JP2002514045A - Secreted proteins and polynucleotides encoding them - Google Patents

Abstract

(57) Abstract Disclosed are novel polynucleotides and proteins encoded by them.

Description

DETAILED DESCRIPTION OF THE INVENTION             Secreted proteins and polynucleotides encoding them   This application is a part of application Ser. No. 08/667231, filed Jul. 9, 1996. No. 08 / 421,488, filed Sep. 27, 1996, which is a continuation-in-part application. Partial continuation application.                                Field of the invention   The present invention relates to novel polynucleotides and the use of such polynucleotides. Proteins, and therapeutic and diagnostic methods for these polynucleotides and proteins Provides catastrophic and research utility.                                Background of the Invention   Protein factors such as lymphokines, interferons, CSFs and The method aimed at the discovery of cytokines such as Matured rapidly over the years. Nowadays conventional hybridization cloning And expression cloning methods provide information directly related to the discovered protein (ie, In the case of hybridization cloning, the partial DNA / amino acid sequence of the protein Column; the activity of the protein in the case of expression cloning) The new polypeptide is cloned "directly". Signal sequence cloning ( DNA sequence based on the presence of the currently well-recognized secretory leader sequence motif And hybridization by various PCRs or with low stringency More recent "indirect" cloning methods, such as the dicing cloning method Is secreted in the case of cloning of the leader sequence. Or biological activity depending on the cell or tissue source in the case of the PCR method. Access to numerous DNA / amino acid sequences for proteins known to be Has improved the status quo. The present invention provides these proteins and their Mode Which are directed to the polynucleotide.                                Summary of the Invention   In one embodiment, the present invention provides an isolated polypeptide selected from the group consisting of: Provide a composition comprising a renucleotide:   (A) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1;   (B) the nucleotide sequence from nucleotide 477 to nucleotide 752 of SEQ ID NO: 1 A polynucleotide comprising a reotide sequence;   (C) of clone AX568 deposited under accession number ATCC 98101 A polynucleotide comprising the nucleotide sequence of the full-length protein coding sequence;   (D) of clone AX568 deposited under accession number ATCC 98101 Polynucleotide encoding full-length protein encoded by cDNA insert Tide;   (E) of clone AX568 deposited under accession number ATCC 98101 A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence;   (F) of clone AX568 deposited under accession number ATCC 98101; Polynucleotide encoding mature protein encoded by cDNA insert Tide;   (G) a polynucleotide encoding a protein comprising the amino acid sequence of SEQ ID NO: 2 Tide;   (H) including a fragment of the amino acid sequence of SEQ ID NO: 2 having biological activity A polynucleotide encoding a protein;   (I) of the isolate AX5628 deposited under accession number ATCC 98180 A polynucleotide comprising the nucleotide sequence of the full-length protein coding sequence;   (J) of isolate AX5628 deposited under accession number ATCC 98180 Polynucleotide encoding full-length protein encoded by cDNA insert Tide;   (K) of isolate AX5628 deposited under accession number ATCC 98180 A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence;   (L) of isolate AX5628 deposited under accession number ATCC 98180 Polynucleotide encoding mature protein encoded by cDNA insert Tide;   (M) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 5;   (N) the nucleotide sequence from nucleotide 307 to nucleotide 402 of SEQ ID NO: 5 A polynucleotide comprising a reotide sequence;   (O) a polynucleotide encoding a protein comprising the amino acid sequence of SEQ ID NO: 6 Tide;   (P) including a fragment of the amino acid sequence of SEQ ID NO: 6 having biological activity A polynucleotide encoding a protein;   (Q) Alleles of the polynucleotides (a) to (e) or (i) to (n) above A polynucleotide that is a genetic variant;   (R) encodes a species homolog of the protein of (g), (h), (o) or (p). Polynucleotide.   Preferably, such a polynucleotide is nucleotide 477 of SEQ ID NO: 1. Accession number ATCC 98101; Of the full-length protein coding sequence of clone AX568 deposited as Clone AX5 deposited under accession number ATCC 98101; It contains the nucleotide sequence of 68 mature protein coding sequences. Other preferred equipment In an embodiment, the polynucleotide has been deposited under accession number ATCC 98101. Length encoded by the cDNA insert of clone AX568 Encodes a mature protein.   Another preferred embodiment is an isolate deposited under accession number ATCC 98180. Polynucleic acid comprising the nucleotide sequence of the full-length protein coding sequence of AX5628 Reotide; isolate AX56 28 deposited under accession number ATCC 98180 Encoding a full-length protein encoded by a cDNA insert Maturation of the isolate AX5628 deposited under accession number ATCC 98180 A polynucleotide comprising the nucleotide sequence of a protein coding sequence; accession number A CDNA insert of isolate AX5628 deposited as TCC98180 Encoding the mature protein encoded by SEQ ID NO: 5 A polynucleotide comprising a nucleotide sequence; nucleotide 307 of SEQ ID NO: 5 A polynucleotide comprising the nucleotide sequence of SEQ ID NO: 6 A polynucleotide that encodes a protein containing a noic acid sequence; or has biological activity Encoding a protein comprising a fragment of the amino acid sequence of SEQ ID NO: 6 And nucleotides.   In another embodiment, the present invention relates to an amino acid selected from the group consisting of: A set comprising a protein comprising an acid sequence and substantially free of other mammalian proteins Serve the product:   (A) the amino acid sequence of SEQ ID NO: 2;   (B) a fragment of the amino acid sequence of SEQ ID NO: 2; and   (C) of clone AX568 deposited under accession number ATCC 98101 an amino acid sequence encoded by the cDNA insert;   (D) the amino acid sequence of SEQ ID NO: 6;   (E) a fragment of the amino acid sequence of SEQ ID NO: 6; and   (F) of isolate AX5628 deposited under accession number ATCC 98180 Amino acid sequence encoded by the cDNA insert. Preferably, such a protein Comprises the amino acid sequence of SEQ ID NO: 2 or SEQ ID NO: 6.   Another specific example is the cDNA sequence of SEQ ID NO: 1 or SEQ ID NO: 4, SEQ ID NO: 4. 5 or a gene corresponding to the cDNA sequence of SEQ ID NO: 7 is provided.   In certain preferred embodiments, the polynucleotide is operable with an expression control sequence. Linked to Noh. The present invention also provides a method for transforming such a polynucleotide composition. Also provided are host cells, including bacterial, yeast, insect and mammalian cells.   Further, a method for producing a protein,   (A) culturing a host cell culture transformed with such a polynucleotide composition; Grown in the appropriate medium;   (B) purifying the protein from the culture There is also provided a method comprising: The protein produced by such a method is also It is provided by Ming. In a preferred embodiment, such a method is used to produce And the protein to be produced is a mature form of the protein.   The protein composition of the present invention may further contain a pharmaceutically acceptable carrier. Or Compositions comprising antibodies specifically reactive with such proteins are also provided by the present invention.   Administering a therapeutically effective amount of a composition comprising a protein of the invention and a pharmaceutically acceptable carrier. For preventing, treating or ameliorating a medical condition, including administering to an animal subject A law is also provided.                             BRIEF DESCRIPTION OF THE FIGURES   FIG. 1 shows the pED6 and pNotS used to deposit the clones disclosed herein. It is a schematic diagram of a vector.                                Detailed description Isolated proteins and polynucleotides   Nucleotides for each of the clones and proteins disclosed herein And amino acid sequences are reported below. In some cases, the array is Contains some inaccurate or unclear bases or amino acids Maybe. By sequencing the deposited clones by known methods, The actual nucleotide sequence of the protein can be easily determined. Then the estimated net The amino acid sequence (both full length and mature) can be determined from such a nucleotide sequence. Can be. Express the clone in an appropriate host cell, collect the protein, and then Of the protein encoded by the individual clones The acid sequence can also be determined.   For each protein disclosed, Applicants provide information on the sequences available at the time of filing. The one that was determined to be the best identifiable reading frame used was identified. Reported The sequence of the reported protein is It includes the symbol “Xaa”. These "Xaa" codes correspond to (1) the nucleotide sequence A residue that cannot be identified because the sequence was unclear, or (2) (nucleotide sequence Decision that the applicant should not be present) Shows the stop codon in the leotide sequence.   As used herein, the term "secreted" protein refers to a membrane when expressed in a suitable host cell. Refers to those that are transported through, and the result of the signal sequence in that amino acid sequence Transportation is also included. "Secreted" proteins are secreted entirely by the cells in which they are expressed. Proteins (eg, soluble proteins) or partially secreted proteins (eg, receptors) ), But not limited thereto. Also, "secreted" proteins pass through the membrane of the endoplasmic reticulum Including but not limited to those transported byClone “AX568”   The polynucleotide of the present invention was identified as clone "AX568". AX 568 were synthesized using methods selective for the cDNA encoding the secreted protein. It was isolated from a human testis cDNA library. AX568 is a full-length clone. And all the secretory proteins (also referred to herein as "AX568" proteins). Includes marking sequences.   The nucleotide sequence of AX568 determined this time is shown in SEQ ID NO: 1. Current Applicant acknowledges that the correct reading frame and AX568 protein corresponding to the above nucleotide sequence And those considered to be deduced amino acid sequences are shown in SEQ ID NO: 2.   The EcoRI / NotI restriction plasmid obtained from the deposit containing clone AX568. The fragment should be about 730 bp.   The nucleotide sequence disclosed herein for AX568 is provided by BLASTA / BLASTX and And a search against the GenBank database using the FASTA search protocol. Day Nothing hit during the tab. The amino acid sequence of AX568 May have some homology to chicken cytotactin Indicates that   Clone "AX56 28" is an additional isolate of AX568. This time decided The nucleotide sequence of the 5 'portion of AX5628 thus obtained is shown in SEQ ID NO: 4. An additional internal nucleotide sequence for AX5628 is shown in SEQ ID NO: 1. Currently, we distribute what we consider to be the correct reading frame for the coding area. Column number: 2 Estimation of AX5628 protein corresponding to the above nucleotide sequence The amino acid sequence is shown in SEQ ID NO: 2. AX56 28-3, including poly A tail The additional nucleotide sequence from the 'portion is shown in SEQ ID NO: 7.   