200 205 Ser Ala Glu Ile Asn Arg Pro Asp Tyr Leu Asp Phe Ala Glu Ser Gly     210 215 220 Gln Val Tyr Phe Gly Ile Ile Ala Leu 225 230 <210> 28 <211> 235 <212> PRT <213> Mus musculus <400> 28 Met Ser Thr Glu Ser Met Ile Arg Asp Val Glu Leu Ala Glu Glu Ala   1 5 10 15 Leu Pro Gln Lys Met Gly Gly Phe Gln Asn Ser Arg Arg Cys Leu Cys              20 25 30 Leu Ser Leu Phe Ser Phe Leu Le A Val Ala Gly Ala Thr Thr Leu Phe          35 40 45 Cys Leu Leu Asn Phe Gly Val Ile Gly Pro Gln Arg Asp Glu Lys Phe      50 55 60 Pro Asn Gly Leu Pro Leu Ile Ser Ser Ale Gln Thr Leu Thr Leu  65 70 75 80 Arg Ser Ser Ser Gln Asn Ser Ser Asp Lys Pro Val Ala His Val Val                  85 90 95 Ala Asn His Gln Val Glu Glu Gln Leu Glu Trp Leu Ser Gln Arg Ala             100 105 110 Asn Ala Leu Leu Ala Asn Gly Met Asp Leu Lys Asp Asn Gln Leu Val         115 120 125 Val Pro Ala Asp Gly Leu Tyr Leu Val Tyr Ser Gln Val Leu Phe Lys     130 135 140 Gly Gln Gly Cys Pro Asp Tyr Val Leu Leu Thr His Thr Val Ser Arg 145 150 155 160 Phe Ala Ile Ser Tyr Gln Glu Lys Val Asn Leu Leu Ser Ala Val Lys                 165 170 175 Ser Pro Cys Pro Lys Asp Thr Pro Glu Gly Ala Glu Leu Lys Pro Trp             180 185 190 Tyr Glu Pro Ile Tyr Leu Gly Gly Val Phe Gln Leu Glu Lys Gly Asp         195 200 205 Gln Leu Ser Ala Glu Val Asn Leu Pro Lys Tyr Leu Asp Phe Ala Glu     210 215 220 Ser Gly Gln Val Tyr Phe Gly Val Ile Ala Leu 225 230 235 <210> 29 <211> 219 <212> PRT <213> Mus musculus <400> 29 Met Ser Thr Glu Ser Met Ile Arg Asp Val Glu Leu Ala Glu Glu Ala   1 5 10 15 Leu Pro Gln Lys Met Gly Gly Phe Gln Asn Ser Arg Arg Cys Leu Cys              20 25 30 Leu Ser Leu Phe Ser Phe Leu Le A Val Ala Gly Ala Thr Thr Leu Phe          35 40 45 Cys Leu Leu Asn Phe Gly Val Ile Gly Pro Gln Arg Asp Glu Lys Phe      50 55 60 Pro Asn Gly Leu Pro Leu Ile Ser Ser Ale Gln Thr Leu Thr Leu  65 70 75 80 Thr Asn His Gln Val Glu Glu Gln Leu Glu Trp Leu Ser Gln Arg Ala                  85 90 95 Asn Ala Leu Leu Ala Asn Gly Met Asp Leu Lys Asp Asn Gln Leu Val             100 105 110 Val Pro Ala Asp Gly Leu Tyr Leu Val Tyr Ser Gln Val Leu Phe Lys         115 120 125 Gly Gln Gly Cys Pro Asp Tyr Val Leu Leu Thr His Thr Val Ser Arg     130 135 140 Phe Ala Ile Ser Tyr Gln Glu Lys Val Asn Leu Leu Ser Ala Val Lys 145 150 155 160 Ser Pro Cys Pro Lys Asp Thr Pro Glu Gly Ala Glu Leu Lys Pro Trp                 165 170 175 Tyr Glu Pro Ile Tyr Leu Gly Gly Val Phe Gln Leu Glu Lys Gly Asp             180 185 190 Gln Leu Ser Ala Glu Val Asn Leu Pro Lys Tyr Leu Asp Phe Ala Glu         195 200 205 Ser Gly Gln Val Tyr Phe Gly Val Ile Ala Leu     210 215 <210> 30 <211> 587 <212> PRT <213> Homo sapiens <400> 30 Met Ala Glu Lys Gly Asp Cys Ile Ala Ser Val Tyr Gly Tyr Asp Leu   1 5 10 15 Gly Gly Arg Phe Val Asp Phe Gln Pro Leu Gly Phe Gly Val Asn Gly              20 25 30 Leu Val Leu Ser Ala Val Asp Ser Arg Ala Cys Arg Lys Val Ala Val          35 40 45 Lys Lys Ile Ala Leu Ser Asp Ala Arg Ser Met Lys His Ala Leu Arg      50 55 60 Glu Ile Lys Ile Ile Arg Arg Leu Asp His Asp Asn Ile Val Lys Val  65 70 75 80 Tyr Glu Val Leu Gly Pro Lys Gly Thr Asp Leu Gln Gly Glu Leu Phe                  85 90 95 Lys Phe Ser Val Ala Tyr Ile Val Gln Glu Tyr Met Glu Thr Asp Leu             100 105 110 Ala Arg Leu Leu Glu Gln Gly Thr Leu Ala Glu Glu His Ala Lys Leu         115 120 125 Phe Met Tyr Gln Leu Leu Arg Gly Leu Lys Tyr Ile His Ser Ala Asn     130 135 140 Val Leu His Arg Asp Leu Lys Pro Ala Asn Ile Phe Ile Ser Thr Glu 145 150 155 160 Asp Leu Val Leu Lys Ile Gly Asp Phe Gly Leu Ala Arg Ile Val Asp                 165 170 175 Gln His Tyr Ser His Lys Gly Tyr Leu Ser Glu Gly Leu Val Thr Lys             180 185 190 Trp Tyr Arg Ser Pro Arg Leu Leu Leu Ser Pro Asn Asn Tyr Thr Lys         195 200 205 Ala Ile Asp Met Trp Ala Ala Gly Cys Ile Leu Ala Glu Met Leu Thr     210 215 220 Gly Arg Met Leu Phe Ala Gly Ala His Glu Leu Glu Gln Met Gln Leu 225 230 235 240 Ile Leu Glu Thr Ile Pro Val Ile Arg Glu Glu Asp Lys Asp Glu Leu                 245 250 255 Leu Arg Val Met Pro Ser Phe Val Ser Ser Thr Trp Glu Val Lys Arg             260 265 270 Pro Leu Arg Lys Leu Leu Pro Glu Val Asn Ser Glu Ala Ile Asp Phe         275 280 285 Leu Glu Lys Ile Leu Thr Phe Asn Pro Met Asp Arg Leu Thr Ala Glu     290 295 300 Met Gly Leu Gln His Pro Tyr Met Ser Pro Tyr Ser Cys Pro Glu Asp 305 310 315 320 Glu Pro Thr Ser Gln His Pro Phe Arg Ile Glu Asp Glu Ile Asp Asp                 325 330 335 Ile Val Leu Met Ala Ala Asn Gln Ser Gln Leu Ser Asn Trp Asp Thr             340 345 350 Cys Ser Ser Arg Tyr Pro Val Ser Leu Ser Ser Asp Leu Glu Trp Arg         355 360 365 Pro Asp Arg Cys Gln Asp Ala Ser Glu Val Gln Arg Asp Pro Arg Ala     370 375 380 Gly Ser Ala Pro Leu Ala Glu Asp Val Gln Val Asp Pro Arg Lys Asp 385 390 395 400 Ser His Ser Ser Glu Arg Phe Leu Glu Gln Ser His Ser Ser Met                 405 410 415 Glu Arg Ala Phe Glu Ala Asp Tyr Gly Arg Ser Cys Asp Tyr Lys Val             420 425 430 Gly Ser Pro Ser Tyr Leu Asp Lys Leu Leu Trp Arg Asp Asn Lys Pro         435 440 445 His His Tyr Ser Glu Pro Lys Leu Ile Leu Asp Leu Ser His Trp Lys     450 455 460 Gln Ala Ala Gly Ala Pro Pro Thr Ala Thr Gly Leu Ala Asp Thr Gly 465 470 475 480 Ala Arg Glu Asp Glu Pro Ala Ser Leu Phe Leu Glu Ile Ala Gln Trp                 485 490 495 Val Lys Ser Thr Gln Gly Gly Pro Glu His Ala Ser Pro Pro Ala Asp             500 505 510 Asp Pro Glu Arg Arg Leu Ser Ala Ser Pro Pro Gly Arg Pro Ala Pro         515 520 525 Val Asp Gly Gly Ala Ser Pro Gln Phe Asp Leu Asp Val Phe Ile Ser     530 535 540 Arg Ala Leu Lys Leu Cys Thr Lys Pro Glu Asp Leu Pro Asp Asn Lys 545 550 555 560 Leu Gly Asp Leu Asn Gly Ala Cys Ile Pro Gly His Pro Gly Asp Leu                 565 570 575 Val Gln Thr Glu Ala Phe Ser Lys Glu Arg Trp             580 585 <210> 31 <211> 233 <212> PRT <213> Homo sapiens <400> 31 Met Ala Glu Lys Gly Asp Cys Ile Ala Ser Val Tyr Gly Tyr Asp Leu   1 5 10 15 Gly Gly Arg Phe Val Asp Phe Gln Pro Leu Gly Phe Gly Val Asn Gly              20 25 30 Leu Val Leu Ser Ala Val Asp Ser Arg Ala Cys Arg Lys Val Ala Val          35 40 45 Lys Lys Ile Ala Leu Ser Asp Ala Arg Ser Met Lys His Ala Leu Arg      50 55 60 Glu Ile Lys Ile Ile Arg Arg Leu Asp His Asp Asn Ile Val Lys Val  65 70 75 80 Tyr Glu Val Leu Gly Pro Lys Gly Thr Asp Leu Gln Gly Glu Leu Phe                  85 90 95 Lys Phe Ser Val Ala Tyr Ile Val Gln Glu Tyr Met Glu Thr Asp Leu             100 105 110 Ala Arg Leu Leu Glu Gln Gly Thr Leu Ala Glu Glu His Ala Lys Leu         115 120 125 Phe Met Tyr Gln Leu Leu Arg Gly Leu Lys Tyr Ile His Ser Ala Asn     130 135 140 Val Leu His Arg Asp Leu Lys Pro Ala Asn Ile Phe Ile Ser Thr Glu 145 150 155 160 Asp Leu Val Leu Lys Ile Gly Asp Phe Gly Leu Ala Arg Ile Val Asp                 165 170 175 Gln His Tyr Ser His Lys Gly Tyr Leu Ser Glu Gly Leu Val Thr Lys             180 185 190 Trp Tyr Arg Ser Pro Arg Leu Leu Leu Ser Pro Asn Asn Tyr Thr Lys         195 200 205 Ala Ile Asp Met Trp Ala Ala Gly Cys Ile Leu Ala Glu Met Leu Thr     210 215 220 Gly Arg Met Leu Phe Ala Gly Thr Leu 225 230 <210> 