MXPA00011729A

MXPA00011729A - Interleukins-21 and 22

Info

Publication number: MXPA00011729A
Application number: MXPA/A/2000/011729A
Authority: MX
Inventors: Steven M Ruben; Reinhard Ebner
Original assignee: Reinhard Ebner; Human Genome Sciences Inc; Steven M Ruben
Priority date: 1998-05-29
Filing date: 2000-11-28
Publication date: 2001-11-21

Abstract

The present invention relates to novel human proteins designated Interleukin-21 (IL-21) and Interleukin-22 (IL-22), and isolated polynucleotides encoding these proteins. Also provided are vectors, host cells, antibodies,and recombinant methods for producing these human proteins. The invention further relates to diagnostic and therapeutic methods useful for diagnosing and treating disorders related to these novel human proteins.

Description

INTERLEUCINAS 21 AND 22 FIELD OF THE INVENTION The present invention relates to two novel human genes, each of which codes for a polypeptide that is a member of the Interleukin family. More specifically, the present invention relates to a polynucleotide that codes for a novel human polypeptide called Interleukin 21 or "IL-21". The present invention also relates to a polynucleotide that codes for a novel human polynucleotide called Interleukin 22 or "IL-22". This invention also relates to IL-21 and IL-22 polypeptides, as well as to vectors, to host cells, antibodies directed against IL-21 and IL-22 polypeptides, and recombinant methods for producing them. Diagnostic methods are also provided to detect disorders related to the immune system, and therapeutic methods to treat such disorders. The invention further relates to screening methods for identifying agonists and antagonists of the activity of IL-21 and I L-22.

Ref: 125088 BACKGROUND OF THE INVENTION Cytokines typically exert their respective biochemical and physiological effects by binding to specific receptor molecules. The link to the receptor then stimulates the pathways of specific signal transduction (Kishimoto, T., et al., Cell 76: 253-262 (1994)). The specific interactions of cytokines with their receptors are often the primary regulators of a wide variety of cellular processes including activation, proliferation, and differentiation (Arai, K.-I, et al., Ann.Rev. Bi. Ochem., 59 : 783-836 (1990), Paul, WE and Seder, RA, Cel l 1 6: 241-251 (1994)). Human interleukin (IL) -17, a homolog closely related to the molecules of the present invention, was only recently identified. IL-17 is a 155 amino acid polypeptide that was molecularly cloned from the CD4 + T-cell cDNA library (Yao, Z., et al., J. Immun ol. 155; 5483-5486 (1995)). The IL-17 polypeptide contains an N-terminal signal peptide and contains approximately 72% identity at the amino acid level with a herpesvirus gene üi & ifia ^. ^ ftj «« ite-W6e-a && ^.