A search was made for additional sequence information obtained from isolate AX5628. And "yz42a06.s1 Homo sapiens cDNA clone 285682 3 '" (N67571, Fasta) and " e05023t Homo sapiens cDNA clone e05023 5 'end "(T19080, BlastN) At least some identity to the performed EST was shown.   Depositing clones   Clone AX568 was obtained on July 9, 1996 at the American Type Culture Coll. Deposited with Accession No. ATCC 98101, from which the clone was obtained. It is possible. Each clone is deposited in a separate bacterial cell (E. coli) as this complex deposit. Was transfected. Additional isolates of AX568 ("AX568 8 ") on September 26, 1996, under accession number ATCC 98180. Deposited with the American Type Culture Collection. This clone also Available.   EcoRI / NotI digestion (5 'site, EcoRI; 3' site, NotI) And perform an appropriate size fragment (approximately Depositing each clone by obtaining the Can be taken from a vector that has been Each clone was designated pED6 or Deposited in any of the pNotS vectors. In some cases, the deposit Clones can "flip" (ie, reverse) in the deposited isolate There is a potential. Even in such cases, digest with EcoRI and NotI Thus, cDNA insert can be isolated. However, on the spot The correct cDNA arrangement for expression in a suitable vector. NotI will generate a 5 'site and EcoRI will generate a 3' site. The c DNA may be expressed from the deposited vector. O289 1 to Bluescript vector You may put it in a deposit and deposit it. In such cases, XhoI (5 'site) and Not The cDNA insert can be removed from the vector by digestion at I (3 'site). Wear.   Bacterial cells containing a particular clone can be obtained from a composite deposit as follows. it can:   The oligonucleotide probe (s) are known as specific clones Should be designed for This sequence may be any of the sequences herein or their It can be derived from a combination of sequences. Used to isolate each full-length clone The sequence of the inserted oligonucleotide probe is identified as Should be the most reliable in isolating clones.clone Probe sequence AX56 8 SEQ ID NO: 3   In the above sequence containing N at position 2, that position is the preferred probe / primer In one example, one nucleotide (for example, biotin phosphoramidite (1- Dimethoxytrityloxy-2- (N-biotinyl-4-aminobutyl) -propyl 3-O- (2-cyanoethyl)-(N, N-diisopropyl) -phosphorua Madite) (Glen Research, cat.no. 10-1953). ) Rather than biotinylated phosphoramidite residues.   Preferably, the design of the oligonucleotide probe follows these parameters. Should be:   (A) The region of the sequence where the number of unclear bases (N), if any, is minimal Should be designed to:   (B) T_mAbout 80 ° C (2 ° C for A or T, G or C respectively) About 4 ° C.).   Preferably, a method widely used for oligonucleotide labeling is used, The oligonucleotide is g-³²P-ATP (specific activity 6000 Ci / mmol). You can use other labeling methods Wear. Preferably, unincorporated labels are separated by gel filtration chromatography or Should be removed by other established methods. Incorporated in the probe Radioactivity should be quantified by measuring with a scintillation counter. You. Preferably, the specific activity of the probe obtained is about 4e + 6 dpm / picomolar. There should be.   Preferably, a bacterial culture containing a pool of full-length clones is thawed and 100 μl of Stock was added to 25 ml of sterile L-broth containing 100 μg / ml ampicillin Should be used to inoculate sterile culture flasks containing. Preferably, the culture is It should be grown at 37 ° C. to saturation, and preferably, the saturated culture should be fresh L Should be diluted with broth. Preferably, some of these dilutions are plated 100 μg / ml ampicillin in a 150 mm Petri dish and Grow overnight at 37 ° C. on solid bacterial culture medium containing L broth containing 5% agar Dilution yields approximately 5000 distinct and well-separated colonies And volume should be determined. To get clear, well-separated colonies Other known methods can also be used.   The colonies are then nitted using standard colony hybridization techniques. Transfer to a low cellulose filter for dissolution, denaturation and baking. You.   Then, preferably, 0.5% SDS, 100 μg / ml yeast RNA and 6X SSC containing 10 mM EDTA (20X stock is 175.3 g NaC 1 / liter, 88.2 g sodium citrate / liter, pH 7.0 with NaOH. 0) (about 10ml per 150mm filter) Incubate the filter at 65 ° C. for 1 hour. Then preferred Alternatively, the probe is added to the hybridization mixture to give a concentration of 1e + 6dp. m / ml or more. Then gently stir at 65 ° C overnight. Incubation is preferred. The filter is then preferably brought to room temperature. I Wash with 500 ml of 2X SSC / 0.5% SDS without stirring, preferably Subsequently, 500 ml of 2X SSC / 0.1 with gentle shaking at room temperature. Wash with% SDS for 15 minutes. The third wash is performed at 65 ° C. for 30 minutes to 1 hour, 0 hours. . Optimally, it is performed with 1X SSC / 0.5% SDS. Then, preferably The filter is dried and subjected to autoradiography for a sufficient time to remove positive Visualize on the film. Other known hybridization methods may be used. it can.   Positive colonies are picked, grown in culture medium, and then added using standard procedures. Isolate the mid DNA. Then, restriction analysis, hybridization analysis or Clones can be verified by DNA sequencing.   A fragment of the protein of the present invention that can exhibit biological activity is also included in the present invention. It is. Protein fragments may be linear or, for example, H.U.Sarago vi, etal., Bio / Technology 10, 773-778 (1992) and R.S. McDowell, et al., J . Amer. Chem. Soc. 114, 9245-9253 (1992) (both references are incorporated herein by reference). Cyclization using known methods such as those described in Good. For many purposes, including increasing the number of valencies at the protein binding site, Such a fragment may be fused to a carrier molecule such as an immunoglobulin. For example, a fragment of a protein can be linked to the immunoglobulin Fc via a “linker” sequence. It may be fused to parts. For a bivalent form of the protein, such a fusion would be an IgG molecule. For the Fc portion of Using other immunoglobulin isotypes Such fusion may be performed. For example, a 10-valent form of a book A protein of the invention will result.   The invention also provides the full length and mature forms of the disclosed proteins. Full length form of such a protein Is identified in the sequence listing by translation of the nucleotide sequence of the disclosed clone. I have. The mature form of such a protein can be found in a suitable mammalian cell or other host cell. To release the disclosed full-length polynucleotides (preferably those deposited with the ATCC). It may be obtained by exposing. Amino acid sequence of full-length form It may be determined from the column.   The present invention also provides a gene corresponding to the cDNA sequence disclosed herein. Corresponding genes can be isolated by known methods using the sequence information disclosed herein. Or Such methods involve the use of genes in a suitable genomic library or other genomic material source. From the disclosed sequence information for the identification and / or amplification of Preparing a primer.   When the protein of the present invention is membrane-bound (for example, when it is a receptor), the present invention Also provides soluble forms of such proteins. In such a form, the protein Cells in which the protein has been expressed by deleting some or all of its transmembrane domain To be sufficiently secreted from Intracellular and transmembrane domains of the protein of the invention Is identified by a known method for determining such a domain from sequence information. Can be.   Species homologs of the disclosed proteins are also provided by the present invention. Shown here Create an appropriate probe or primer from the sequence and select the appropriate nucleic acid from the desired species Species homologs may be isolated and identified by screening the source.   The invention also relates to allelic variants of the disclosed polynucleotides or proteins. Is included. That is, the present invention provides a protein encoded by the polynucleotide. Isolated polynucleotide encoding the same, homologous or related protein as white And other forms existing in the above.   The isolated polynucleotide of the present invention can be prepared as described in Kaufman et al., Nucleic Acids Res. 19 Expression vector such as the pMT2 or pED expression vector disclosed in The protein may be recombinantly produced by operably linking it to the sequence. Many fit Various expression control sequences are known in the art. General method for expressing recombinant proteins Are also known and are described, for example, in R. Kaufman, Methods in Enzymology 185, 537-566. (1990). "Operably linked," as defined herein, refers to the present invention. The isolated polynucleotide and expression control sequence are placed in a vector or cell, Transformation with the polynucleotide / expression control sequence linked to the protein (transfection Means that it is to be expressed by the transformed host cell.   Many types of cells can serve as suitable host cells for expression of the protein. Mammalian host Cells include, for example, monkey COS cells, Chinese hamster ovary (CHO) cells , Human kidney 293 cells, human epidermal A431 cells, human Colo205 cells, 3T3 Cells, CV-1 cells, other transformed primate cell lines, normal diploid cells, primary Cell lines derived from in vitro culture of tissues, primary explants, HeLa cells, mouse L Cells, BHK, HL-60, U937, HaK or Jurkat cells.   Alternatively, in lower eukaryotic cells such as yeast or prokaryotic cells such as bacteria. It is also possible to produce proteins. A potentially suitable yeast strain is Saccharomyces ce revisiae, Schizosaccharomyces pombe, Kluyveromyces strain, Candida, or Yeast strains capable of expressing a seed protein are included. A potentially suitable bacterial strain is Escheric hia coli, Bacillus subtilis, Salmonella typhimurium, or heterologous protein Includes competent bacterial strains. When the protein is produced in yeast or bacteria The protein produced therein, for example, by phosphorylation at appropriate sites or glycosylation May need to be modified to obtain a functional protein. Known chemical Alternatively, such covalent bonding may be performed using enzymatic methods.   The isolated polynucleotide of the present invention may be contained in one or more insect expression vectors. The protein is obtained by operably linking it to an appropriate control sequence and using an insect expression system. Is also good. Materials and methods for baculovirus / insect cell expression systems are described, for example, in Kit form from Tnvitrogen, San Diego, California, USA (MaxBac^Rkit) And such methods are well known in the art and are available from Summers. And Smith, Texas Agricultural Experiment Station Bulletin No. 1555 (198 7) (such as those described herein by reference) is there. As used herein, a conjugate having the ability to express the polynucleotide of the present invention is used. Insect cells have been "transformed".   Culturing transformed host cells under culture conditions suitable for expressing the recombinant protein The protein of the present invention may be produced more. Next, the obtained expressed protein was subjected to gel filtration and filtration. Such cultures using known purification processes, such as ion exchange chromatography (I.e., culture medium or cell extract). Also, protein Purification is performed using an affinity column containing an agent that will bind to the protein; Canavalin A-agarose, heparin-Toyopearl^ROr Cibacrom blue 3GA Sepharose^ROne or more columns with affinity columns such as D. Trees such as phenyl ether, butyl ether, or propyl ether One or more steps using hydrophobic interaction chromatography with fats Or alternatively, immunoaffinity chromatography.   