32 <211> 376 <212> PRT <213> Homo sapiens <400> 32 Met Trp Ala Ala Gly Cys Ile Leu Ala Glu Met Leu Thr Gly Arg Met   1 5 10 15 Leu Phe Ala Gly Ala His Glu Leu Glu Gln Met Gln Leu Ile Leu Glu              20 25 30 Thr Ile Pro Val Ile Arg Glu Glu Asp Lys Asp Glu Leu Leu Arg Val          35 40 45 Met Pro Ser Phe Val Ser Ser Thr Trp Glu Val Lys Arg Pro Leu Arg      50 55 60 Lys Leu Leu Pro Glu Val Asn Ser Glu Ala Ile Asp Phe Leu Glu Lys  65 70 75 80 Ile Leu Thr Phe Asn Pro Met Asp Arg Leu Thr Ala Glu Met Gly Leu                  85 90 95 Gln His Pro Tyr Met Ser Pro Tyr Ser Cys Pro Glu Asp Glu Pro Thr             100 105 110 Ser Gln His Pro Phe Arg Ile Glu Asp Glu Ile Asp Asp Ile Val Leu         115 120 125 Met Ala Ala Asn Gln Ser Gln Leu Ser Asn Trp Asp Thr Cys Ser Ser     130 135 140 Arg Tyr Pro Val Ser Leu Ser Ser Asp Leu Glu Trp Arg Pro Asp Arg 145 150 155 160 Cys Gln Asp Ala Ser Glu Val Gln Arg Asp Pro Arg Ala Gly Ser Ala                 165 170 175 Pro Leu Ala Glu Asp Val Gln Val Asp Pro Arg Lys Asp Ser His Ser             180 185 190 Ser Ser Glu Arg Phe Leu Glu Gln Ser His Ser Ser Met Glu Arg Ala         195 200 205 Phe Glu Ala Asp Tyr Gly Arg Ser Cys Asp Tyr Lys Val Gly Ser Pro     210 215 220 Ser Tyr Leu Asp Lys Leu Leu Trp Arg Asp Asn Lys Pro His His Tyr 225 230 235 240 Ser Glu Pro Lys Leu Ile Leu Asp Leu Ser His Trp Lys Gln Ala Ala                 245 250 255 Gly Ala Pro Pro Thr Ala Thr Gly Leu Ala Asp Thr Gly Ala Arg Glu             260 265 270 Asp Glu Pro Ala Ser Leu Phe Leu Glu Ile Ala Gln Trp Val Lys Ser         275 280 285 Thr Gln Gly Gly Pro Glu His Ala Ser Pro Pro Ala Asp Asp Pro Glu     290 295 300 Arg Arg Leu Ser Ala Ser Pro Pro Gly Arg Pro Ala Pro Val Asp Gly 305 310 315 320 Gly Ala Ser Pro Gln Phe Asp Leu Asp Val Phe Ile Ser Arg Ala Leu                 325 330 335 Lys Leu Cys Thr Lys Pro Glu Asp Leu Pro Asp Asn Lys Leu Gly Asp             340 345 350 Leu Asn Gly Ala Cys Ile Pro Glu His Pro Gly Asp Leu Val Gln Thr         355 360 365 Glu Ala Phe Ser Lys Glu Arg Trp     370 375 <210> 33 <211> 721 <212> PRT <213> Homo sapiens <400> 33 Met Ala Glu Lys Phe Glu Ser Leu Met Asn Ile His Gly Phe Asp Leu   1 5 10 15 Gly Ser Arg Tyr Met Asp Leu Lys Pro Leu Gly Cys Gly Gly Asn Gly              20 25 30 Leu Val Phe Ser Ala Val Asp Asn Asp Cys Asp Lys Arg Val Ala Ile          35 40 45 Lys Lys Ile Val Leu Thr Asp Pro Gln Ser Val Lys His Ala Leu Arg      50 55 60 Glu Ile Lys Ile Ile Arg Arg Leu Asp His Asp Asn Ile Val Lys Val  65 70 75 80 Phe Glu Ile Leu Gly Pro Ser Gly Ser Gln Leu Thr Asp Asp Val Gly                  85 90 95 Ser Leu Thr Glu Leu Asn Ser Val Tyr Ile Val Gln Glu Tyr Met Glu             100 105 110 Thr Asp Leu Ala Asn Val Leu Glu Gln Gly Pro Leu Leu Glu Glu His         115 120 125 Ala Arg Leu Phe Met Tyr Gln Leu Leu Arg Gly Leu Lys Tyr Ile His     130 135 140 Ser Ala Asn Val Leu His Arg Asp Leu Lys Pro Ala Asn Leu Phe Ile 145 150 155 160 Asn Thr Glu Asp Leu Val Leu Lys Ile Gly Asp Phe Gly Leu Ala Arg                 165 170 175 Ile Met Asp Pro His Tyr Ser His Lys Gly His Leu Ser Glu Gly Leu             180 185 190 Val Thr Lys Trp Tyr Arg Ser Pro Arg Leu Leu Leu Ser Pro Asn Asn         195 200 205 Tyr Thr Lys Ala Ile Asp Met Trp Ala Ala Gly Cys Ile Phe Ala Glu     210 215 220 Met Leu Thr Gly Lys Thr Leu Phe Ala Gly Ala His Glu Leu Glu Gln 225 230 235 240 Met Gln Leu Ile Leu Glu Ser Ile Pro Val Val His Glu Glu Asp Arg                 245 250 255 Gln Glu Leu Leu Ser Val Ile Pro Val Tyr Ile Arg Asn Asp Met Thr             260 265 270 Glu Pro His Lys Pro Leu Thr Gln Leu Leu Pro Gly Ile Ser Arg Glu         275 280 285 Ala Leu Asp Phe Leu Glu Gln Ile Leu Thr Phe Ser Pro Met Asp Arg     290 295 300 Leu Thr Ala Glu Glu Ala Leu Ser His Pro Tyr Met Ser Ile Tyr Ser 305 310 315 320 Phe Pro Met Asp Glu Pro Ile Ser Ser His Pro Phe His Ile Glu Asp                 325 330 335 Glu Val Asp Asp Ile Leu Leu Met Asp Glu Thr His Ser His Ile Tyr             340 345 350 Asn Trp Glu Arg Tyr His Asp Cys Gln Phe Ser Glu His Asp Trp Pro         355 360 365 Val His Asn Asn Phe Asp Ile Asp Glu Val Gln Leu Asp Pro Arg Ala     370 375 380 Leu Ser Asp Val Thr Asp Glu Glu Glu Val Gln Val Asp Pro Arg Lys 385 390 395 400 Tyr Leu Asp Gly Asp Arg Glu Lys Tyr Leu Glu Asp Pro Ala Phe Asp                 405 410 415 Thr Asn Tyr Ser Thr Glu Pro Cys Trp Gln Tyr Ser Asp His His Glu             420 425 430 Asn Lys Tyr Cys Asp Leu Glu Cys Ser His Thr Cys Asn Tyr Lys Thr         435 440 445 Arg Ser Ser Ser Tyr Leu Asp Asn Leu Val Trp Arg Glu Ser Glu Val     450 455 460 Asn His Tyr Tyr Glu Pro Lys Leu Ile Ile Asp Leu Ser Asn Trp Lys 465 470 475 480 Glu Gln Ser Lys Glu Lys Ser Asp Lys Lys Gly Lys Ser Lys Cys Glu                 485 490 495 Arg Asn Gly Leu Val Lys Ala Gln Ile Ala Leu Glu Glu Ala Ser Gln             500 505 510 Gln Leu Ala Gly Lys Glu Arg Glu Lys Asn Gln Gly Phe Asp Phe Asp         515 520 525 Ser Phe Ile Ala Gly Thr Ile Gln Leu Ser Ser Gln His Glu Pro Thr     530 535 540 Asp Val Val Asp Lys Leu Asn Asp Leu Asn Ser Ser Val Ser Gln Leu 545 550 555 560 Glu Leu Lys Ser Leu Ile Ser Lys Ser Val Ser Gln Glu Lys Gln Glu                 565 570 575 Lys Gly Met Ala Asn Leu Ala Gln Leu Glu Ala Leu Tyr Gln Ser Ser             580 585 590 Trp Asp Ser Gln Phe Val Ser Gly Gly Glu Asp Cys Phe Phe Ile Asn         595 600 605 Gln Phe Cys Glu Val Arg Lys Asp Glu Gln Val Glu Lys Glu Asn Thr     610 615 620 Tyr Thr Ser Tyr Leu Asp Lys Phe Phe Ser Arg Lys Glu Asp Thr Glu 625 630 635 640 Met Leu Glu Thr Glu Pro Val Glu Asp Gly Lys Leu Gly Glu Arg Gly                 645 650 655 His Glu Glu Gly Phe Leu Asn Asn Ser Gly Glu Phe Leu Phe Asn Lys             660 665 670 Gln Leu Glu Ser Ile Gly Ile Pro Gln Phe His Ser Pro Val Gly Ser         675 680 685 Pro Leu Lys Ser Ile Gln Ala Thr Leu Thr Ser Ser Ala Met Lys Ser     690 695 700 Ser Pro Gln Ile Pro His Gln Thr Tyr Ser Ser Ile Leu Lys His Leu 705 710 715 720 Asn     <210> 34 <211> 478 <212> PRT <213> Mus musculus <400> 34 Met Ser Leu His Phe Leu Tyr Tyr Cys Ser Glu Pro Thr Leu Asp Val   1 5 10 15 Lys Ile Ala Phe Cys Gln Gly Phe Asp Lys His Val Asp Val Ser Ser              20 25 30 Ile Ala Lys His Tyr Asn Met Ser Lys Ser Lys Val Asp Asn Gln Phe          35 40 45 Tyr Ser Val Glu Val Gly Asp Ser Thr Phe Thr Val Leu Lys Arg Tyr      50 55 60 Gln Asn Leu Lys Pro Ile Gly Ser Gly Ala Gln Gly Ile Val Cys Ala  65 70 75 80 Ala Tyr Asp Ala Val Leu Asp Arg Asn Val Ala Ile Lys Lys Leu Ser                  85 90 95 Arg Pro Phe Gln Asn Gln Thr His Ala Lys Arg Ala Tyr Arg Glu Leu             100 105 110 Val Leu Met Lys Cys Val Asn His Lys Asn Ile Ile Ser Leu Leu Asn         115 120 125 Val Phe Thr Pro Gln Lys Thr Leu Glu Glu Phe Gln Asp Val Tyr Leu     130 135 140 Val Met Glu Leu Met Asp Ala Asn Leu Cys Gln Val Ile Gln Met Glu 145 150 155 160 Leu Asp His Glu Arg Met Ser Tyr Leu Leu Tyr Gln Met Leu Cys Gly                 165 170 175 Ile Lys His Leu His Ser Ala Gly Ile Ile His Arg Asp Leu Lys Pro             180 185 190 Ser Asn Ile Val Val Lys Ser Asp Cys Thr Leu Lys Ile Leu Asp Phe         195 200 205 Gly Leu Ala Arg Thr Ala Gly Thr Ser Phe Met Met Thr Pro Tyr Val     210 215 220 Val Thr Arg Tyr Tyr Arg Ala Pro Glu Val Ile Leu Gly Met Gly Tyr 225 230 235 240 Lys Glu Asn Val Asp Ile Trp Ser Val Gly Cys Ile Met Gly Glu Met                 245 250 255 Val Arg His Lys Ile Leu Phe Pro Gly Arg Asp Tyr Ile Asp Gln Trp             260 265 270 Asn Lys Val Ile Glu Gln Leu Gly Thr Pro Cys Pro Glu Phe Met Lys         275 280 285 Lys Leu Gln Pro Thr Val Arg Asn Tyr Val Glu Asn Arg Pro Lys Tyr     290 295 300 Ala Gly Leu Thr Phe Pro Lys Leu Phe Pro Asp Ser Leu Phe Pro Ala 305 310 315 320 Asp Ser Glu His Asn Lys Leu Lys Ala Ser Gln Ala Arg Asp Leu Leu                 325 330 335 Ser Lys Met Leu Val Ile Asp Pro Ala Lys Arg Ile Ser Val Asp Asp             340 345 350 Ala Leu Gln His Pro Tyr Ile Asn Val Trp Tyr Asp Pro Ala Glu Val         355 360 365 Glu Ala Pro Pro Gln Ile Tyr Asp Lys Gln Leu Asp Glu Arg Glu     370 375 380 His Thr Ile Glu Glu Trp Lys Glu Leu Ile Tyr Lys Glu Val Met Asn 385 390 395 400 Ser Glu Glu Lys Thr Lys Asn Gly Val Val Lys Gly Gln Pro Ser Pro                 405 410 415 Ser Gly Ala Ala Val Asn Ser Ser Glu Ser Leu Pro Ser Ser Ser             420 425 430 Val Asn Asp Ile Ser Ser Met Ser Thr Asp Gln Thr Leu Ala Ser Asp         435 440 445 Thr Asp Ser Ser Leu Glu Ala Ser Ala Gly Pro Leu Gly Cys Cys Arg     450 455 460 Xaa Leu Ala Cys Leu Arg Asn Pro Ala Phe Phe Arg Arg 465 470 475 <210> 35 <211> 720 <212> PRT <213> Mus musculus <400> 35 Met Ala Glu Lys Phe Glu Ser Leu Met Asn Ile His Gly Phe Asp Leu   1 5 10 15 Gly Ser Arg Tyr Met Asp Leu Lys Pro Leu Gly Cys Gly Gly Asn Gly              20 25 30 Leu Val Phe Ser Ala Val Asp Asn Asp Cys Asp Lys Arg Val Ala Ile          35 40 45 Lys Lys Ile Val Leu Thr Asp Pro Gln Ser Val Lys His Ala Leu Arg      50 55 60 Glu Ile Lys Ile Ile Arg Arg Leu Asp His Asp Asn Ile Val Lys Val  65 70 75 80 Phe Glu Ile Leu Gly Pro Ser Gly Ser Gln Leu Thr Asp Asp Val Gly                  85 90 95 Ser Leu Thr Glu Leu Asn Ser Val Tyr Ile Val Gln Glu Tyr Met Glu             100 105 110 Thr Asp Leu Ala Asn Val Leu Glu Gln Gly Pro Leu Leu Glu Glu His         115 120 125 Ala Arg Leu Phe Met Tyr Gln Leu Leu Arg Gly Leu Lys Tyr Ile His     130 135 140 Ser Ala Asn Val Leu His Arg Asp Leu Lys Pro Ala Asn Leu Phe Ile 145 150 155 160 Asn Thr Glu Asp Leu Val Leu Lys Ile Gly Asp Phe Gly Leu Ala Arg                 165 170 175 Ile Met Asp Pro His Tyr Ser His Lys Gly His Leu Ser Glu Gly Leu             180 185 190 Val Thr Lys Trp Tyr Arg Ser Pro Arg Leu Leu Leu Ser Pro Asn Asn         195 200 205 Tyr Thr Lys Ala Ile Asp Met Trp Ala Ala Gly Cys Ile Phe Ala Glu     210 215 220 Met Leu Thr Gly Lys Thr Leu Phe Ala Gly Ala His Glu Leu Glu Gln 225 230 235 240 Met Gln Leu Ile Leu Asp Ser Ile Pro Val Val His Glu Glu Asp Arg                 245 250 255 Gln Glu Leu Leu Ser Val Ile Pro Val Tyr Ile Arg Asn Asp Met Thr             260 265 270 Glu Pro His Arg Pro Leu Thr Gln Leu Leu Pro Gly Ile Ser Arg Glu         275 280 285 Ala Leu Asp Phe Leu Glu Gln Ile Leu Thr Phe Ser Pro Met Asp Arg     290 295 300 Leu Thr Ala Glu Glu Ala Leu Ser His Pro Tyr Met Ser Ile Tyr Ser 305 310 315 320 Phe Pro Thr Asp Glu Pro Ile Ser Ser His Pro Phe His Ile Glu Asp                 325 330 335 Glu Val Asp Asp Ile Leu Leu Met Asp Glu Thr His Ser His Ile Tyr             340 345 350 Asn Trp Glu Arg Tyr His Asp Cys Gln Phe Ser Glu His Asp Trp Pro         355 360 365 Ile His Asn Asn Phe Asp Ile Asp Glu Val Gln Leu Asp Pro Arg Ala     370 375 380 Leu Ser Asp Val Thr Asp Glu Glu Glu Val Gln Val Asp Pro Arg Lys 385 390 395 400 Tyr Leu Asp Gly Asp Arg Glu Lys Tyr Leu Glu Asp Pro Ala Phe Asp                 405 410 415 Thr Ser Tyr Ser Ala Glu Pro Cys Trp Gln Tyr Pro Asp His His Glu             420 425 430 Asn Lys Tyr Cys Asp Leu Glu Cys Ser His Thr Cys Asn Tyr Lys Thr         435 440 445 Arg Ser Ser Pro Tyr Leu Asp Asn Leu Val Trp Arg Glu Ser Glu Val     450 455 460 Asn His Tyr Tyr Glu Pro Lys Leu Ile Ile Asp Leu Ser Asn Trp Lys 465 470 475 480 Glu Gln Ser Lys Glu Lys Ser Asp Lys Arg Gly Lys Ser Lys Cys Glu                 485 490 495 Arg Asn Gly Leu Val Lys Ala Gln Ile Ala Leu Glu Glu Ala Ser Gln             500 505 510 Gln Leu Ala Glu Arg Glu Arg Gly Gln Gly Phe Asp Phe Asp Ser Phe         515 520 525 Ile Ala Gly Thr Ile Gln Leu Ser Ala Gln His Gln Ser Ala Asp Val     530 535 540 Val Asp Lys Leu Asn Asp Leu Asn Ser Ser Val Ser Gln Leu Glu Leu 545 550 555 560 Lys Ser Leu Ile Ser Lys Ser Val Ser Arg Glu Lys Gln Glu Lys Gly                 565 570 575 Arg Ala Asn Leu Ala Gln Leu Gly Ala Leu Tyr Gln Ser Ser Trp Asp             580 585 590 Ser Gln Phe Val Ser Gly Gly Glu Glu Cys Phe Leu Ile Ser Gln Phe         595 600 605 Cys Cys Glu Val Arg Lys Asp Glu His Ala Glu Lys Glu Asn Thr Tyr     610 615 620 Thr Ser Tyr Leu Asp Lys Phe Phe Ser Arg Lys Glu Asp Ser Glu Met 625 630 635 640 Leu Glu Thr Glu Pro Val Glu Glu Gly Lys Arg Gly Glu Arg Gly Arg                 645 650 655 Glu Ala Gly Leu Leu Ser Gly Gly Gly Glu Phe Leu Leu Ser Lys Gln             660 665 670 Leu Glu Ser Ile Gly Thr Pro Gln Phe His Ser Pro Val Gly Ser Pro         675 680 685 Leu Lys Ser Ile Gln Ala Thr Leu Thr Pro Ser Ala Met Lys Ser Ser     690 695 700 Pro Gln Ile Pro His Lys Thr Tyr Ser Ser Le Leu Lys His Leu Asn 705 710 715 720 <210> 36 <211> 720 <212> PRT <213> Mus musculus <400> 36 Met Ala Glu Lys Phe Glu Ser Leu Met Asn Ile His Gly Phe Asp Leu   1 5 10 15 Gly Ser Arg Tyr Met Asp Leu Lys Pro Leu Gly Cys Gly Gly Asn Gly              20 25 30 Leu Val Phe Ser Ala Val Asp Asn Asp Cys Asp Lys Arg Val Ala Ile          35 40 45 Lys Lys Ile Val Leu Thr Asp Pro Gln Ser Val Lys His Ala Leu Arg      50 55 60 Glu Ile Lys Ile Ile Arg Arg Leu Asp His Asp Asn Ile Val Lys Val  65 70 75 80 Phe Glu Ile Leu Gly Pro Ser Gly Ser Gln Leu Thr Asp Asp Val Gly                  85 90 95 Ser Leu Thr Glu Leu Asn Ser Val Tyr Ile Val Gln Glu Tyr Met Glu             100 105 110 Thr Asp Leu Ala Asn Val Leu Glu Gln Gly Pro Leu Leu Glu Glu His         115 120 125 Ala Arg Leu Phe Met Tyr Gln Leu Leu Arg Gly Leu Lys Tyr Ile His     130 135 140 Ser Ala Asn Val Leu His Arg Asp Leu Lys Pro Ala Asn Leu Phe Ile 145 150 155 160 Asn Thr Glu Asp Leu Val Leu Lys Ile Gly Asp Phe Gly Leu Ala Arg                 165 170 175 Ile Met Asp Pro His Tyr Ser His Lys Gly His Leu Ser Glu Gly Leu             180 185 190 Val Thr Lys Trp Tyr Arg Ser Pro Arg Leu Leu Leu Ser Pro Asn Asn         195 200 205 Tyr Thr Lys Ala Ile Asp Met Trp Ala Ala Gly Cys Ile Phe Ala Glu     210 215 220 Met Leu Thr Gly Lys Thr Leu Phe Ala Gly Ala His Glu Leu Glu Gln 225 230 235 240 Met Gln Leu Ile Leu Asp Ser Ile Pro Val Val His Glu Glu Asp Arg                 245 250 255 Gln Glu Leu Leu Ser Val Ile Pro Val Tyr Ile Arg Asn Asp Met Thr             260 265 270 Glu Pro His Arg Pro Leu Thr Gln Leu Leu Pro Gly Ile Ser Arg Glu         275 280 285 Ala Leu Asp Phe Leu Glu Gln Ile Leu Thr Phe Ser Pro Met Asp Arg     290 295 300 Leu Thr Ala Glu Glu Ala Leu Ser His Pro Tyr Met Ser Ile Tyr Ser 305 310 315 320 Phe Pro Thr Asp Glu Pro Ile Ser Ser His