Alternatively, the protein of the invention may be expressed in an easily purified form. For example, Lactose binding protein (MBP), glutathione-S-tonspherase (GST) Or expressed as a fusion protein such as a fusion protein with thioredoxin (TRX) You may. Kits for expression and purification of such fusion proteins are available from New En from glan BioLab (Beverly, MA), Pharmacia (Piscataway, NJ) and InVitrogen It is commercially available. Tag a protein with an epitope, and then tag such epitope Purification can also be achieved by using directed, specific antibodies. It takes 1 Pitopes ("flags") are commercially available from Kodak (New Haeven, CT).   Finally, a hydrophobic RP-HPLC medium such as a pendant methyl or other aliphatic group is added. One or more reversed-phase high quality liquid chromatography using silica gel with The protein can be further purified using the-(RP-PLC) step. A few Or a substantially homogeneous isolation system using various combinations of all of the above purification steps. A recombinant protein can also be obtained. The protein thus purified can be derived from other mammalian proteins. It is not qualitatively included and is defined by the present invention as an "isolated protein".   The protein of the present invention may be used as a product of a transgenic animal. Characterized by somatic or germ cells containing the nucleotide sequence Expressed as an ingredient in milk of transgenic cows, goats, pigs, or sheep Is also good.   The protein may be produced by known conventional chemical synthesis. The present invention by synthetic means Methods for constructing proteins are known to those skilled in the art. The synthetically constructed protein sequence is primary Sharing secondary or tertiary structure and / or conformational properties Thus, they may have common biological properties, including protein activity. Yo For the screening of therapeutic compounds and the generation of antibodies for the production of antibodies. Process to replace the biological activity or immunological replacement of the naturally occurring purified protein Use May be.   The protein shown in the present specification has an amino acid sequence similar to that of the purified protein. Which are modified naturally or by derivatization Is also good. For example, modification of a peptide or DNA sequence can be accomplished by one of ordinary skill in the art using known methods. More can be done. The desired modification in the protein sequence depends on the choice in the coding sequence. Includes amino acid residue changes, substitutions, exchanges, insertions or deletions. For example, up to one Or more cysteine residues have been deleted or replaced with another amino acid. The conformation of the child may be changed. Such changes, replacements, exchanges, insertions, etc. Methods for deletion or deletion are well known to those skilled in the art (see, for example, US Pat. No. 8584). Preferably, such changes, substitutions, exchanges, insertions or deletions It retains the desired activity of white.   Expected to retain protein activity in whole or in part, thus screening Or other fragments of the protein sequence that may be useful for immunological or other immunological methods. Derivatives can also be made by those skilled in the art from the disclosure herein. Such modifications are the subject of the present invention. Is believed to be encompassed byApplications and biological activities   The polynucleotides and proteins of the present invention may have one or more of the following uses or Exhibiting biological activity (including those associated with the assays cited herein) Conceivable. The uses or activities described for the proteins of the present invention may be attributed to the administration of such proteins. Supply or use, or of a polynucleotide encoding such a protein. Administration or use (eg, for any vector suitable for gene therapy or DNA transfer) C).Research applications and usefulness   The polynucleotide provided by the present invention can be used for various purposes depending on the research population. Can be used. Preferential expression of the corresponding protein for analysis, characterization or therapeutic use (Constitutively, at a specific stage of tissue differentiation or development, or As tissue markers (expressed in disease states) as well as Southern gel molecules As a quantity marker, it can be used to identify chromosomes or map related gene locations. Compared to the patient's endogenous DNA as a chromosome marker or tag (if labeled) Probes to identify potential genetic diseases Genetic fingerprinting to discover new related DNA sequences As a source of information for obtaining PCR primers for Probes to subtract known sequences in the process of nucleotide discovery And select the oligomer to bind to a “gene chip” or other support To test for expression patterns, use DNA immunization to To generate antibodies, as well as to generate anti-DNA antibodies, or Polynucleotides can be used as antigens to induce an immune response You. Binds or potentially binds another protein (e.g., If the protein encodes a protein that binds to Or interaction trap assays to identify inhibitors of binding interactions A (for example, as described in Gyuris et al., Cell 75: 791-803 (1993)). And polynucleotides can be used.   The proteins provided by the present invention are a family of multiple proteins for high-throughput screening. For assays to determine biological activity, including proteins of interest; production of antibodies or other A protein (or its receptor) in a biological fluid Reagents (including labeling reagents) in assays designed to quantitatively determine The corresponding protein is preferentially expressed (constitutively or with tissue differentiation or Is expressed at a particular stage of development or in a disease state) As well as used to, of course, isolate related receptors or ligands You may. Bind when a protein binds or potentially binds to another protein Proteins to identify other proteins and / or inhibitors of binding interactions White can be used. Using proteins involved in these binding interactions, Of peptide or small molecular inhibitors or agonists for binding interactions Training can also be performed.   Any or all of these research uses may be commercialized as research products. Can be developed into reagent grade or kit formats.   Methods for performing the above applications are well known to those skilled in the art. Open this method References shown are "Molecular Cloning: A Laboratory Manual", 2nd ed., Cold Spr. ing Harbor Laboratory Press, Sambrook, J., E.F. Fritsch and T. Maniatis eds., 1989, and "Methods in Enzymology: Guide to Molecular Cloning Techn. iques ", Academic Press, Berger, S.L. and A.R. Kimmel eds., 1987 But not limited to these.   Nutritional uses   The protein of the present invention can be used as a nutrient source or additive. Such uses , As a protein or amino acid additive, as a carbon source, as a nitrogen source And use as a carbohydrate source. Take In some cases, the protein or polynucleotide of the present invention may be added as a food for a particular organism. Well or separate, such as in the form of a powder, pill, solution, suspension or capsule It can also be administered as an individual or as a liquid formulation. In the case of microorganisms, The protein or polynucleotide of the present invention can be added to the medium to be cultured.   Cytokines and cell proliferation / differentiation activity   The protein of the present invention has cytokine activity, cell growth activity (induction or inhibition) or cell activity. Can show blast differentiation activity (induction or inhibition) or in certain cell populations To induce the production of other cytokines. Many proteins found to date Sex factors (including all known cytokines) are dependent on one or more factors Showing activity in cell proliferation assays, and thus the assay Serves as a convenient way to confirm activity. The activity of the protein of the present invention is determined in cell lines (32D, DA 2, DA1G, T10, B9, B9 / 11, BaF3, MC9 / G, M + (pr eB M +), 2E8, RB5, DA1, 123, T1165, HT2, CTL Including, but not limited to, L2, TF-1, Mo7e and CMK) Confirmed by any of the many conventional factor-dependent cell proliferation assays for You.   The activity of the protein of the invention may be measured, inter alia, by the following method:   Assays for T cell or thymocyte proliferation are described in Current Protocols in I mmunology, Ed by J. E. Coligan, A.M. Kruisbeek, D.H. Margulies, E.M. She vach, W Strober, Pub. Greene Publishing Associates and Wiley-Interscienc e (Chapter 3, In Vitroassays for Mouse Lymphocyte Function 3.1-3.19; Chap ter 7, Immunologic studies in Humans); Takai et al. Immunol. 137: 3494 -3500, 1986; Bertagnolli et al., J. Am. Immunol. 145: 1706-1712, 1990; Bertagno lli et al., Cellular Immunology 133: 327-341, 1991; Bertagnolli, et al., J . Immunol. 149: 3778-3783, 1992; Bowman et al., J. Am. Immunol. 152: 1756-1761 , 1994, but not limited thereto.   Cytokine production and / or expansion of spleen cells, lymph node cells or thymocytes Assays for propagation are described in Polyclonal T cell stimulation, Kruisbeek, A.M. and  Shevach, E.M. In Current Protocols in Immunology. J.E.e.a. Coligan eds . Vol 1 pp. 3.12.1-3.12.14, John Wiley and Sons, Toronto. 1994; and Measu rement of mouse and human Interferon γ, Schreiber, R.D. In Current Prot ocols in Immunology. J.E.e.a. Coligan eds. Vol 1 pp. 6.8.1-6.8.8, John W iley and Sons, Toronto. Includes, but is not limited to those described in 1994 No.   Assays for proliferation and differentiation of hematopoietic and lymphogenic cells ent of Human and Murine Interleukin 2 and Interleukin 4, Bottomly, K., D avis, L.S. and Lipsky, P.E. In Current Protocols in Immunology. J.E.e.a . Coliganeds. Vol 1 pp. 6.3.1-6.3.12, John Wiley and Sons, Toronto. 1991 ; deVries et al., J .; Exp. Med. 173: 1205-1211, 1991; Moreau et al., Nature 336: 690-692, 1988; Greenberger et al., Proc. Natl. Acad. Sci. U.S.A. 80: 2 931-2938, 1983; Measurement of mouse and human interleukin 6-Nordan, R.A. In Current Protocols in Immunology. J.E.e.a. Coligan eds. Vol 1 pp. 6.6. 1-6.6.5, John Wiley and Sons, Toronto. 1991; Smith et al., Proc. Natl. Ac ad. Sci. U.S.A. 83: 1857-1861, 1986; Measurement of human Interleukin 11 -Bennett, F., Giannotti, J., Clark, S.C. and Turner, K. J. In Current P rotocols in Immunology. J.E.e.a. Coliganeds. Vol 1 pp. 6.15.1 John Wiley  and Sons, Toronto. 1991; Measurement of mouse and human Interleukin 9-C iarletta, A., Giannotti, J., Clark, S.C. and Turner, K.J. In Current Pro tocols in Immunology. J.E.e.a. Collgan eds. Vol 1 pp. 6.13.1, John Wiley  and Sons, Toronto. Includes, but is not limited to, those described in 1991.   Assays for the response of T cell clones to antigens (particularly proliferation and APC-T cell interaction as well as direct (Identifying the effect of T cells) is described in Current Protocols in Immunology, Ed. E. Coligan, A.M. Kruisbeek, D.H. Margulies, E.M. Shevach, W Strober, Pub. G reene Publishing Associates and Wiley-Interscience (Chapter 3, In Vitro a ssays for Mouse Lymphocyte Function; Chapter 6, Cytokines and their cellu lar receptors; Chapter 7, Immunologic studies in Humans); Weinberger et al ., Proc. Natl. Acad. Sci. USA 77: 6091-6095, 1980; Weinberger et al., Eur . J. Immun. 11: 405-411, 1981; Takai et al., J. Am. Immunol. 137: 3494-3500, 1 986; Takai et al., J. Am. Immunol. 