Pro Phe His Ile Glu Asp                 325 330 335 Glu Val Asp Asp Ile Leu Leu Met Asp Glu Thr His Ser His Ile Tyr             340 345 350 Asn Trp Glu Arg Tyr His Asp Cys Gln Phe Ser Glu His Asp Trp Pro         355 360 365 Ile His Asn Asn Phe Asp Ile Asp Glu Val Gln Leu Asp Pro Arg Ala     370 375 380 Leu Ser Asp Val Thr Asp Glu Glu Glu Val Gln Val Asp Pro Arg Lys 385 390 395 400 Tyr Leu Asp Gly Asp Arg Glu Lys Tyr Leu Glu Asp Pro Ala Phe Asp                 405 410 415 Thr Ser Tyr Ser Ala Glu Pro Cys Trp Gln Tyr Pro Asp His His Glu             420 425 430 Asn Lys Tyr Cys Asp Leu Glu Cys Ser His Thr Cys Asn Tyr Lys Thr         435 440 445 Arg Ser Ser Pro Tyr Leu Asp Asn Leu Val Trp Arg Glu Ser Glu Val     450 455 460 Asn His Tyr Tyr Glu Pro Lys Leu Ile Ile Asp Leu Ser Asn Trp Lys 465 470 475 480 Glu Gln Ser Lys Glu Lys Ser Asp Lys Arg Gly Lys Ser Lys Cys Glu                 485 490 495 Arg Asn Gly Leu Val Lys Ala Gln Ile Ala Leu Glu Glu Ala Ser Gln             500 505 510 Gln Leu Ala Glu Arg Glu Arg Gly Gln Gly Phe Asp Phe Asp Ser Phe         515 520 525 Ile Ala Gly Thr Ile Gln Leu Ser Ala Gln His Gln Ser Ala Asp Val     530 535 540 Val Asp Lys Leu Asn Asp Leu Asn Ser Ser Val Ser Gln Leu Glu Leu 545 550 555 560 Lys Ser Leu Ile Ser Lys Ser Val Ser Arg Glu Lys Gln Glu Lys Gly                 565 570 575 Arg Ala Asn Leu Ala Gln Leu Gly Ala Leu Tyr Gln Ser Ser Trp Asp             580 585 590 Ser Gln Phe Val Ser Gly Gly Glu Glu Cys Phe Leu Ile Ser Gln Phe         595 600 605 Cys Cys Glu Val Arg Lys Asp Glu His Ala Glu Lys Glu Asn Thr Tyr     610 615 620 Thr Ser Tyr Leu Asp Lys Phe Phe Ser Arg Lys Glu Asp Ser Glu Met 625 630 635 640 Leu Glu Thr Glu Pro Val Glu Glu Gly Lys Arg Gly Glu Arg Gly Arg                 645 650 655 Glu Ala Gly Leu Leu Ser Gly Gly Gly Glu Phe Leu Leu Ser Lys Gln             660 665 670 Leu Glu Ser Ile Gly Thr Pro Gln Phe His Ser Pro Val Gly Ser Pro         675 680 685 Leu Lys Ser Ile Gln Ala Thr Leu Thr Pro Ser Ala Met Lys Ser Ser     690 695 700 Pro Gln Ile Pro His Lys Thr Tyr Ser Ser Le Leu Lys His Leu Asn 705 710 715 720 <210> 37 <211> 367 <212> PRT <213> Mus musculus <400> 37 Met Ser Ser Pro Pro Ala Arg Lys Gly Phe Tyr Arg Gln Glu Val   1 5 10 15 Thr Lys Thr Ala Trp Glu Val Arg Ala Val Tyr Gln Asp Leu Gln Pro              20 25 30 Val Gly Ser Gly Ala Tyr Gly Ala Val Cys Ser Ala Val Asp Ser Arg          35 40 45 Thr Gly Asn Lys Val Ala Ile Lys Lys Leu Tyr Arg Pro Phe Gln Ser      50 55 60 Glu Leu Phe Ala Lys Arg Ala Tyr Arg Glu Leu Arg Leu Leu Lys His  65 70 75 80 Met Arg His Glu Asn Val Ile Gly Leu Leu Asp Val Phe Thr Pro Asp                  85 90 95 Glu Ser Leu Asp Asp Phe Thr Asp Phe Tyr Leu Val Met Pro Phe Met             100 105 110 Gly Thr Asp Leu Gly Lys Leu Met Lys His Glu Thr Leu Ser Glu Asp         115 120 125 Arg Ile Gln Phe Leu Val Tyr Gln Met Leu Lys Gly Leu Lys Tyr Ile     130 135 140 His Ala Gly Val Ile His Arg Asp Leu Lys Pro Gly Asn Leu Ala 145 150 155 160 Val Asn Glu Asp Cys Glu Leu Lys Ile Leu Asp Phe Gly Leu Ala Arg                 165 170 175 Gln Ala Asp Ser Glu Met Thr Gly Tyr Val Val Thr Arg Trp Tyr Arg             180 185 190 Ala Pro Glu Val Ile Leu Asn Trp Met Arg Tyr Thr Gln Thr Val Asp         195 200 205 Ile Trp Ser Val Gly Cys Ile Met Ala Glu Met Ile Thr Gly Lys Ile     210 215 220 Leu Phe Lys Gly Asn Asp His Leu Asp Gln Leu Lys Glu Ile Met Lys 225 230 235 240 Ile Thr Gly Thr Pro Pro Pro Glu Phe Val Gln Lys Leu Gln Ser Ala                 245 250 255 Glu Ala Lys Asn Tyr Met Glu Gly Leu Pro Glu Leu Glu Lys Lys Asp             260 265 270 Phe Ala Ser Val Leu Thr Asn Ala Ser Pro Gln Ala Val Asn Leu Leu         275 280 285 Glu Arg Met Leu Val Leu Asp Ala Glu Gln Arg Val Thr Ala Ala Glu     290 295 300 Ala Leu Thr His Pro Tyr Phe Glu Ser Leu Arg Asp Thr Glu Asp Glu 305 310 315 320 Pro Lys Ala Gln Lys Tyr Asp Asp Ser Phe Asp Asp Val Asp Arg Thr                 325 330 335 Leu Glu Glu Trp Lys Arg Val Thr Tyr Lys Glu Val Leu Ser Phe Lys             340 345 350 Pro Pro Arg Gln Leu Gly Ala Arg Val Pro Lys Glu Thr Ala Leu         355 360 365 <210> 38 <211> 366 <212> PRT <213> Mus musculus <400> 38 Met Ser Leu Thr Arg Lys Arg Gly Phe Tyr Lys Gln Asp Ile Asn Lys   1 5 10 15 Thr Ala Trp Glu Leu Pro Lys Thr Tyr Leu Ala Pro Ala His Val Gly              20 25 30 Ser Gly Ala Tyr Gly Ala Val Cys Ser Ala Ile Asp Lys Arg Thr Gly          35 40 45 Glu Lys Val Ala Ile Lys Lys Leu Ser Arg Pro Phe Gln Ser Glu Ile      50 55 60 Phe Ala Lys Arg Ala Tyr Arg Glu Leu Leu Leu Leu Lys His Met His  65 70 75 80 His Glu Asn Val Ile Gly Leu Leu Asp Val Phe Thr Pro Ala Ser Ser                  85 90 95 Leu Arg Ser Phe His Asp Phe Tyr Leu Val Met Pro Phe Met Gln Thr             100 105 110 Asp Leu Gln Lys Ile Met Gly Met Glu Phe Ser Glu Asp Lys Val Gln         115 120 125 Tyr Leu Val Tyr Gln Met Leu Lys Gly Leu Lys Tyr Ile His Ser Ala     130 135 140 Gly Ile Val His Arg Asp Leu Lys Pro Gly Asn Leu Ala Val Asn Glu 145 150 155 160 Asp Cys Glu Leu Lys Ile Leu Asp Phe Gly Leu Ala Arg His Thr Asp                 165 170 175 Thr Glu Met Thr Gly Tyr Val Val Thr Arg Trp Tyr Arg Ala Pro Glu             180 185 190 Val Ile Leu Ser Trp Met His Tyr Asn Gln Thr Val Asp Ile Trp Ser         195 200 205 Val Gly Cys Ile Met Ala Glu Met Leu Thr Gly Lys Thr Leu Phe Lys     210 215 220 Gly Lys Asp Tyr Leu Asp Gln Leu Thr Gln Ile Leu Lys Val Thr Gly 225 230 235 240 Val Pro Gly Ala Glu Phe Val Gln Lys Leu Lys Asp Lys Ala Ala Lys                 245 250 255 Ser Tyr Ile Gln Ser Leu Pro Gln Ser Pro Lys Lys Asp Phe Thr Gln             260 265 270 Leu Phe Pro Arg Ala Ser Pro Gln Ala Ala Asp Leu Leu Asp Lys Met         275 280 285 Leu Glu Leu Asp Val Asp Lys Arg Leu Thr Ala Ala Gln Ala Leu Ala     290 295 300 His Pro Phe Phe Glu Pro Phe Arg Asp Pro Glu Glu Glu Thr Glu Ala 305 310 315 320 Gln Gln Pro Phe Asp Asp Ala Leu Glu His Glu Lys Leu Ser Val Asp                 325 330 335 Glu Trp Lys Gln His Ile Tyr Lys Glu Ile Ser Asn Phe Ser Pro Ile             340 345 350 Ala Arg Lys Asp Ser Arg Arg Arg Ser Gly Met Lys Leu Gln         355 360 365 <210> 39 <211> 364 <212> PRT <213> Mus musculus <400> 39 Met Ser Gly Pro Arg Ala Gly Phe Tyr Arg Gln Glu Leu Asn Lys Thr   1 5 10 15 Val Trp Glu Val Pro Gln Arg Leu Gln Gly Leu Arg Pro Val Gly Ser              20 25 30 Gly Ala Tyr Gly Ser Val Cys Ser Ala Tyr Asp Ala Arg Leu Arg Gln          35 40 45 Lys Val Ala Val Lys Lys Leu Ser Arg Pro Phe Gln Ser Leu Ile His      50 55 60 Ala Arg Arg Thr Tyr Arg Glu Leu Arg Leu Leu Lys His Leu Lys His  65 70 75 80 Glu Asn Val Ile Gly Leu Leu Asp Val Phe Thr Pro Ala Thr Ser Ile                  85 90 95 Glu Asp Phe Ser Glu Val Tyr Leu Val Thr Thr Leu Met Gly Ala Asp             100 105 110 Leu Asn Asn Ile Val Lys Cys Gln Ala Leu Ser Asp Glu His Val Gln         115 120 125 Phe Leu Val Tyr Gln Leu Leu Arg Gly Leu Lys Tyr Ile His Ser Ala     130 135 140 Gly Ile Ile His Arg Asp Leu Lys Pro Ser Asn Val Ala Val Asn Glu 145 150 155 160 Asp Cys Glu Leu Arg Ile Leu Asp Phe Gly Leu Ala Arg Gln Ala Asp                 165 170 175 Glu Glu Met Thr Gly Tyr Val Ala Thr Arg Trp Tyr Arg Ala Pro Glu             180 185 190 Ile Met Leu Asn Trp Met His Tyr Asn Gln Thr Val Asp Ile Trp Ser         195 200 205 Val Gly Cys Ile Met Ala Glu Leu Leu Gln Gly Lys Ala Leu Phe Pro     210 215 220 Gly Asn Asp Tyr Ile Asp Gln Leu Lys Arg Ile Met Glu Val Val Gly 225 230 235 240 Thr Pro Ser Pro Glu Val Leu Ala Lys Ile Ser Ser Glu His Ala Arg                 245 250 255 Thr Tyr Ile Gln Ser Leu Pro Pro Met Pro Gln Lys Asp Leu Ser Ser             260 265 270 Val Phe His Gly Ala Asn Pro Leu Ala Ile Asp Leu Leu Gly Arg Met         275 280 285 Leu Val Leu Asp Ser Asp Gln Arg Val Ser Ala Ala Glu Ala Leu Ala     290 295 300 His Ala Tyr Phe Ser Gln Tyr His Asp Pro Asp Asp Glu Pro Glu Ala 305 310 315 320 Glu Pro Tyr Asp Glu Ser Val Glu Ala Lys Glu Arg Thr Leu Glu Glu                 325 330 335 Trp Lys Glu Leu Thr Tyr Gln Glu Val Leu Ser Phe Lys Pro Leu Glu             340 345 350 Pro Ser Gln Leu Pro Gly Thr His Glu Ile Glu Gln         355 360 <210> 40 <211> 360 <212> PRT <213> Homo sapiens <400> 40 Met Ser Gln Glu Arg Pro Thr Phe Tyr Arg Gln Glu Leu Asn Lys Thr   1 5 10 15 Ile Trp Glu Val Pro Glu Arg Tyr Gln Asn Leu Ser Pro Val Gly Ser              20 25 30 Gly Ala Tyr Gly Ser Val Cys Ala Ala Phe Asp Thr Lys Thr Gly Leu          35 40 45 Arg Val Ala Val Lys Lys Leu Ser Arg Pro Phe Gln Ser Ile Ile His      50 55 60 Ala Lys Arg Thr Tyr Arg Glu Leu Arg Leu Leu Lys His Met Lys His  65 70 75 80 Glu Asn Val Ile Gly Leu Leu Asp Val Phe Thr Pro Ala Arg Ser Leu                  85 90 95 Glu Glu Phe Asn Asp Val Tyr Leu Val Thr His Leu Met Gly Ala Asp             100 105 110 Leu Asn Asn Ile Val Lys Cys Gln Lys Leu Thr Asp Asp His Val Gln         115 120 125 Phe Leu Ile Tyr Gln Ile Leu Arg Gly Leu Lys Tyr Ile His Ser Ala     130 135 140 Asp Ile Ile His Arg Asp Leu Lys Pro Ser Asn Leu Ala Val Asn Glu 145 150 155 160 Asp Cys Glu Leu Lys Ile Leu Asp Phe Gly Leu Ala Arg His Thr Asp                 165 170 175 Asp Glu Met Thr Gly Tyr Val Ala Thr Arg Trp Tyr Arg Ala Pro Glu             180 185 190 Ile Met Leu Asn Trp Met His Tyr Asn Gln Thr Val Asp Ile Trp Ser         195 200 205 Val Gly Cys Ile Met Ala Glu Leu Leu Thr Gly Arg Thr Leu Phe Pro     210 215 220 Gly Thr Asp His Ile Asn Gln Leu Gln Gln Ile Met Arg Leu Thr Gly 225 230 235 240 Thr Pro Pro Ala Tyr Leu Ile Asn Arg Met Pro Ser Ser Glu Ala Arg                 245 250 255 Asn Tyr Ile Gln Ser Leu Thr Gln Met Pro Lys Met Asn Phe Ala Asn             260 265 270 Val Phe Ile Gly Ala Asn Pro Leu Ala Val Asp Leu Leu Glu Lys Met         275 280 285 Leu Val Leu Asp Ser Asp Lys Arg Ile Thr Ala Ala Gln Ala Leu Ala     290 295 300 His Ala Tyr Phe Ala Gln Tyr His Asp Pro Asp Asp Glu Pro Val Ala 305 310 315 320 Asp Pro Tyr Asp Gln Ser Phe Glu Ser Arg Asp Leu Leu Ile Asp Glu                 325 330 335 Trp Lys Ser Leu Thr Tyr Asp Glu Val Ile Ser Phe Val Pro Pro             340 345 350 Leu Asp Gln Glu Glu Met Glu Ser         355 360 <210> 41 <211> 360 <212> PRT <213> Mus musculus <400> 41 Met Ser Gln Glu Arg Pro Thr Phe Tyr Arg Gln Glu Leu Asn Lys Thr   1 5 10 15 Ile Trp Glu Val Pro Glu Arg Tyr Gln Asn Leu Ser Pro Val Gly Ser              20 25 30 Gly Ala Tyr Gly Ser Val Cys Ala Ala Phe Asp Thr Lys Thr Gly His          35 40 45 Arg Val Ala Val Lys Lys Leu Ser Arg Pro Phe Gln Ser Ile Ile His      50 55 60 Ala Lys Arg Thr Tyr Arg Glu Leu Arg Leu Leu Lys His Met Lys His  65 70 75 80 Glu Asn Val Ile Gly Leu Leu Asp Val Phe Thr Pro Ala Arg Ser Leu                  85 90 95 Glu Glu Phe Asn Asp Val Tyr Leu Val Thr His Leu Met Gly Ala Asp             100 105 110 Leu Asn Asn Ile Val Lys Cys Gln Lys Leu Thr Asp Asp His Val Gln         115 120 125 Phe Leu Ile Tyr Gln Ile Leu Arg Gly Leu Lys Tyr Ile His Ser Ala     130 135 140 Asp Ile Ile His Arg Asp Leu Lys Pro Ser Asn Leu Ala Val Asn Glu 145 150 155 160 Asp Cys Glu Leu Lys Ile Leu Asp Phe Gly Leu Ala Arg His Thr Asp                 165 170 175 Asp Glu Met Thr Gly Tyr Val Ala Thr Arg Trp Tyr Arg Ala Pro Glu             180 185 190 Ile Met Leu Asn Trp Met His Tyr Asn Gln Thr Val Asp Ile Trp Ser         195 200 205 Val Gly Cys Ile Met Ala Glu Leu Leu Thr Gly Arg Thr Leu Phe Pro     210 215 220 Gly Thr Asp His Ile Asp Gln Leu Lys Leu Ile Leu Arg Leu Val Gly 225 230 235 240 Thr Pro Gly Ala Glu Leu Leu Lys Lys Ile Ser Ser Glu Ser Ala Arg                 245 250 255 Asn Tyr Ile Gln Ser Leu Ala Gln Met Pro Lys Met Asn Phe Ala Asn             260 265 270 Val Phe Ile Gly Ala Asn Pro Leu Ala Val Asp Leu Leu Glu Lys Met         275 280 285 Leu Val Leu Asp Ser Asp Lys Arg Ile Thr Ala Ala Gln Ala Leu Ala     290 295 300 His Ala Tyr Phe Ala Gln Tyr His Asp Pro Asp Asp Glu Pro Val Ala 305 310 315 320 Asp Pro Tyr Asp Gln Ser Phe Glu Ser Arg Asp Leu Leu Ile Asp Glu                 325 330 335 Trp Lys Ser Leu Thr Tyr Asp Glu Val Ile Ser Phe Val Pro Pro             340 345 350 Leu Asp Gln Glu Glu Met Glu Ser         355 360 <210> 42 <211> 352 <212> PRT <213> Homo sapiens <400> 42 Met Asp Tyr Gln Val Ser Ser Pro Ile Tyr Asp Ile Asn Tyr Tyr Thr   1 5 10 15 Ser Glu Pro Cys Gln Lys Ile Asn Val Lys Gln Ile Ala Ala Arg Leu              20 25 30 Leu Pro Pro Leu Tyr Ser Leu Val Phe Ile Phe Gly Phe Val Gly Asn          35 40 45 Met Leu Val Ile Leu Ile Leu Ile Asn Cys Lys Arg Leu Lys Ser Met      50 55 60 Thr Asp Ile Tyr Leu Leu Aspen Leu Aspen Leu Phe Phe Leu  65 70 75 80 Leu Thr Val Pro Phe Trp Ala His Tyr Ala Ala Gln Trp Asp Phe                  85 90 95 Gly Asn Thr Met Cys Gln Leu Leu Thr Gly Leu Tyr Phe Ile Gly Phe             100 105 110 Phe Ser Gly Ile Phe Phe Ile Ile Leu Leu Thr Ile Asp Arg Tyr Leu         115 120 125 Ala Val Valp His Ala Val Phe Ala Leu Lys Ala Arg Thr Val Thr Phe     130 135 140 Gly Val Val Thr Ser Val Ile Thr Trp Val Val Ala Val Phe Ala Ser 145 150 155 160 Leu Pro Gly Ile Ile Phe Thr Arg Ser Gln Lys Glu Gly Leu His Tyr                 165 170 175 Thr Cys Ser Ser His Phe Pro Tyr Ser Gln Tyr Gln Phe Trp Lys Asn             180 185 190 Phe Gln Thr Leu Lys Ile Val Ile Leu Gly Leu Val Leu Pro Leu Leu         195 200 205 Val Met Val Ile Cys Tyr Ser Gly Ile Leu Lys Thr Leu Leu Arg Cys     210 215 220 Arg Asn Glu Lys Lys Arg His Arg Ala Val Arg Leu Ile Phe Thr Ile 225 230 235 240 Met Ile Val Tyr Phe Leu Phe Trp Ala Pro Tyr Asn Ile Val Leu Leu                 245 250 255 Leu Asn Thr Phe Gln Glu Phe Phe Gly Leu Asn Asn Cys Ser Ser Ser             260 265 270 Asn Arg Leu Asp Gln Ala Met Gln Val Thr Glu Thr Leu Gly Met Thr         275 280 285 His Cys Cys Ile Asn Pro Ile Ile Tyr Ala Phe Val Gly Glu Lys Phe     290 295 300 Arg Asn Tyr Leu Leu Val Phe Phe Gln Lys His Ile Ala Lys Arg Phe 305 310 315 320 Cys Lys Cys Cys Ser Ile Phe Gln Gln Glu Ala Pro Glu Arg Ala Ser                 325 330 335 Ser Val Tyr Thr Arg Ser Thr Gly Glu Gln Glu Ile Ser Val Gly Leu             340 345 350 <210> 43 <211> 354 <212> PRT <213> Mus musculus <400> 43 Met Asp Phe Gln Gly Ser Val Pro Thr Tyr Ser Tyr Asp Ile Asp Tyr   1 5 10 15 Gly Met Ser Ala Pro Cys Gln Lys Ile Asn Val Lys Gln Ile Ala Ala              20 25 30 Gln Leu Leu Pro Pro Leu