140: 508-512, including those described in 1988 But not limited to these.   Immunostimulatory or suppressive activity   In addition, the protein of the present invention has an activity (including but not limited to ), And may exhibit immunostimulatory or immunosuppressive activity. Protein is In various deficiencies and disorders, including severe immunodeficiency complications (SCID) May be useful, for example, to increase the proliferation of T and / or B lymphocytes In regulating or down-regulating, and N Useful in enhancing the cytolytic activity of K cells and other cell populations It is possible. These immune deficiencies can be genetic and can include viral ( HIV, as well as those caused by bacterial or fungal infections. Or may be caused by an autoimmune disease. Yo More specifically, the susceptibility caused by viruses, bacteria, fungi or other infectious agents Infectious diseases, e.g. For example, HIV, hepatitis virus, herpes virus, mycobacteria, reshmani A variety of fungal infections, such as A, Malaria and Candida species, are Is treatable. Of course, in this regard, a boost to the immune system dictates In other words, the protein of the present invention may be used in the treatment of cancer.   Autoimmune diseases that may be treated using the protein of the present invention include, for example, multiple sclerosis, Systemic deep elitomades, rheumatoid arthritis, autoimmune pneumonia, Guilla in-Barre syndrome, autoimmune thyroiditis, insulin-dependent diabetes, myasthenia gravis , Graft-versus-host disease and autoimmune inflammatory eye diseases. Of the present invention Such proteins can be used to treat allergic reactions such as asthma or other respiratory disorders and It may be used to treat symptoms. Other conditions for which immunosuppression is desired (eg, asthma (especially Allergic asthma) and related respiratory problems) using the protein of the present invention. You may be treated. Other conditions in which immunosuppression is desired (including, for example, organ transplantation) The treatment may be performed using the inventive protein.   The proteins of the present invention can also be used to enable an immune response in a number of ways. Da Unregulation can block or block an immune response already in progress Or includes interfering with the induction of an immune response. There may be. By suppressing the T cell response, or Inhibits activated T cell function by inducing resistance or both May be. Generally, immunosuppression of T cell responses is an active, non-antigen specific Process, which requires continuous exposure of T cells to the inhibitor. Resistance and Means non-responsive or anergic in T cells, generally antigen-specific In that it persists after exposure to the tolerizing agent has ceased. Operationally, the lack of a T cell response when exposed to a specific antigen in the absence of a resistance agent More resistance may be shown.   One or more antigen functions (eg, B lymphocyte antigen function (eg, B7) Including, but not limited to, down-regulating For example, blocking high levels of lymphokine synthesis by activated T cells Useful in tissue, skin and organ transplantation and graft versus host disease (GVHD) There will be. For example, blocking T cell function reduces tissue destruction in tissue transplantation Should be. Typically, in tissue transplants, graft rejection is Initiated by T cells being recognized as foreign and then breaking the graft A destructive immune response occurs. B7 Lymphocyte Antigen on Immune Cells and Its Native Riga A molecule that inhibits or blocks interaction with another B lymphocyte antigen (eg, , B7-1, B7-3), in the form of a monomer having the activity of Or a single, soluble, monomeric peptide having B7-2 activity) Pre-implant administration is essential for immune cells without transmitting responsive costimulatory signals. May induce binding of the molecule to its ligand. B lymphocytes in this regard Blocking antigen function depends on cytokine synthesis by immune cells such as T cells. It interferes with growth and thus acts as an immunosuppressant. Besides, lack of co-stimulation May be sufficient to cause a loss of T cell response. B lymphocyte antigen blocking test The induction of long-term tolerance by drugs allows repeated administration of these blocking reagents. The need can be avoided. To achieve sufficient immunosuppression or tolerance in the subject May also need to block the function of the B lymphocyte antigen combination No.   Individual blocking in preventing organ transplant rejection or GVHD Evaluate the effectiveness of the reagents using animal models that can predict their effectiveness in humans be able to. Examples of suitable systems that can be used include allografts in rats and allografts. Xenogeneic pancreatic islet cell grafts in mice and Was used to test the immunosuppressive effect of the CTLA4Ig fusion protein (Lenscho w et al., Science 257: 789-792 (1992) and Turka et al., Proc Natl. Acad. Sci. USA, 89: 11102-11105 (1992)). In addition, GVHD mice Mimodel (Paul ed., Fundamental Immunology, Ravan Press, New York, 1989) , Pp. 846-847) using B lymphocytes to progress the disease in vivo. The blocking effect of the antigen function can be determined.   In addition, blocking antigen function is therapeutically useful for treating autoimmune diseases It can be. Many autoimmune diseases are responsive to self- Inappropriate activity of T cells to promote the production of cytokines and autoantibodies involved in It is the result of sexualization. Interfering with the activation of self-reactive T cells reduces the symptoms of the disease. Less or may be eliminated. Disrupting B lymphocyte antigen receptor: ligand interaction Inhibit T cell activation using administration of a reagent that blocks T cell costimulation Production of autoantibodies or T cell-derived cytokines that can harm and participate in the disease process Can interfere with life. In addition, blocking reagents can provide long-term remission of disease It may induce antigen-specific resistance of autoreactive T cells that may lead. Autoimmune disease The effectiveness of blocking reagents in the prevention or amelioration of Can be determined using a number of well-characterized animal models You. Examples are murine experimental autoimmune encephalitis, MRLlpr / lpr mice or N Systemic deep erythematosus and murine self-immunity in ZB hybrid mice Plague collagen arthritis, diabetes in NOD mice and BB rats, and Experimental rat myasthenia gravis (Paul ed., Fundamental Immunology, Raven Press) , New York, 1989, pp. 840-856).   Antigen function as a means of up-regulating the immune response (preferred Alternatively, up-regulation of B lymphocyte antigen function) is also therapeutically useful. Up-regulation of the immune response may be May be in a form that eliminates the initial immune response. For example, B lymphocyte antigen function Enhancing the immune response by stimulating may be useful in the case of a viral infection. In addition, systemic viral such as influenza, common cold, and encephalitis Disease may be ameliorated by systemic administration of a stimulatory form of B lymphocyte antigen .   Alternatively, T cells are taken from a patient and tagged with ACPs that express a peptide of the invention. Co-stimulate T cells in vitro with added viral antigens, or Is combined with a stimulating form of the soluble peptide of the invention and then activated in vitro Re-introduced T cells into the patient, the anti-virus in infected patients It may enhance the immune response. Another method of promoting an antiviral immune response is Infected cells are taken from the patient and the nucleic acid encoding the protein of the invention described herein is To allow cells to express all or part of the protein on their surface. And then reintroduce the transfected cells into the patient. U. The infected cells then transmit a costimulatory signal to the T cells in vivo. Could thereby activate T cells.   In another application, antigen function (preferably B lymphocyte antigen function) Up-regulation or promotion may be useful in inducing tumor immunity . Transfection with a nucleic acid encoding at least one peptide of the invention Tumor cells (eg, sarcoma, melanoma, lymphoma, leukemia, neuroblastoma, cancer Tumor) can be administered to a subject to overcome tumor-specific resistance in the subject. Desired Transfect tumor cells to express a combination of peptides be able to. For example, a tumor cell obtained from a patient is transformed into a peptide having B7-2-like activity. Peptide having only tide or B7-1-like activity and / or B7-3-like activity Expression vector that directs expression in combination with Can be transfected. Transfected tumor cells To the patient to express the peptide on the surface of the transfected cells. Let Alternatively, transfection in vivo using gene therapy methods To target tumor cells.   Existence of the peptide of the present invention having B lymphocyte antigen activity on the surface of tumor cells Provides the necessary co-stimulatory signals for T cells to provide T cell-mediated trafficking. Induces an immune response against the transfected tumor cells. In addition, MHC Tumor cells lacking class I or MHC class II molecules, or sufficient MHC Tumor cells that cannot reexpress class I or MHC class II molecules are I α chain protein and β_TwoMicroglobulin protein or MHC class II α chain or Or all or part of the MHC class II β chain protein (eg, Transfection with the nucleic acid encoding the Class I or class II MHC proteins can be expressed on the cell surface . A marker having the activity of a B lymphocyte antigen (eg, B7-1, B7-2, B7-3) Expression of the appropriate class I or class II HMC in combination with the peptide is Elicits an immune response to transfected tumor cells mediated by vesicles Lead. If desired, expression of MHC class II binding proteins, such as invariant chains, can be blocked. Check A gene encoding an antisense construct that expresses B lymphocyte antigen activity. Co-transfection with DNA encoding the peptide It can also promote presentation of tumor-associated antigens and induce tumor-specific immunity. Therefore, Induction of an immune response mediated by T cells in a subject It may be sufficient to overcome heterogeneity.   The activity of the protein of the invention may be measured, inter alia, by the following method:   Suitable assays for thymocyte or spleen cell cytotoxicity are described in Current Protocols in Immunology, Ed by J. E. Coligan, A.M. Kruisbeek, D.H. Margulies, E.M . Shevach, W Strober, Pub. Greene Publishing Associates and Wiley-Inters cience (Chapter 3, In Vitro assays for Mouse Lymphocyte Function 3.1-3.1 9; Chapter 7, Immunologic studies in Humans); Herrmann et al., Proc. Natl . Acad. Sci. USA 78: 2488-2492, 1981; Herrmann et al., J. Am. Immunol. 128: 196 8-1974, 1982; Handa et al. Imrnunol. 135: 1564-1572, 1985; Takai et al. , J. et al. Immunol. 137: 3494-3500, 1986; Takai et al., J. Am. Immunol. 140: 508-512 Herrmann et al., Proc. Natl. Acad. Sci. USA 78: 2488-2492, 1981; He rrmann et al., J. Amer. Immunol. 128: 1968-1974, 1982; Handa et al., J. Am. Immunol . 135: 1564-1572, 1985; Takai et al., J. Am. Imnlunol. 137: 3494-3500, 1986; Bow manet al., J. et al. Virology 61: 1992-1998; Takai et al., J. Am. Immunol. 140: 508-51 2, 1988; Bertagnolliet al., Cellular Immunology 133: 327-341, 1991; Brown e t al., J.S. Immunol. 153: 3079-3092, including the assay described in 1994, Not limited to these.   Updates for T cell-dependent immunoglobulin response and isotype switching Say (especially to modulate the T cell-dependent antibody response to a Th1 / Th2 profile Influencing proteins are identified in Maliazewski, J. Immunol. 