Tyr Ser Leu Val Phe Ile Phe Gly Phe Val          35 40 45 Gly Asn Met Met Val Phe Leu Ile Leu Ile Ser Cys Lys Lys Leu Lys      50 55 60 Ser Val Thr Asp Ile Tyr Leu Leu Asn Leu Ala Ile Ser Asp Leu Leu  65 70 75 80 Phe Leu Leu Thr Leu Pro Phe Trp Ala His Tyr Ala Ala Asn Glu Trp                  85 90 95 Val Phe Gly Asn Ile Met Cys Lys Val Phe Thr Gly Leu Tyr His Ile             100 105 110 Gly Tyr Phe Gly Gly Ile Phe Phe Ile Ile Leu Leu Thr Ile Asp Arg         115 120 125 Tyr Leu Ala Ile Val Ala Val Phe Ala Leu Lys Val Arg Thr Val     130 135 140 Asn Phe Gly Val Ile Thr Ser Val Val Thr Trp Ala Val Ala Val Phe 145 150 155 160 Ala Ser Leu Pro Glu Ile Ile Phe Thr Arg Ser Gln Lys Glu Gly Phe                 165 170 175 His Tyr Thr Cys Ser Pro His Phe Pro His Thr Gln Tyr His Phe Trp             180 185 190 Lys Ser Phe Gln Thr Leu Lys Met Val Ile Leu Ser Leu Ile Leu Pro         195 200 205 Leu Leu Val Met Val Ile Cys Tyr Ser Gly Ile Leu His Thr Leu Phe     210 215 220 Arg Cys Arg Asn Glu Lys Lys Arg His Arg Ala Val Arg Leu Ile Phe 225 230 235 240 Ala Ile Met Ile Val Tyr Phe Leu Phe Trp Thr Pro Tyr Asn Ile Val                 245 250 255 Leu Leu Leu Thr Thr Phe Gln Glu Phe Phe Gly Leu Asn Asn Cys Ser             260 265 270 Ser Ser Asn Arg Leu Asp Gln Ala Met Gln Ala Thr Glu Thr Leu Gly         275 280 285 Met Thr His Cys Cys Leu Asn Pro Val Ile Tyr Ala Phe Val Gly Glu     290 295 300 Lys Phe Arg Ser Tyr Leu Ser Val Phe Phe Arg Lys His Met Val Lys 305 310 315 320 Arg Phe Cys Lys Arg Cys Ser Ile Phe Gln Gln Asp Asn Pro Asp Arg                 325 330 335 Ala Ser Ser Val Tyr Thr Arg Ser Thr Gly Glu His Glu Val Ser Thr             340 345 350 Gly Leu         <210> 44 <211> 136 <212> PRT <213> Homo sapiens <400> 44 Met Cys Glu Ser Ser Lys Glu Ala Leu Ala Glu Asn Asn Leu Asn Leu   1 5 10 15 Pro Lys Met Ala Glu Lys Asp Gly Cys Phe Gln Ser Gly Phe Asn Glu              20 25 30 Glu Thr Cys Leu Val Lys Ile Ile Thr Gly Leu Leu Glu Phe Glu Val          35 40 45 Tyr Leu Glu Tyr Leu Gln Asn Arg Phe Glu Ser Ser Glu Glu Gln Ala      50 55 60 Arg Ala Val Gln Met Ser Thr Lys Val Leu Ile Gln Phe Leu Gln Lys  65 70 75 80 Lys Ala Lys Asn Leu Asp Ala Ile Thr Thr Pro Asp Pro Thr Thr Asn                  85 90 95 Ala Ser Leu Leu Thr Lys Leu Gln Ala Gln Asn Gln Trp Leu Gln Asp             100 105 110 Met Thr Thr His Leu Ile Leu Arg Ser Phe Lys Glu Phe Leu Gln Ser         115 120 125 Ser Leu Arg Ala Leu Arg Gln Met     130 135 <210> 45 <211> 211 <212> PRT <213> Mus musculus <400> 45 Met Lys Phe Leu Ser Ala Arg Asp Phe His Pro Val Ala Phe Leu Gly   1 5 10 15 Leu Met Leu Val Thr Thr Thr Ala Phe Pro Thr Ser Gln Val Arg Arg              20 25 30 Gly Asp Phe Thr Glu Asp Thr Thr Pro Asn Arg Pro Val Tyr Thr Thr          35 40 45 Ser Gln Val Gly Gly Leu Ile Thr His Val Leu Trp Glu Ile Val Glu      50 55 60 Met Arg Lys Glu Leu Cys Asn Gly Asn Ser Asp Cys Met Asn Asn Asp  65 70 75 80 Asp Ala Leu Ala Glu Asn Asn Leu Lys Leu Pro Glu Ile Gln Arg Asn                  85 90 95 Asp Gly Cys Tyr Gln Thr Gly Tyr Asn Gln Glu Ile Cys Leu Leu Lys             100 105 110 Ile Ser Ser Gly Leu Leu Glu Tyr His Ser Tyr Leu Glu Tyr Met Lys         115 120 125 Asn Asn Leu Lys Asp Asn Lys Lys Asp Lys Ala Arg Val Leu Gln Arg     130 135 140 Asp Thr Glu Thr Leu Ile His Ile Phe Asn Gln Glu Val Lys Asp Leu 145 150 155 160 His Lys Ile Val Leu Pro Thr Pro Ile Ser Asn Ala Leu Leu Thr Asp                 165 170 175 Lys Leu Glu Ser Gln Lys Glu Trp Leu Arg Thr Lys Thr Ile Gln Phe             180 185 190 Ile Leu Lys Ser Leu Glu Glu Phe Leu Lys Val Thr Leu Arg Ser Thr         195 200 205 Arg Gln Thr     210 <210> 46 <211> 269 <212> PRT <213> Mus musculus <400> 46 Met Ala Glu Val Glu Leu Ala Ser Glu Met Met Ala Tyr Tyr Ser   1 5 10 15 Gly Asn Glu Asp Asp Leu Phe Phe Glu Ala Asp Gly Pro Lys Gln Met              20 25 30 Lys Cys Ser Phe Gln Asp Leu Asp Leu Cys Pro Leu Asp Gly Gly Ile          35 40 45 Gln Leu Arg Ile Ser Asp His His Tyr Ser Lys Gly Phe Arg Gln Ala      50 55 60 Ala Ser Val Val Ala Met Asp Lys Leu Arg Lys Met Leu Val Pro  65 70 75 80 Cys Pro Gln Thr Phe Gln Glu Asn Asp Leu Ser Thr Phe Phe Pro Phe                  85 90 95 Ile Phe Glu Glu Glu Pro Ile Phe Phe Asp Thr Trp Asp Asn Glu Ala             100 105 110 Tyr Val His Asp Ala Pro Val Arg Ser Leu Asn Cys Thr Leu Arg Asp         115 120 125 Ser Gln Gln Lys Ser Leu Val Met Ser Gly Pro Tyr Glu Leu Lys Ala     130 135 140 Leu His Leu Gln Gly Gln Asp Met Glu Gln Gln Val Val Phe Ser Met 145 150 155 160 Ser Phe Val Gln Gly Glu Glu Ser Asn Asp Lys Ile Pro Val Ala Leu                 165 170 175 Gly Leu Lys Glu Lys Asn Leu Tyr Leu Ser Cys Val Leu Lys Asp Asp             180 185 190 Lys Pro Thr Leu Gln Leu Glu Ser Val Asp Pro Lys Asn Tyr Pro Lys         195 200 205 Lys Lys Met Glu Lys Arg Phe Val Phe Asn Lys Ile Glu Ile Asn Asn     210 215 220 Lys Leu Glu Phe Glu Ser Ala Gln Phe Pro Asn Trp Tyr Ile Ser Thr 225 230 235 240 Ser Gln Ala Glu Asn Met Pro Val Phe Leu Gly Gly Thr Lys Gly Gly                 245 250 255 Gln Asp Ile Thr Asp Phe Thr Met Gln Phe Val Ser Ser             260 265 <210> 47 <211> 269 <212> PRT <213> Mus musculus <400> 47 Met Ala Thr Val Pro Glu Leu Asn Cys Glu Met Pro Pro Phe Asp Ser   1 5 10 15 Asp Glu Asn Asp Leu Phe Phe Glu Val Asp Gly Pro Gln Lys Met Lys              20 25 30 Gly Cys Phe Gln Thr Phe Asp Leu Gly Cys Pro Asp Glu Ser Ile Gln          35 40 45 Leu Gln Ile Ser Gln Gln His Ile Asn Lys Ser Phe Arg Gln Ala Val      50 55 60 Ser Leu Ile Val Ala Val Glu Lys Leu Trp Gln Leu Pro Val Ser Phe  65 70 75 80 Pro Trp Thr Phe Gln Asp Glu Asp Met Ser Thr Phe Phe Ser Phe Ile                  85 90 95 Phe Glu Glu Glu Pro Ile Leu Cys Asp Ser Trp Asp Asp Asp Asp Asn             100 105 110 Leu Leu Val Cys Asp Val Pro Ile Arg Gln Leu His Tyr Arg Leu Arg         115 120 125 Asp Glu Gln Gln Lys Ser Leu Val Leu Ser Asp Pro Tyr Glu Leu Lys     130 135 140 Ala Leu His Leu Asn Gly Gln Asn Ile Asn Gln Gln Val Ile Phe Ser 145 150 155 160 Met Ser Phe Val Gln Gly Glu Pro Ser Asn Asp Lys Ile Pro Val Ala                 165 170 175 Leu Gly Leu Lys Gly Lys Asn Leu Tyr Leu Ser Cys Val Met Lys Asp             180 185 190 Gly Thr Pro Thr Leu Gln Leu Glu Ser Val Asp Pro Lys Gln Tyr Pro         195 200 205 Lys Lys Lys Met Glu Lys Arg Phe Val Phe Asn Lys Ile Glu Val Lys     210 215 220 Ser Lys Val Glu Phe Glu Ser Ala Glu Phe Pro Asn Trp Tyr Ile Ser 225 230 235 240 Thr Ser Gln Ala Glu His Lys Pro Val Phe Leu Gly Asn Asn Ser Gly                 245 250 255 Gln Asp Ile Ile Asp Phe Thr Met Glu Ser Val Ser Ser             260 265 <210> 48 <211> 1034 <212> PRT <213> Homo sapiens <400> 48 Met Ala Ser Thr Arg Cys Lys Leu Ala Arg Tyr Leu Glu Asp Leu Glu   1 5 10 15 Asp Val Asp Leu Lys Lys Phe Lys Met His Leu Glu Asp Tyr Pro Pro              20 25 30 Gln Lys Gly Cys Ile Pro Leu Pro Arg Gly Gln Thr Glu Lys Ala Asp          35 40 45 His Val Asp Leu Ala Thr Leu Met Ile Asp Phe Asn Gly Glu Glu Lys      50 55 60 Ala Trp Ala Met Ala Val Trp Ile Phe Ala Ala Ile Asn Arg Arg Asp  65 70 75 80 Leu Tyr Glu Lys Ala Lys Arg Asp Glu Pro Lys Trp Gly Ser Asp Asn                  85 90 95 Ala Arg Val Ser Asn Pro Thr Val Ile Cys Gln Glu Asp Ser Ile Glu             100 105 110 Glu Glu Trp Met Gly Leu Leu Glu Tyr Leu Ser Arg Ile Ser Ile Cys         115 120 125 Lys Met Lys Lys Asp Tyr Arg Lys Lys Tyr Arg Lys Tyr Val Arg Ser     130 135 140 Arg Phe Gln Cys Ile Glu Asp Arg Asn Ala Arg Leu Gly Glu Ser Val 145 150 155 160 Ser Leu Asn Lys Arg Tyr Thr Arg Leu Arg Leu Ile Lys Glu His Arg                 165 170 175 Ser Gln Gln Glu Arg Glu Glu Glu Leu Leu Ala Ile Gly Lys Thr Lys             180 185 190 Thr Cys Glu Ser Pro Val Ser Pro Ile Lys Met Glu Leu Leu Phe Asp         195 200 205 Pro Asp Asp Glu His Ser Glu Pro Val His Thr Val Val Phe Gln Gly     210 215 220 Ala Ala Gly Ile Gly Lys Thr Ile Leu Ala Arg Lys Met Met Leu Asp 225 230 235 240 Trp Ala Ser Gly Thr Leu Tyr Gln Asp Arg Phe Asp Tyr Leu Phe Tyr                 245 250 255 Ile His Cys Arg Glu Val Ser Leu Val Thr Gln Arg Ser Leu Gly Asp             260 265 270 Leu Ile Met Ser Cys Cys Pro Asp Pro Asn Pro Pro Ile His Lys Ile         275 280 285 Val Arg Lys Pro Ser Arg Ile Leu Phe Leu Met Asp Gly Phe Asp Glu     290 295 300 Leu Gln Gly Ala Phe Asp Glu His Ile Gly Pro Leu Cys Thr Asp Trp 305 310 315 320 Gln Lys Ala Glu Arg Gly Asp Ile Leu Leu Ser Ser Leu Ile Arg Lys                 325 330 335 Lys Leu Leu Pro Glu Ala Ser Leu Leu Ile Thr Thr Arg Pro Val Ala             340 345 350 Leu Glu Lys Leu Gln His Leu Leu Asp His Pro Arg His Val Glu Ile         355 360 365 Leu Gly Phe Ser Glu Ala Lys Arg Lys Glu Tyr Phe Phe Lys Tyr Phe     370 375 380 Ser Asp Glu Ala Gln Ala Arg Ala Ala Phe Ser Leu Ile Gln Glu Asn 385 390 395 400 Glu Val Leu Phe Thr Met Cys Phe Ile Pro Leu Val Cys Trp Ile Val                 405 410 415 Cys Thr Gly Leu Lys Gln Gln Met Glu Ser Gly Lys Ser Leu Ala Gln             420 425 430 Thr Ser Lys Thr Thr Thr Ala Val Tyr Val Phe Le Le Ser Ser Leu         435 440 445 Leu Gln Pro Arg Gly Gly Ser Gln Glu His Gly Leu Cys Ala His Leu     450 455 460 Trp Gly Leu Cys Ser Leu Ala Ala Asp Gly Ile Trp Asn Gln Lys Ile 465 470 475 480 Leu Phe Glu Glu Ser Asp Leu Arg Asn His Gly Leu Gln Lys Ala Asp                 485 490 495 Val Ser Ala Phe Leu Arg Met Asn Leu Phe Gln Lys Glu Val Asp Cys             500 505 510 Glu Lys Phe Tyr Ser Phe Ile His Met Thr Phe Gln Glu Phe Phe Ala         515 520 525 Ala Met Tyr Tyr Leu Leu Glu Glu Glu Lys Glu Gly Arg Thr Asn Val     530 535 540 Pro Gly Ser Arg Leu Lys Leu Pro Ser Arg Asp Val Thr Val Leu Leu 545 550 555 560 Glu Asn Tyr Gly Lys Phe Glu Lys Gly Tyr Leu Ile Phe Val Val Arg                 565 570 575 Phe Leu Phe Gly Leu Val Asn Gln Glu Arg Thr Ser Tyr Leu Glu Lys             580 585 590 Lys Leu Ser Cys Lys Ile Ser Gln Gln Ile Arg Leu Glu Leu Leu Lys         595 600 605 Trp Ile Glu Val Lys Ala Lys Ala Lys Lys Leu Gln Ile Gln Pro Ser     610 615 620 Gln Leu Glu Leu Phe Tyr Cys Leu Tyr Glu Met Gln Glu Glu Asp Phe 625 630 635 640 Val Gln Arg Ala Met Asp Tyr Phe Pro Lys Ile Glu Ile Asn Leu Ser                 645 650 655 Thr Arg Met Asp His Met Val Ser Ser Phe Cys Ile Glu Asn Cys His             660 665 670 Arg Val Glu Ser Leu Ser Leu Gly Phe Leu His Asn Met Pro Lys Glu         675 680 685 Glu Glu Glu Glu Glu Lys Glu Gly Arg His Leu Asp Met Val Gln Cys     690 695 700 Val Leu Pro Ser Ser Ser His Ala Ala Cys Ser His Gly Leu Val Asn 705 710 715 720 Ser His Leu Thr Ser Ser Phe Cys Arg Gly Leu Phe Ser Ser Leu Ser                 725 730 735 Thr Ser Gln Ser Leu Thr Glu Leu Asp Leu Ser Asp Asn Ser Leu Gly             740 745 750 Asp Pro Gly Met Arg Val Leu Cys Glu Thr Leu Gln His Pro Gly Cys         755 760 765 Asn Ile Arg Arg Leu Trp Leu Gly Arg Cys Gly Leu Ser His Glu Cys     770 775 780 Cys Phe Asp Ile Ser Leu Val Leu Ser Ser Asn Gln Lys Leu Val Glu 785 790 795 800 Leu Asp Leu Ser Asp Asn Ala Leu Gly Asp Phe Gly Ile Arg Leu Leu                 805 810 815 Cys Val Gly Leu Lys His Leu Leu Cys Asn Leu Lys Lys Leu Trp Leu             820 825 830 Val Ser Cys Cys Leu Thr Ser Ala Cys Cys Gln Asp Leu Ala Ser Val         835 840 845 Leu Ser Thr Ser His Ser Leu Thr Arg Leu Tyr Val Gly Glu Asn Ala     850 855 860 Leu Gly Asp Ser Gly Val Ala Leu Cys Glu Lys Ala Lys Asn Pro 865 870 875 880 Gln Cys Asn Leu Gln Lys Leu Gly Leu Val Asn Ser Gly Leu Thr Ser                 885 890 895 Val Cys Cys Ser Ala Leu Ser Ser Val Leu Ser Thr Asn Gln Asn Leu             900 905 910 Thr His Leu Tyr Leu Arg Gly Asn Thr Leu Gly Asp Lys Gly Ile Lys         915 920 925 Leu Leu Cys Glu Gly Leu Leu His Pro Asp Cys Lys Leu Gln Val Leu     930 935 940 Glu Leu Asp Asn Cys Asn Leu Thr Ser His Cys Cys Trp Asp Leu Ser 945 950 955 960 Thr Leu Leu Thr Ser Ser Gln Ser Leu Arg Lys Leu Ser Leu Gly Asn                 965 970 975 Asn Asp Leu Gly Asp Leu Gly Val Met Met Phe Cys Glu Val Leu Lys             980 985 990 Gln Gln Ser Cys Leu Leu Gln Asn Leu Gly Leu Ser Glu Met Tyr Phe         995 1000 1005 Asn Tyr Glu Thr Lys Ser Ala Leu Glu Thr Leu Gln Glu Glu Lys Pro    1010 1015 1020 Glu Leu Thr Val Val Phe Glu Pro Ser Trp 1025 1030 <210> 49 <211> 1033 <212> PRT <213> Mus musculus <400> 49 Met Thr Ser Val Arg Cys Lys Leu Ala Gln Tyr Leu Glu Asp Leu Glu   1 5 10 15 Asp Val Asp Leu Lys Lys Phe Lys Met His Leu Glu Asp Tyr Pro Pro              20 25 30 Glu Lys Gly Cys Ile Pro Val Pro Arg Gly Gln Met Glu Lys Ala Asp          35 40 45 His Leu Asp Leu Ala Thr Leu Met Ile Asp Phe Asn Gly Glu Glu Lys      50 55 60 Ala Trp Ala Met Ala Val Trp Ile Phe Ala Ala Ile Asn Arg Arg Asp  65 70 75 80 Leu Trp Glu Lys Ala Lys Lys Asp Gln Pro Glu Trp Asn Asp Thr Cys                  85 90 95 Thr Ser His Ser Ser Met Val Cys Gln Glu Asp Ser Leu Glu Glu Glu             100 105 110 Trp Met Gly Leu Leu Gly Tyr Leu Ser Arg Ile Ser Ile Cys Lys Lys         115 120 125 Lys Lys Asp Tyr Cys Lys Met Tyr Arg Arg His Val Arg Ser Ser Phe     130 135 140 Tyr Ser Ile Lys Asp Arg Asn Ala Arg Leu Gly Glu Ser Val Asp Leu 145 150 155 160 Asn Ser Arg Tyr Thr Gln Leu Gln Leu Val Lys Glu His Pro Ser Lys                 165 170 175 Gln Glu Arg Glu His Glu Leu Leu Thr Ile Gly Arg Thr Lys Met Arg             180 185 190 Asp Ser Pro Met Ser Ser Leu Lys Leu Glu Leu Leu Phe Glu Pro Glu         195 200 205 Asp Gly His Ser Glu Pro Val His Thr Val Val Phe Gln Gly Ala Ala     210 215 220 Gly Ile Gly Lys Thr Ile Leu Ala Arg Lys Ile Met Leu Asp Trp Ala 225 230 235 240 Leu Gly Lys Leu Phe Lys Asp Lys Phe Asp Tyr Leu Phe Phe Ile His                 245 250 255 Cys Arg Glu Val Ser Leu Arg Thr Pro Arg Ser Leu Ala Asp Leu Ile             260 265 270 Val Ser Cys Trp Pro Asp Pro Asn Pro Pro Val Cys Lys Ile Leu Arg         275 280 285 Lys Pro Ser Arg Ile Leu Phe Leu Met Asp Gly Phe Asp Glu Leu Gln     290 295 300 Gly Ala Phe Asp Glu His Ile Gly Glu Val Cys Thr Asp Trp Gln Lys 305 310 315 320 Ala Val Arg Gly Asp Ile Leu Leu Ser Ser Leu Ile Arg Lys Lys Leu                 325 330 335 Leu Pro Lys Ala Ser Leu Leu Ile Thr Thr Arg Pro Val Ala Leu Glu             340 345 350 Lys Leu Gln His Leu Leu Asp His Pro Arg His Val Glu Ile Leu Gly         355 360 365 Phe Ser Glu Ala Lys