144: 3028-3033, 1990. Includes, but is not limited to, the assays described, and further enhances B cell function Say, In vitro antibody production, Mond, J.J. and Brunswick, M. In Current  Protocols in Immunology J.E.Coliganeds.Val 1 pp.3.8.1-3.8.16, John Wiley  and Sons, Tronto 1994.   Mixed lymphocyte reaction (MLR) assays (especially primarily Th1 and CTL Identify the protein that produces the answer), Current Protocols in Immunology, Ed by  J. E. Coligan, A.M. Kruisbeek, D.H. Margulies, E.M. Shevach, W Strober, Pub . Greene Publishing Associates and Wiley-Interscience (Chapter 3, In Vitr o assays for Mouse Lymphocyte Function 3.1-3.19; Chapter 7, Immunologics tudies in Humans); Takai et al. Immunol. 137: 3494-3500, 1986; Takai et  al., J. et al. Immunol. 140: 508-512, 1988; Bertagnolli et al., J. Am. Immunol. 149: 3778-3783, including but not limited to the assays described in 1992.   Dendritic cell-dependent assays (especially for dendritic cells that activate untreated T cells) To identify proteins that are more expressed) are described in Guery et al., J. Am. Immunol. 134: 536-544 Inaba et al., Journal of Experimental Medicine 173: 549-559, 1991; Macatonia et al., Journal of Immunology 154: 5071-5079, 1995; Porgador et al., Journal of Experimental Medicine 182: 255-260, 1995; Nair et al., Jou rnal of Virology 67: 4062-4069, 1993; Huang et al., Science 264: 961-965, 1 994; Macatonia et al., Journal of Experimental Medicine 169: 1255-1264, 19 89; Bhardwaj et al., Journal of Clinical Investigation 94: 797-807, 1994; a nd Inaba et al., Journal of Experimental Medicine 172: 631-640, 1990. Includes, but is not limited to, the assays listed.   Lymphocyte survival / apoptosis assays (especially apoptosis after superantibody induction) Identify Proteins That Prevent Lysis and that Regulate Lymphocyte Homeostasis ) Is Darzynkiewicz et al., Cytometry 13: 795-808, 1992; Gorczyca et al., Leukemia 7: 659-670, 1993; Gorczyca et al., Cancer Research 53: 1945-1951, 1993; Itoh et al., Cell 66: 233-243, 1991; Zacharchuk, Journal of Immunolog. y 145: 4037-4045, 1990; Zamal et al., Cytometry 14: 891-897, 1993; Gorczyca et al., International Journal of Oncology 1: 639-648, 1992. Includes, but is not limited to:   Early stage T cell commitment and development assays Antica et al., Blood 84: 111-117, 1994; Fine et al., Cellular Immunology 155: 111-122, 1994; Galy et al., Blood 85: 2770-2778, 1995; Toki et al., Proc. Natl. A cad.Sci. USA 88: 7548-7551, 1991, including but not limited to Absent.Hematopoietic regulatory activity   The proteins of the present invention are useful in regulating hematopoiesis and are therefore Useful in the treatment of bulb deficiency. Colony forming cells or factor-dependent cell lines Even minimal biological activity in support of has been implicated in regulating hematopoiesis. For example, to support the growth of erythroid progenitor cells alone, or to In combination with, for example, treatment of various anemias, or release Production of erythroid progenitors and / or erythroblasts in combination with radiation therapy / chemotherapy Stimulating viability; proliferation of bone marrow cells such as granulocytes and monocytes / macrophages Support (ie, traditional CSF activity), eg, in combination with chemotherapy In addition, it is useful in preventing subsequent myelosuppression; Breeding, then supporting platelet growth, and thereby thrombocytopenia It is useful for preventing or treating various platelet diseases, and Used instead of or in addition to platelet infusion; and / or hematopoietic stem Cells (mature to all of the above hematopoietic cells, and therefore various stem cell diseases ( Diseases that are usually treated by transplantation, such as aplastic anemia and development It is useful for supporting the growth of the hemoglobinuria And even after radiation / chemotherapy in vivo or ex vivo (ie, Or combined with bone marrow transplantation) or genetically engineered for gene therapy It is also useful in later repopulation of the stem cell compartment as normal cells.   In particular, the activity of the protein of the present invention may be measured by the following method.   Assays suitable for proliferation and differentiation of various hematopoietic cell lines have already been cited .   Assays for embryonic stem cell differentiation, specifically identifying proteins that affect embryonic hematopoiesis ) Is Johansson et al. Cellular Biology 15: 141-151, 1995; Keller et al., Mol ecular and Cellular Biology 13: 473-486,1993; McClanahan et al., Blood81: 29 Includes, but is not limited to, the assays described in 03-2915,1993.   Assays for stem cell survival and differentiation, specifically identifying proteins that regulate lympho-hematopoiesis ) Is described in Methylcellulose colony forming assays, Freshney, M.G. In Cultu re of Hematopoietic Cells. R.I. Freshney, et al. eds. Vol pp. 265-268, W iley-Liss, Inc., New York, NY. 1994; Hirayama et al., Proc. Natl. Acad. Sci. USA 89: 5907-5 911, 1992; Primitive hematopoietic colony forming cells with high prolife rative potential, McNiece, I.K. and Briddell, R.A. In Culture of Hematop oietic Cells. R.I. Freshney, et al. eds. Vol pp. 23-39, Wiley-Liss, Inc. New York, NY. 1994; Neben et al., Experimental Hematology 22: 353-359, 1 994; Cobblestone area forming cell assay, Ploemacher, R.E. In Culture of Hematopoietic Cells. R.I. Freshney, et al. eds. Vol pp. 1-21, Wiley-Liss , Inc .., New York, NY. 1994; Long term bone marrow cultures in the presen ce of stromal cells, Spooncer, E., Dexter, M. and Allen, T .; In Culture o f Hematopoietic Cells. R.I. Freshney, et al. eds. Vol pp. 163-179, Wiley -Liss, Inc., New York, NY. 1994; Long term culture initiating cell assay Sutherland, H.J. In Culture of Hematopoietic Cells. R.I. Freshney, et al. eds. Vol pp. 139-162, Wiley-Liss, Inc., New York, NY. Described in 1994 Assays, including but not limited to:   Tissue proliferation activity   The protein of the present invention may be used to grow or regenerate bone, cartilage, keys, ligaments and / or nerve tissue. And compositions used for wound healing and tissue repair, further including burns, lacerations And has utility in the treatment of ulcers.   Book that induces cartilage and / or bone growth in an environment where bone is not formed normally Inventive proteins cure healing of fractures and cartilage damage or defects in humans and other animals There are applications in Such formulations using the protein of the present invention include closed fractures and It is useful for reducing open fractures and improving fixation of artificial joints. Bone De novo bone formation induced by plasticizers can be congenital, traumatic or tumor Contributes to the repair of craniofacial deficits induced by radiological resection and further It is also useful in general surgery.   The proteins of the present invention may be used in the treatment of periodontal disease and other tooth restoration processes. Heel Agents provide an environment to attract osteogenic cells, stimulate the growth of osteogenic cells, Alternatively, it can induce osteogenic cell precursor differentiation. For example, the protein of the present invention And / or mediated by inflammatory or inflammatory processes Block the process of tissue destruction (collagenase activity, osteoclast activity, etc.) This may be useful in the treatment of osteoporosis or osteoarthritis.   Another category of tissue developmental activity that may be attributed to the proteins of the invention is the key. / Ligament formation. Environments where key / ligament-like or other tissues are not formed properly The protein of the present invention that induces the formation of such a tissue in humans is Applied to healing of key or ligament lacerations, deformations and other key or ligament defects You. Such a formulation using a key / ligament-like tissue-inducing protein may be used for key or ligament-like tissue. To prevent key or ligament fixation to bone or other tissue It may be. The de novo key / ligament-like tissue formation induced by the composition of the present invention Sexual, traumatic, or oncological resection-induced craniofacial defects Cosmetic surgery for contributing to the repair and also for attaching or repairing keys or ligaments It is also useful in The composition of the present invention attracts key- or ligament-forming cells Provide an environment for stimulating the growth of key- or ligament-forming cells, Induction of precursors of band-forming cells or tissue repair when returned to vivo. Ex vivo can induce the growth of key / ligament cells or progenitors ex vivo. The compositions of the present invention can be used to treat keratitis, carpal tunnel syndrome and other key or ligament defects. Is also useful. The composition of the present invention may comprise a suitable matrix and / or It may include a sequestering agent, such as a well-known carrier.   The protein of the present invention is used for proliferation of nerve cells and regeneration of nerve and brain tissue, , Central and peripheral nervous system diseases and neuropathy and mechanical diseases and Treatment of traumatic diseases (including degeneration, death or trauma to nerve cells or nerve tissue) May also be useful for treatment. More specifically, peripheral nerve injury, peripheral neuropathy and Diseases of the peripheral nervous system such as neuropathy and localized neuropathy, and Alzheimer's disease , Parkinson's disease, Huntington's disease, amyotrophic lateral spinal sclerosis and Shy-Drager Proteins may be used in the treatment of diseases of the central nervous system, such as the syndrome. By the present invention Additional conditions that may be treated include mechanical disorders such as spinal cord disorders and traumatic disorders And cerebrovascular diseases such as head trauma and stroke. Chemotherapy or other of Peripheral neuropathy caused by medical therapy is cured using the protein of the present invention. Can be treated.   The protein of the present invention may also be used for pressure ulcer, ulcer associated with vascular dysfunction, Non-healing wounds, including but not limited to wounds due to trauma, etc. It may also be useful for promoting quick closure.   The protein of the present invention includes organs (eg, pancreas, liver, intestine, kidney, skin, endothelium). ), Muscle (smooth, skeletal or cardiac) and vascular (including vascular endothelium) tissue To promote the development of other tissues or the proliferation of cells that make up such tissues. Activity. Part of the desired effect is by suppressing fibrotic scars. Regeneration of normal tissue. The proteins of the present invention may also exhibit angiogenic activity.   The protein of the present invention is useful for protecting or regenerating intestine, and fibrosis of lung or liver, Treatment of tissue reperfusion injury and symptoms caused by systemic cytokine damage May also be useful for treatment.   The protein of the present invention promotes or suppresses differentiation of the above-mentioned tissue from precursor tissue or cells; Alternatively, it may be useful for suppressing the growth of the tissue.   The activity of the protein of the invention may be measured, inter alia, by the following method:   Assay for tissue regeneration activity is described in WO 95/16035 (bone, cartilage, key) International Publication WO95 / 05846 (nerves, neurons); International Publication WO91 / 05 7491 (skin, endothelium), including but not limited to .   Wound healing activity assays are described in Maibach, HI and Rovee, DT, eds., Year Book M edical Publishers, Inc., Chicago (Eaglstein and Mertz, J. Invest. Dermato l 71: 382-384 (modified by 1978)). Not limited to these.   