Arg Lys Glu Tyr Phe Phe Lys Tyr Phe Ser Asn     370 375 380 Glu Leu Gln Ala Arg Glu Ala Phe Arg Leu Ile Gln Glu Asn Glu Val 385 390 395 400 Leu Phe Thr Met Cys Phe Ile Pro Leu Val Cys Trp Ile Val Cys Thr                 405 410 415 Gly Leu Lys Gln Gln Met Glu Thr Gly Lys Ser Leu Ala Gln Thr Ser             420 425 430 Lys Thr Thr Thr Ala Val Tyr Val Phe Leu Ser Ser Leu Leu Gln         435 440 445 Ser Arg Gly Gly Ile Glu Glu His Leu Phe Ser Asp Tyr Leu Gln Gly     450 455 460 Leu Cys Ser Leu Ala Ala Asp Gly Ile Trp Asn Gln Lys Ile Leu Phe 465 470 475 480 Glu Glu Cys Asp Leu Arg Lys His Gly Leu Gln Lys Thr Asp Val Ser                 485 490 495 Ala Phe Leu Arg Met Asn Val Phe Gln Lys Glu Val Asp Cys Glu Arg             500 505 510 Phe Tyr Ser Phe Ser His Met Thr Phe Gln Glu Phe Phe Ala Ala Met         515 520 525 Tyr Tyr Leu Leu Glu Glu Glu Ala Glu Gly Glu Thr Val Arg Lys Gly     530 535 540 Pro Gly Gly Cys Ser Asp Leu Leu Asn Arg Asp Val Lys Val Leu Leu 545 550 555 560 Glu Asn Tyr Gly Lys Phe Glu Lys Gly Tyr Leu Ile Phe Val Val Arg                 565 570 575 Phe Leu Phe Gly Leu Val Asn Gln Glu Arg Thr Ser Tyr Leu Glu Lys             580 585 590 Lys Leu Ser Cys Lys Ile Ser Gln Gln Val Arg Leu Glu Leu Leu Lys         595 600 605 Trp Ile Glu Val Lys Ala Lys Ala Lys Lys Leu Gln Trp Gln Pro Ser     610 615 620 Gln Leu Glu Leu Phe Tyr Cys Leu Tyr Glu Met Gln Glu Glu Asp Phe 625 630 635 640 Val Gln Ser Ala Met Asp His Phe Pro Lys Ile Glu Ile Asn Leu Ser                 645 650 655 Thr Arg Met Asp His Val Val Ser Ser Phe Cys Ile Lys Asn Cys His             660 665 670 Arg Val Lys Thr Leu Ser Leu Gly Phe Phe His Asn Ser Pro Lys Glu         675 680 685 Glu Glu Glu Arg Arg Gly Gly Arg Pro Leu Asp Gln Val Gln Cys     690 695 700 Val Phe Pro Asp Thr His Val Ala Cys Ser Ser Arg Leu Val Asn Cys 705 710 715 720 Cys Leu Thr Ser Ser Phe Cys Arg Gly Leu Phe Ser Ser Leu Ser Thr                 725 730 735 Asn Arg Ser Leu Thr Glu Leu Asp Leu Ser Asp Asn Thr Leu Gly Asp             740 745 750 Pro Gly Met Arg Val Leu Cys Glu Ala Leu Gln His Pro Gly Cys Asn         755 760 765 Ile Gln Arg Leu Trp Leu Gly Arg Cys Gly Leu Ser His Gln Cys Cys     770 775 780 Phe Asp Ile Ser Ser Val Leu Ser Ser Ser Gln Lys Leu Val Glu Leu 785 790 795 800 Asp Leu Ser Asp Asn Ala Leu Gly Asp Phe Gly Ile Arg Leu Leu Cys                 805 810 815 Val Gly Leu Lys His Leu Leu Cys Asn Leu Gln Lys Leu Trp Leu Val             820 825 830 Ser Cys Cys Leu Thr Ser Ala Cys Cys Gln Asp Leu Ala Leu Val Leu         835 840 845 Ser Ser Asn His Ser Leu Thr Arg Leu Tyr Ile Gly Glu Asn Ala Leu     850 855 860 Gly Asp Ser Gly Val Gln Val Leu Cys Glu Lys Met Lys Asp Pro Gln 865 870 875 880 Cys Asn Leu Gln Lys Leu Gly Leu Val Asn Ser Gly Leu Thr Ser Ile                 885 890 895 Cys Cys Ser Ala Leu Thr Ser Val Leu Lys Thr Asn Gln Asn Phe Thr             900 905 910 His Leu Tyr Leu Arg Ser Asn Ala Leu Gly Asp Thr Gly Leu Arg Leu         915 920 925 Leu Cys Glu Leu Leu His Pro Asp Cys Lys Leu Gln Met Leu Glu     930 935 940 Leu Asp Asn Cys Ser Leu Thr Ser His Ser Cys Trp Asn Leu Ser Thr 945 950 955 960 Ile Leu Thr His Asn His Ser Leu Arg Lys Leu Asn Leu Gly Asn Asn                 965 970 975 Asp Leu Gly Asp Leu Cys Val Val Thr Leu Cys Glu Val Leu Lys Gln             980 985 990 Gln Gly Cys Leu Leu Gln Ser Leu Gln Leu Gly Glu Met Tyr Leu Asn         995 1000 1005 Arg Glu Thr Lys Arg Ala Leu Glu Ala Leu Glu Glu Glu Lys Pro Glu    1010 1015 1020 Leu Thr Ile Val Phe Glu Ile Ser Trp 1025 1030 <210> 50 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Ccr5 primer <400> 50 gtttgcctct ctcccagaaa ta 22 <210> 51 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Ccr5 primer <400> 51 ctgagtagca gatgaccatg ac 22 <210> 52 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> Il-6 primer <400> 52 tggatgctac caaactggat ataa 24 <210> 53 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Il-6 primer <400> 53 aggactctgg ctttgtcttt c 21 <210> 54 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Csf3 primer <400> 54 cagaaagccc tttccagata gt 22 <210> 55 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Csf3 primer <400> 55 agccttctct ctctgctcta a 21 <210> 56 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Il-1beta primer <400> 56 tcattgtggc tgtggagaag 20 <210> 57 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> Il-1beta primer <400> 57 gcctgtagtg cagttgtcta a 21 <210> 58 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Clec4d primer <400> 58 tggagagcct tccagtctaa 20 <210> 59 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Clec4d primer <400> 59 atcagcaagt cccaggaaat aa 22 <210> 60 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> Ctla2b primer <400> 60 gactcatgtg ggaggagaat aag 23 <210> 61 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Ctla2b primer <400> 61 ggcaaatcag gagccatttc 20 <210> 62 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Nlrp3 primer <400> 62 ccgtctacgt cttcttcctt tc 22 <210> 63 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> Nlrp3 primer <400> 63 cgcagatcac actcctcaaa ta 22

Claims (19)

In vitro, incubating the cells with palmitic acid;
Incubating the palmitic acid and incubated cells with minimally-oxidized low density lipoprotein (mMLDL) and lipopolysaccharides (LPS);
Contacting the cell with a test substance;
Measuring the expression level of the Clec4d gene or the activity level of the protein in the cell contacted with the test substance;
Comparing the expression level of the measured gene or the activity level of the protein with the level of the control group; And
And determining the test substance as a candidate for a therapeutic agent for a cardiovascular disease when the expression level of the gene or the activity level of the protein is lower than the level of the control group. delete delete The method according to claim 1, wherein the concentration of palmitic acid is 10 to 1000 μM and the concentration of mMLDL is 5 to 500 μg / ml. The method of claim 1, wherein the concentration of LPS is between 1 and 100 ng / ml. The method according to claim 1, wherein the cell is a monocyte, a vascular endothelial cell, or a macrophage. The method according to claim 1, wherein the expression level or protein activity level of at least one gene selected from the group consisting of CXCL2, TNF-alpha, pERK, pp38, Ccr5, IL-6, IL- Measuring;
Comparing the expression level of the measured gene or the activity level of the protein with the level of the control group; And
And determining the test substance as a candidate for a therapeutic agent for cardiovascular disease when the expression level of the gene or the activity level of the protein is lower than the level of the control group. The method according to claim 1, wherein the cardiovascular disease is at least one selected from the group consisting of atherosclerosis, hypertension, myocardial infarction and angina. delete delete delete delete delete delete delete delete delete delete delete

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