Activin / inhibin activity   Also, the protein of the present invention may exhibit activin- or inhibin-related activity. I Inhibins are characterized by their ability to inhibit follicle stimulating hormone (FSH) release, Activins are characterized by their ability to stimulate the release of follicle stimulating hormone (FSH). You. Therefore, the protein of the present invention may be used alone or as a member of the inhibin α family. In the form of a heterodimer with a female mammal, which reduces the fertility of a female mammal May be useful as an infertility drug based on the ability of inhibin to reduce spermatogenesis in animals You. Administration of sufficient amounts of other inhibins may result in infertility in these mammals. Can be guided. Alternatively, the proteins of the present invention may be homodimers or Is a heterodimer with other protein subunits of the inhibin-β group Based on the ability of activin molecules to stimulate FSH release from anterior pituitary cells And may be useful as a therapeutic agent that induces fertility. For example, US Pat. See 98885. In addition, the protein of the present invention is useful for reproduction in sexually immature mammals. It may be useful for enhancement and, as a result, homes such as cattle, sheep and pigs Increased fertility during the life of the animal.   The activity of the protein of the invention may be measured, inter alia, by the following method:   Assays for activin / inhibin activity are described in Vale et al., Endocrinology 91: 5. 62-572,1972; Ling et al., Nature 321: 779-782,1986; Vale et al., Nature 321: 7 76-779, 1986; Mason et al., Nature 318: 659-663, 1985; Forage et al., Proc. Natl Acad. Sci. USA 83: 3091-3095, 1986, including but not limited to No.   Chemotaxis / chemokinesis activity   The protein of the present invention can be used for mammalian cells such as monocytes, neutrophils, T cells, mast cells, and eosinophils. Chemotactic or chemokinetic activity of spheres and / or endothelial cells (eg, Acting as in). Uses chemotactic and chemokinetic proteins And recruit or attract the desired cell population to the desired site of action. Chemotaxis Alternatively, chemokinesis proteins may be used to treat and localize tissue injuries and other trauma. It is particularly advantageous for treating localized infections. For example, tumors of lymphocytes, monocytes or neutrophils Attraction to tumor or infected site may improve immune response to tumor or infectious agent You.   Certain proteins or peptides have chemotactic activity for specific cell populations. Direct or indirect, if stimulated, the direction or behavior of such cell populations. Command movement. Preferably, the protein or peptide has a directed movement of the cell. Have the ability to directly stimulate Specific proteins have chemotactic activity on cell populations Is used to determine whether such proteins or peptides have known chemotaxis Can be easily determined.   The activity of the protein of the invention may be measured, inter alia, by the following method:   Assay for chemotaxis activity (an assay to identify proteins that induce or interfere with chemotaxis) Say) describes the ability of the protein to induce cell translocation across the membrane as well as the Includes assays that measure the ability of a protein to induce adhesion to other cell populations. Moving Assays suitable for and adhesion are described in Current Protocols in Immunology, Ed by J.E. Coligan, A.M. Kruisbeek, D.H. Margulies, E.M. Shevach, W. Strober, Pub. G reene Publishing Associates and Wiley-Interscience (Chapter 6.12, Measur ement of alpha and beta Chemokines 6.12.1-6.12.28; Taub et al. J. Clin. I nvest. 95: 1370-1376, 1995; Lind et al. APMIS 103: 140-146, 1995; Muller et al Eur. J. Immunol. 25: 1744-1748; Gruber et al. J. of Immunol. 152: 5860-5 867, 1994; Johnston et al. J. of Immunol. 153: 1762-1768, 1994. Includes but is not limited to Say.   Hemostasis and thrombolytic activity   In addition, the protein of the present invention may exhibit hemostatic or thrombolytic activity. As a result Thus, such proteins are useful in the treatment of various clotting disorders, including genetic disorders such as hemophilia. Or in the treatment of injury caused by trauma, surgery or other causes. Blood clots and other hemostasis. The protein of the present invention may be used to dissolve or form a thrombus. Growth inhibition and the symptoms resulting therefrom (eg, myocardial infarction and central nervous system blood) It may be useful in the treatment and prevention of vascular infarcts (eg, stroke).   The activity of the protein of the invention may be measured, inter alia, by the following method:   Assays for haemostatic and thrombolytic activity are described by Linet et al., J. Clin. Pharmacol. 26: 1. 31-140, 1986; Burdick et al., Thrombosis Res. 45: 413-419, 1987; Humphrey et al., Fibrinolysis 5: 71-79 (1991); Schaub, Pr ostaglandins 35: 467-474, 1988, including but not limited to No.   Receptor / ligand activity   Further, the protein of the present invention may be a receptor, a receptor ligand or a receptor / ligand interaction. May exhibit activity as inhibitors or agonists. Such receptors and Examples of gand are cytokine receptors and their ligands, receptor kinases and And their ligands, receptor phosphatases and their ligands, cells Receptors involved in cell interactions and their ligands (cell adhesion molecules (SELEC , Integrin and their ligands) and antigen presentation, antigens Receptor / ligand pairs involved in the development of cognitive, cellular and humoral immune responses Inclusive) but not limited to. Receptors and ligands are related receptors Screening of potential peptide or small molecule inhibitors for body / ligand interactions It is also useful for training. The protein of the present invention (receptor and ligand fragments Include, but are not limited to, blocking the receptor / ligand interaction itself. May be useful as a harmful agent.   The activity of the protein of the invention may be measured, inter alia, by the following method:   Suitable assays for receptor-ligand activity are described in Current Protocols in Immunolog. y, Ed by J.E. Coligan, A.M. Kruisbeek, D.H. Margulies, E.M. Shevach, W.S trober, Pub. Greene Publishing Associates and Wiley-Interscience (Chapter  7.28, Measurement of Cellular Adhesion under static conditions 7.28.1-7 .28.22), Takai et al., Proc. Natl. Acad. Sci. USA 84: 6864-6868, 1987; Bie reretal., J. Exp. Med. 168: 1145-1156, 1988; Rosenstein et al., J. Am. Exp. Me d. 169: 149-160 1989; Stoltenborg et al., J. Am. Immunol. Methods 175: 59-68, 1 994; Stitt et al., Cell 80: 661-670, 1995. Not limited to these.   Anti-inflammatory activity   The protein of the present invention can exhibit anti-inflammatory activity. Anti-inflammatory activity is involved in the stimulus response By stimulating cells, cell-cell interaction (eg, attachment) Chemotaxis of cells involved in the inflammatory process by inhibiting or promoting By inhibiting or promoting, by inhibiting or promoting cell extravasation, Alternatively, it stimulates the production of other factors that more directly inhibit or promote the inflammatory response or Is exhibited by suppressing. Symptoms of inflammation ( Includes chronic or acute symptoms, infection-related inflammation (including septic shock, sepsis) Or systemic inflammatory response syndrome (SIRS)), ischemia-reperfusion injury, Fatal injury by dotoxin, arthritis, hyperacute rejection mediated by complement, Nephritis, lung injury induced by cytokines or chemokines, inflammatory bowel disease Disease, Crohn's disease or overproduction of cytokines such as TNF or IL-1 (Including but not limited to diseases resulting from). Departure Myelin protein is a cure for anaphylaxis and hypersensitivity to antigenic substances or materials. May also be useful for treatment.   Tumor inhibitory activity   In addition to the activities described above for the immunological treatment or prevention of tumors, the protein of the present invention Can exhibit antitumor activity. Certain proteins directly or indirectly affect tumor growth (eg, (Via ADCC). Certain proteins are found in tumor or tumor precursor tissue Act to inhibit the formation of tissue necessary to support tumor growth (eg, angiogenesis) Other factors, agents or cell types that inhibit tumor growth Factors, agents or agents that cause the production of Alternatively, a tumor inhibitory activity may be exhibited by removing or inhibiting a cell type.   Other activities   The proteins of the present invention may exhibit one or more of the following additional activities or effects: Infectious agents, including but not limited to bacteria, viruses, fungi and other parasites Killing of factors; height, weight, hair color, eye color, skin or other tissue pigmentation, Or the size or form of an organ or body part (eg, breast augmentation or vice versa) Influence on the physical properties involved (suppression or promotion); consumed fat, protein or carbohydrates Effects of chloride on digestion; appetite, libido, stress, cognition (including cognitive disorders), depression ( Behavioral characteristics including (but not limited to) depressive illness and violent behavior Of analgesic or other pain-relieving effects; embryos in non-hematopoietic lineages Promotion of stem cell differentiation and proliferation; hormonal or endocrine activity; Correcting deficiency and treating related disorders; hyperproliferative disorders (eg, psoriasis) Therapy; immunoglobulin-like activity (eg, the ability to bind antibodies or complement); Such a protein or such a protein acting as an antigen in a vaccine composition Ability to generate an immune response to other substances or entities that cross react with.Administration and dose   The protein of the invention (from any source, recombinant and non-recombinant) Methods, including, but not limited to, those described above) with a pharmaceutically acceptable carrier. They may be used together in a pharmaceutical composition. Such compositions comprise (in addition to the protein and carrier, ), Diluents, fillers, salts, buffers, stabilizers, solubilizers, and the like. It may contain other substances well known in the above. The term "pharmaceutically acceptable" A non-toxic substance that does not interfere with the effectiveness of the biological activity of the active ingredient. Characteristics of carrier Sex depends on the route of administration. The pharmaceutical composition of the present invention comprises M-CSF, GM-CSF, T NF, IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL- 7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IFN, TNF0, TNF1, TNF2, G-CSF , Meg-CSF, thrombopoietin, stem cell factor, and erythroboyet Contain cytokines, lymphokines, or other hematopoietic factors, such as Good. The pharmaceutical composition further enhances the protein activity or its activity in therapy. Or it may contain other agents that supplement its usefulness. Such additional factors and A synergistic effect with the protein of the present invention by containing Or minimize side effects. Conversely, certain cytokines, Lymphokines, other hematopoietic factors, thrombolytic or antithrombotic factors, or anti-inflammatory agents Including the protein of the present invention in the formulation of, cytokines, lymphokines, other hematopoietic factors May minimize the side effects of thrombolytic or antithrombotic factors, or anti-inflammatory agents .   The protein of the present invention may be a multimer (for example, a heterodimer or a homodimer) or It may be active by itself or in a complex with other proteins. As a result, The pharmaceutical composition comprises such a multimeric or complexed form of the protein of the present invention. You may.   The pharmaceutical composition of the present invention may be in the form of a complex of the protein of the present invention and a protein or peptide antigen. It may be in a state. Protein and / or peptide antigens can be It will deliver a stimulus signal to the lymphocytes. B lymphocytes have their surface immunity Will respond to antigen through globulin receptors. T lymphocytes are MHC proteins Will respond to the antigen through the T cell receptor (TCR). MHC and host cell class I and class II MHC genes Structurally related proteins, including those that have been loaded, are peptide-antigens against T lymphocytes. It will help to present the original. The antigen component was a purified MHC-peptide complex only. Or with co-stimulatory molecules that can send signals directly to T cells Is done. Alternatively, it binds to surface immunoglobulins and other molecules on B cells The resulting antibodies can also be mixed with the pharmaceutical compositions of the present invention.   The pharmaceutical composition of the present invention may be in the form of liposome, in which the protein of the present invention is contained. And other pharmaceutically acceptable carriers, amphipathic agents such as lipids (mice in aqueous solution). Agglomerate form, such as insoluble monolayer, liquid crystal or lamellar layer) Have been combined. Suitable lipids for liposome formulations are monoglycerides, diglycerides , Sulfatizide, lysolecithin, phospholipids, saponins, bile acids, etc. However, it is not limited to these. The preparation of such liposome formulations is within the level of ordinary skill in the art. And, for example, U.S. Pat. Nos. 4,235,871, 4501728, 4837028, And 4737323, all of which are described herein by reference. Is considered).   As used herein, the term "therapeutically effective amount" refers to a significant patient benefit, e.g., Sufficient pharmaceutical composition or method to show improvement in symptoms of the condition, healing, increased rate of healing It means the total amount of each active ingredient. Used for individual active ingredients administered alone When used, the term refers only to that component. When used in combinations of active ingredients, The total amount of active ingredients that produces a therapeutic effect, regardless of whether U.   In practicing the treatment method of the present invention, a disease to be treated with a therapeutically effective amount of the protein of the present invention. Administered to a mammal having a morphology. According to the method of the present invention, alone or with a cytokine The protein of the invention together with other therapeutic agents such as lymphokines or other hematopoietic factors. It may be administered. One or more cytokines, lymphokines or other When co-administered with hematopoietic factors, the protein of the present invention may be administered with cytokines, lymphokines, and other It may be administered simultaneously or sequentially with blood, thrombolytic or antithrombotic factors. Gradually When administering the next dose, the attending physician will ask for cytokines, lymphokines, other hematopoietic factors, blood Appropriate administration of thrombolytic factor or antithrombotic factor in combination with the protein of the present invention Will determine the order.   The administration of the protein of the present invention in the pharmaceutical composition of the present invention or used in the practice of the present invention can be carried out by various conventional methods. Administration methods, e.g., oral feeding, inhalation, or intradermal, subcutaneous, or intravenous injection. I can. Intravenous injection into a patient is preferred.   When a therapeutically effective amount of the protein of the present invention is orally administered, the protein of the present invention may contain , Powder, solution or elixir form. When administered in tablet form, The pharmaceutical composition may further comprise a solid carrier such as gelatin or an adjuvant. It may be. Tablets, capsules, and powders contain about 5 to 95% of a protein of the present invention, It preferably contains about 25-90% of the protein of the invention. Place to administer in liquid form Oil, water, petroleum, oils of animal or vegetable origin, such as peanut oil, mineral oil , Soybean oil, sesame oil, or synthetic fats and oils may be added. Pharmaceutical group in liquid form The composition can be a saline solution, dextrose or other saccharide solution, or glycol (Eg, ethylene glycol, propylene glycol or polyethylene glycol) Recall). Pharmaceutical compositions when administered in liquid form Is about 0.5 to 90% by weight of the protein of the present invention, preferably about 1 to 50% Invention Contains white.   When a therapeutically effective amount of the protein of the present invention is administered by intravenous, intradermal or subcutaneous injection, The protein of the present invention may be in the form of a pyrogen-free, parenterally acceptable aqueous solution . Of such a parenterally acceptable protein solution having the appropriate pH, isotonicity, and stability. Preparation is within the skill of the art. Preferred medicament for intravenous, intradermal or subcutaneous injection The composition comprises, in addition to the protein of the present invention, sodium chloride for injection, Ringer's solution, Dextrose, dextrose and sodium chloride solution for injection, lactic acid for injection Should contain Ringer's solution, or other carriers known in the art . The pharmaceutical composition of the present invention may comprise a stabilizer, a preservative, a buffer, an antioxidant, Other additives known to the consumer.   The amount of the protein of the invention in the pharmaceutical composition of the invention depends on the nature and severity of the condition to be treated, As well as the nature of the treatment the patient has already received. Ultimately, your doctor Each patient will determine the amount of the protein of the invention to be treated. First, the doctor in charge Administer an amount of the protein of the invention and observe the patient's response. Until optimal therapeutic effects are obtained Higher doses of the protein of the present invention may be administered and may be used once optimal therapeutic effect is obtained. Do not increase the volume further. Various pharmaceutical compositions for performing the methods of the present invention include About 0.01 μg to about 100 mg of the protein of the present invention (preferably, About 0.1 μg to about 10 mg / kg body weight, more preferably 1 kg / kg body weight 0.1 to about 1 mg of the protein of the present invention.   The duration of treatment by intravenous administration using the composition of the present invention depends on the severity of the disease to be treated. And will vary depending on the condition and response of each patient. Each of the proteins of the present invention The duration of application will range from 12 to 24 hours for continuous intravenous administration . Ultimately, the attending physician will determine the stage of treatment by intravenous administration using the pharmaceutical composition of the present invention. Will decide between.   Immunizing an animal with the protein of the present invention to specifically react with the protein of the present invention; And monoclonal antibodies may be obtained. Whole protein or its fragment Such antibodies may be obtained using immunoglobulin as an immunogen. Carboxy peptide immunogen It may further contain a cysteine residue at the end, and can be used for keyhole rimpet hemoglobin. Shi Binds to haptens such as anine (KLH). Method for synthesizing such a peptide Are known in the art, for example, R.P.Merrifield, J. Amer. Chem. Soc. 85, 2149-2154 (1963); J.L. Krstenansky, et.al., FEBS Lett. 211, 10 (1987). Monoclonal antibodies that bind to the protein of the present invention are useful for immunodetection of the protein of the present invention. It can be a diagnostic. Neutralizing antibodies that bind to the protein of the present invention And may be associated with abnormal expression of the protein of the present invention. May be useful in the treatment of certain tumors in the treatment of some forms of cancer You. In the case of cancer cells or white blood cells, a neutralizing monoclonal antibody against the protein of the present invention The body is useful for detecting and preventing metastatic spread of cancer cells that can be mediated by the proteins of the present invention It can be.   For compositions of the invention useful for regenerating bone, cartilage, tendons or ligaments, the method of treatment comprises: The composition can be administered locally, systemically, or locally as an implant or device. Administration. When administered, the therapeutic composition used in the present invention comprises Of course, it is a pyrogen-free, physiologically acceptable form. In addition, Alternatively, the composition may be encapsulated or injected as a viscous form to provide bone, cartilage or May be delivered to the site of tissue damage. Local administration is suitable for wound healing and tissue repair I do. Therapeutically useful action other than the protein of the present invention which may be contained in the above composition The agent is, alternatively or additionally, administered simultaneously or sequentially with the composition in the method of the present invention. May be. Preferably, for bone and / or cartilage formation, the composition comprises Delivering the white-containing composition to the site of bone and / or cartilage damage, and developing the bone and cartilage; Provides a structure for living, and optimally contains a matrix that can be absorbed into the body Will have. Such matrices are currently used for other implanted medical devices It may be made from the materials that have been used.   The choice of matrix material depends on its biocompatibility, biodegradability, mechanical properties, cosmetic Based on appearance and interface properties. The appropriate formulation will be determined by the particular application of the composition. U. Possible matrices for the composition are biodegradable, chemically known sulfuric acid Lucium, tricalcium phosphate, hydroxyapatite, polylactic acid, polyglyco ー It may be luic acid and polyanhydride. Other usable materials are biodegradable and Well known in the art, for example, bone or skin collagen. Furthermore The tricks may include pure proteins or extracellular matrix components. other Possible matrices include sintered hydroxyapatite, bioglass, aluminum Is not biodegradable, such as metal or other ceramics, but is chemically known It is. The matrix is a combination of materials of the above type, for example polylactic acid and Contains hydroxyapatite or collagen and tricalcium phosphate You may. Bioceramics may be added to the composition, for example, calcium-aluminate. -May be modified in the phosphate and processed to produce pore size, particle size, Child shape and biodegradability may be varied.   Lactic acid in the form of porous particles having a diameter in the range of 150 to 800 microns and A 50:50 (molar weight) copolymer of biglycolic acid is presently preferred. . In some applications, carboxymethylcellulose or autologous Prevent the dissociation of the protein composition from the matrix using a sequestering agent such as a clot And would be useful.   Preferred families of sequestrants are methylcellulose, ethylcellulose, Roxyethyl cellulose, hydroxypropyl cellulose, hydroxypropyl Alkylcells including methylcellose and carboxymethylcellulose With cellulosic materials such as Lurose (including hydroxyalkylcellulose) Yes, cationic salts of carboxymethylcellulose (CMC) are most preferred . Other preferred sequestering agents are hyaluronic acid, sodium alginate, poly (ethylene) Glycol), polyoxyethylene oxide, carboxyvinyl polymer and And poly (vinyl alcohol). The amount of sequestrant useful in the present invention depends on the total formulation. It is 0.5 to 20% by weight, preferably 1 to 10% by weight, and this amount is Prevents protein desorption from the polymer matrix and ensures proper handling of the composition In an amount that allows it, the progenitor cells invade the matrix, To give proteins the opportunity to promote the osteogenic activity of carcinoma cells, there is not much Not to be.   In a further composition, the protein of the present invention comprises a bone and / or cartilage defect, wound, Alternatively, it may be mixed with other agents that are beneficial in treating the tissue. These agents are , Epidermal growth factor (EGF), platelet-derived growth factor (PDGF), transformed growth factor (TGF-α and TGF-β) and insulin-like growth factor (IGF) Include.   At present, the therapeutic compositions are also valuable for veterinary use. For details, In addition to humans, livestock and thoroughbred horses are treated with the protein of the present invention. Is a desirable patient.   Rules of administration of protein-containing pharmaceutical compositions used for tissue regeneration alter the effect of proteins Factors, such as tissue weight, damage site, and damage desired to be formed. The condition of the tissue received, the size of the wound, the type of tissue required (eg, bone), the patient's year Consider age, gender and diet, severity of infection, duration of administration, and other clinical factors And your doctor will decide. The dose depends on the type of matrix used for reconstitution. And depending on the content of other proteins in the pharmaceutical composition. For example, IGF   Addition of other known growth factors, such as I (insulin-like growth factor I) to the final composition Addition can also affect dosage. Tissue / bone growth and / or repair, By regular assessment by morphological survey and tetracycline labeling The progress can be monitored.   The polynucleotide of the present invention can also be used for gene therapy. Such polynuks Leotide is introduced into cells in vivo or ex vivo and expressed in a mammalian subject. Can be made. Other known methods for introducing nucleic acids into cells or organisms The polynucleotide of the present invention may be administered more (in a viral vector, Or in the form of naked DNA, but is not limited thereto).   Culturing and growing the cells ex vivo in the presence of the protein of the invention, or Produce the desired effect on such cells or produce the desired activity in such cells. May be. The treated cells are then introduced in vivo for therapeutic purposes. can do.   The patents and publications cited herein are hereby incorporated by reference in their entirety. It is assumed that

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) A61P 43/00 111 C12P 21/02 C C07K 14/47 C12N 15/00 ZNAA C12N 5/10 5/00 B C12P 21/02 A61K 37/02 (31) Priority claim number 08 / 739,066 (32) Priority date October 28, 1996 (Oct. 28, 1996) (33) Priority claim country United States (US) (81) Designated countries EP (AT, BE, CH, DE, DK, ES, FI, FR, GB, GR, IE, IT, LU, MC, NL, PT, SE), OA (BF, BJ, CF) , CG, CI, CM, GA, GN, ML, MR, NE, SN, TD, TG), AP (GH, KE, LS, MW, SD, SZ, UG, ZW), EA (AM, AZ, B , KG, KZ, MD, RU, TJ, TM), AL, AM, AT, AU, AZ, BA, BB, BG, BR, BY, CA, CH, CN, CU, CZ, DE, DK, EE, ES, FI, GB, GE, HU, IL, IS, JP, KE, KG, KP, KR, KZ, LC, LK, LR, LS, LT, LU, LV, MD, MG, MK, MN, MW , MX, NO, NZ, PL, PT, RO, RU, SD, SE, SG, SI, SK, TJ, TM, TR, TT, UA, UG, UZ, VN (72) Inventor Ravalee, Edward Earl United States 01876 Green Meadow Drive, Tuxbury, Mass. 90 (72) Inventor Lacey, Lisa A. United States 01720 Acton, Mass., School Street 124 (72) Inventor Marberg, Ibid United States 01720 Acton, Massachusetts, Orchard Drive No. 2 (72) Inventor Tracy, Maurice United States 02167 Massachusetts, Chestnut Hill, Walcott Road No. 93 (72) Inventor Spalding, Vickey United States 01821 Billerica, Mass. Meadowbank Road 11

Claims (1)

[Claims] 1. (A) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1;   (B) the nucleotide sequence from nucleotide 477 to nucleotide 752 of SEQ ID NO: 1 A polynucleotide comprising a reotide sequence;   (C) of clone AX568 deposited under accession number ATCC 98101 A polynucleotide comprising the nucleotide sequence of the full-length protein coding sequence;   (D) of clone AX568 deposited under accession number ATCC 98101 Polynucleotide encoding full-length protein encoded by cDNA insert Tide;   (E) of clone AX568 deposited under accession number ATCC 98101 A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence;   (F) of clone AX568 deposited under accession number ATCC 98101; Polynucleotide encoding mature protein encoded by cDNA insert Tide;   (G) a polynucleotide encoding a protein comprising the amino acid sequence of SEQ ID NO: 2 Tide;   (H) including a fragment of the amino acid sequence of SEQ ID NO: 2 having biological activity A polynucleotide encoding a protein;   (I) of the isolate AX5628 deposited under accession number ATCC 98180 A polynucleotide comprising the nucleotide sequence of the full-length protein coding sequence;   (J) of isolate AX5628 deposited under accession number ATCC 98180 Polynucleotide encoding full-length protein encoded by cDNA insert Tide;   (K) of isolate AX5628 deposited under accession number ATCC 98180 A polynucleotide comprising the nucleotide sequence of the mature protein coding sequence;   (L) of isolate AX5628 deposited under accession number ATCC 98180 Polynucleotide encoding mature protein encoded by cDNA insert Tide;   (M) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 5;   (N) the nucleotide sequence from nucleotide 307 to nucleotide 402 of SEQ ID NO: 5 A polynucleotide comprising a reotide sequence;   (O) a polynucleotide encoding a protein comprising the amino acid sequence of SEQ ID NO: 6 Tide;   (P) including a fragment of the amino acid sequence of SEQ ID NO: 6 having biological activity A polynucleotide encoding a protein;   (Q) Alleles of the polynucleotides (a) to (e) or (i) to (n) above A polynucleotide that is a genetic variant;   (R) encodes a species homolog of the protein of (g), (h), (o) or (p). Polynucleotide A composition comprising an isolated polynucleotide selected from the group consisting of:   2. 2. The polynucleotide of claim 1, wherein said polynucleotide is operably linked to an expression control sequence. Composition.   3. A host cell transformed with the composition of claim 2.   4. 4. The host cell of claim 3, wherein said cell is a mammalian cell.   5. (A) growing the culture of the host cell of claim 3 in a suitable medium;   (B) A method for producing a protein, comprising purifying the protein from a culture.   6. A protein produced by the method of claim 5.   7. 7. The protein of claim 6, comprising a mature protein.   8. A protein comprising an amino acid sequence selected from the group consisting of: A composition comprising a protein substantially free of other mammalian proteins:   (A) amino acid sequence of SEQ ID NO: 2:   (B) a fragment of the amino acid sequence of SEQ ID NO: 2; and   (C) of clone AX568 deposited under accession number ATCC 98101 an amino acid sequence encoded by the cDNA insert;   (D) the amino acid sequence of SEQ ID NO: 6;   (E) a fragment of the amino acid sequence of SEQ ID NO: 6; and   (F) of isolate AX5628 deposited under accession number ATCC 98180 Amino acid sequence encoded by the cDNA insert.   9. 9. The composition of claim 8, wherein said protein comprises the amino acid sequence of SEQ ID NO: 2.   10. 9. The composition of claim 8, further comprising a pharmaceutically acceptable carrier.   11. Administering a therapeutically effective amount of the composition of claim 10 to a mammalian subject. How to prevent, treat or ameliorate a medical condition.   12. A gene corresponding to the cDNA sequence of SEQ ID NO: 1.   13. This corresponds to the cDNA sequence of SEQ ID NO: 4, SEQ ID NO: 5 or SEQ ID NO: 7. The corresponding gene.   14． 2. The polynucleotide of claim 1, wherein said polynucleotide comprises the nucleotide sequence of SEQ ID NO: 1. Composition.   15. The polynucleotide is a nucleoside from nucleotide 477 of SEQ ID NO: 1 2. The composition of claim 1, comprising up to 752 nucleotide sequences.   16. The polynucleotide was deposited under accession number ATCC 98101. Comprising the nucleotide sequence of the full length protein coding sequence of clone AX568; The composition of claim 1.   17． The polynucleotide was deposited under accession number ATCC 98101. The full-length protein encoded by the cDNA insert of clone AX568 was The composition of claim 1, comprising a polynucleotide to be loaded.   18. The polynucleotide was deposited under accession number ATCC 98101. Comprising the nucleotide sequence of the mature protein coding sequence of clone AX568. The composition of claim 1.   19. The polynucleotide was deposited under accession number ATCC 98101. The mature protein encoded by the cDNA insert of clone AX568 was The composition of claim 1, comprising a polynucleotide to be loaded.   20. The polynucleotide encodes a protein comprising the amino acid sequence of SEQ ID NO: 2. The composition of claim 1, comprising a polynucleotide to be loaded.   21. The polynucleotide of SEQ ID NO: 2 having biological activity 2. A polynucleotide comprising a polynucleotide encoding a protein comprising a fragment of the sequence. Composition.   22. The polynucleotide was deposited under accession number ATCC 98180. The nucleotide sequence of the full length protein coding sequence of isolate AX5628, The composition of claim 1.   23. The polynucleotide was deposited under accession number ATCC 98180. The full-length protein encoded by the cDNA insert of isolate AX5628 was The composition of claim 1, comprising a polynucleotide to be loaded.   24. The polynucleotide was deposited under accession number ATCC 98180. The nucleotide sequence of the mature protein coding sequence of isolate AX5628; The composition of claim 1.   25. The polynucleotide was deposited under accession number ATCC 98180. The mature protein encoded by the cDNA insert of isolate AX5628 was The composition of claim 1, comprising a polynucleotide to be loaded.   26. 2. The polynucleotide of claim 1, wherein said polynucleotide comprises the nucleotide sequence of SEQ ID NO: 5. Composition.   27. The polynucleotide is a nucleoside from nucleotide 307 of SEQ ID NO: 5 2. The composition of claim 1 comprising a nucleotide sequence up to Pide 402.   28. The polynucleotide encodes a protein comprising the amino acid sequence of SEQ ID NO: 6. The composition of claim 1, comprising a polynucleotide to be loaded.   29. The polynucleotide of SEQ ID NO: 6 having biological activity 2. A polynucleotide comprising a polynucleotide encoding a protein comprising a fragment of the sequence. Composition.   30. Clone AX wherein the protein has been deposited under accession number ATCC 98101. 568. comprising an amino acid sequence encoded by a 568 cDNA insert. The composition of 8.   31. 9. The composition of claim 8, wherein said protein comprises the amino acid sequence of SEQ ID NO: 6.   32. The isolate AX5, wherein the protein has been deposited under accession number ATCC 98180. 628 comprising an amino acid sequence encoded by the 628 cDNA insert. The composition of 8.