T-cell trophic saimiri (HVS) designated HVS13. High levels of IL-17 are secreted from primary peripheral blood leukocytes, positive to CD4 (PBL) after stimulation (Yao, Z., et al., Immuni ty 3: 811-821 (1995)). The treatment of fibroblasts with IL-17, HVS13, or another murine homologue, designated CTLA8, activates the signal transduction pathways, and results in the stimulation of the family of the transcription factor NF-kappaB, the secretion of IL -6 and the co-stimulation of T cell proliferation (Yao, Z., et al., Immuni ty 3: 811-821 (1995)). A fusion protein HVS13-Fc was used to isolate a murine IL-17 receptor molecule that does not appear to belong to any of the families of cytosine receptors previously described (Yao, Z., et al., Immuni ty 3: 811 -821 (nineteen ninety five)). The murine IL-17 receptor (mIL-17R) is predicted to encode a type I transmembrane protein of 864 amino acids with an apparent molecular mass of 97.8 kDa. mIL-17R is predicted to possess an N-terminal signal peptide with a cleavage site between alanine 31 and serine 32. The molecule also contains an extracellular domain of 291 amino acids, a transmembrane domain of 21 amino acids, and a cytoplasmic tail of 521 amino acids. A soluble recombinant IL-17R molecule consisting of 323 amino acids of the extracellular domain of IL-17R fused to the Fe portion of the human IgGl immunoglobulin was able to significantly inhibit the IL-6 production induced by IL-17 by murine NIH- cells. 3T3 (s upra). Interestingly, the expression of the IL-17 gene is highly restricted. This is typically observed mainly in activated T lymphocyte memory cells (Broxmeyer, H. J. Exp. Med. 183: 2411-2415 (1996); Fossiez, F., et al., J. Exp. Med. 183: 2593-2603 (1996)). Conversely, the IL-17 receptor appears to be expressed in a large number of cells and tissues (Rouvier, E., et al., J. Immun ol. 150: 5445-5456 (1993); Yao, Z., et al., J. Immun ol. 155: 5483-5486 (1995)). It remains to be seen, however, if IL-17 itself can play an autocrine role in the expression of IL-17. IL-17 has been implicated as a causative agent in the expression of IL-6, IL-8, G-CSF, Prostaglandin E (PGE2), and the intracellular adhesion molecule (ICAM) -l (Fossiez, F., s upra; Yao, Z., et al., Immuni ty 3: 811-821 (1995)). Each of these molecules possesses highly relevant and potentially therapeutically valuable properties. For example, IL-6 is involved in the regulation of the hematopoietic line and in the growth of progenitor cells and their expansion (Ikebuchi, K., et al., Proc. Nati. Acad. Sci. USA 84: 9035-9039 (1987), Gentile, P. and Roxmeyer, HE Ann., NY Acad. Sci. USA 628: 74-83 (1991)). IL-8 shows myelosuppressive activity for totipotential cells and immature subgroups of myeloid progenitors (Broxmeyer, H.E., et al., Ann. Hematol. 71: 235-246 (1995); Daly, T.J., et al., J. Biol. Chem. 270: 23282-23292 (nineteen ninety five)). G-CSF acts early and late to activate and stimulate hematopoiesis in general, and more specifically on neutrophil hematopoiesis, while PGE2 improves erythropoiesis, suppresses lymphopoiesis and myelopoiesis in general, and strongly suppresses monocytopoiesis (Broxmeyer, HE AMER J. Ped Hematol Oncol 14: 22-30 (1992) Broxmeyer HE and Williams DE CRC Crit. Rev. Oconol. Hematol 8: 173-226 (1988)). Thus, there is a need for polypeptides that function as immunoregulatory molecules and thereby modulate the transfer of an extracellular signal eventually to the cell nucleus, since disturbances of such regulation may be involved in disorders related to cellular activity, to haemostasis, to angiogenesis, to tumor metastasis, to cell migration and ovulation, as well as to neurogenesis. Therefore, there is a need for the identification and characterization of human polypeptides that may play a role in the detection, prevention, amelioration or correction of such disorders.

BRIEF DESCRIPTION OF THE INVENTION The present invention relates to the novel polynucleotides and to the encoded polypeptides of IL-21 and IL-22. In addition, the present invention relates to vectors, cells host, antibodies, and recombinant methods to produce the polypeptides and polynucleotides. Diagnostic methods are also provided for detecting disorders related to the polypeptides, and therapeutic methods for treating such disorders. The invention also relates to ^ .fr ~ * »» > - »selection methods to identify the link partners of 11-21 and IL-22.

BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 shows the partial nucleotide sequence (SEQ ID NO: 1) and the deduced amino acid sequence (SEQ ID NO: 2) of IL-21. The preserved I-IV Domains sites (see below) are underlined and marked as such. Figures 2A and 2B show the nucleotide sequence (SEQ ID NO: 3) and the deduced amino acid sequence (SEQ ID NO: 4) of IL-22. The sites of conserved I-IV domains (see below) are underlined and marked as such. The locations of two potential N-linked glycosylation sites are identified by a bold asparagine symbol (N) accompanied by a bold number sign (#) located above the initial nucleotide of the codon coding for the corresponding asparagine. Figures 3A, 3B and 3C show the regions of identity between the amino acid sequences of: (1) human Interleukin 17 (designated IL-17.aa in the figure; GenBank Accession No. U32659; SEQ ID NO: 5); (2) mouse Interleukin-17 (designated mIL-17.aa in the figure; GenBank Accession No. U43088; SEQ ID NO: 6); (3) viral Interleukin-17 (designated vIL-17.aa in the figure; GenBank Accession No. X64346; SEQ ID NO: 7); (4) IL-20 (designated IL-20.aa in the figure and described in the co-pending US Provisional Patent Application Serial No. 60 / 060,140, filed September 26, 1997, SEQ ID NO: 8); (5) a part-length IL-21 protein (SEQ ID NO: 2); (6) the full length IL-21 protein (designated IL-21FL.aa in the figure); (7) a partial-length protein IL-22 (designated IL-22.aa in the figure), and (8) an IL-22 protein (designated IL22ext.aa in the figure), as determined by the alignment of the sequences using the MegAlign component of the computer program DNA * Star ( DNASTAR, Inc., 1228 S. Park St., Madison, Wl 53715 USA) using the default parameters. Figure 4 shows an analysis of the partial amino acid sequence of IL-21 (SEQ ID NO: 2). The alpha, beta, back and helix regions; hydrophilicity and hydrophobicity; the unfriendly regions; the flexible regions; the antigenic index and the superficial probability are also shown. In the graph "Antigenic Index" or "Jameson-Wolf", the positive peaks indicate the locations of the highly antigenic regions of the IL-21 protein, ie, the regions from which epitope-bearing peptides can be determined. , of the invention. The polypeptides and polynucleotides that code for the polypeptides comprising the domains defined by these graphs are contemplated by the present invention. Figure 5 shows an analysis of the amino acid sequence of IL-22. The alpha, beta, back and helix regions are shown; hydrophilicity and hydrophobicity; the unfriendly regions; the flexible regions; the antigenic index and the superficial probability. In the graph "Antigenic Index" or "Jameson-Wolf", the positive peaks indicate the locations of the highly antigenic regions of the IL-22 protein, ie, the regions from which the peptides of the invention can be determined. that have an epitope. The polypeptides and polynucleotides that code for the polypeptides comprising the domains defined by these graphs are contemplated by the present invention. The data presented in Figure 5 are also represented in tabular form in Table II. The columns are marked with the headings "Res", "Position" and Roman Numbers I-XIII. The column headings refer to the following characteristics of the amino acid sequence presented in Figure 5 and Table II: "Res": amino acid residue of SEQ ID NO: 4 or Figures 2A and 2B; "Position": position of the corresponding residue within SEQ ID NO: 4 or Figures 2A and 2B; I: Alpha-Garnier-Robson regions; II: Alpha-Chou-Fasman regions; III: Beta-Garnier-Robson Regions; IV: Beta-Chou-Fasman Regions; V: Regions of Vuelta-Garnier-Robson; VI: Regions of Vuelta-Chou-Fasman; VII: Regions of Helix-Garnier-Robson; VIII: Hydrophilicity Chart-Kyte-Doolittle; IX: Alpha-Eisenberg Antipathetic Regions; X: Beta-Eisenberg Antipyretic Regions; XI: Flexible Regions-Karplus-Schulz; XII: Antigenic-Jameson-Wolf index; and XIII: Surface Probability Graph-Emini. Figures 6A and 6B show the nucleotide sequence (SEQ ID NO: 28) and the deduced amino acid sequence (SEQ ID NO: 29) of full-length IL-21. The locations of the conserved I-IV domains (identical to those shown in Figure 1) and of the conserved V-VII domains are underlined and marked as such. A signal peptide predicted from methionine-1 to alanine-18 is doubly underlined. Figure 7 shows an analysis of an amino acid sequence of full-length IL-21. The alpha, beta, back and helix regions are shown; hydrophilicity and hydrophobicity; the unfriendly regions; the flexible regions; the antigenic index and the superficial probability. In the graph "Antigenic Index" or "Jameson-Wolf", the positive peaks indicate the locations of the highly antigenic regions of a full length IL-21 protein, ie, the regions from which the peptides can be determined. of the invention that possess epitope. The polypeptides and polynucleotides that code for the polypeptides comprising the domains defined by these graphs are contemplated by the present invention. The data presented in Figure 7 are also represented in tabular form in the ^ i ^ a, Table I. The columns are marked with the headings "Res", "Position", and Roman Numbers I-XIV. The column headings refer to the following characteristics of the amino acid sequence presented in Figure 7 and Table I: "Res": amino acid residue of SEQ ID NO: 29 or Figures 6A and 6B; "Position": position of the corresponding residue within SEQ ID NO: 29 or Figures 6A and 6B; I: Alpha-Garnier-Robson regions; II: Alpha-Chou-Fasman regions; III: Beta-Garnier-Robson Regions; IV: Beta-Chou-Fasman Regions; V: Regions of Vuelta-Garnier-Robson; VI: Regions of Vuelta-Chou-Fasman; VII: Regions of Helix-Garnier-Robson; VIII: Hydrophilicity Chart-Kyte-Doolittle; IX: Antipathic Regions Al fa-Eisenberg; X: Alfa-Eisenberg Antipatic Regions; XI: Antipática Beta-Eisenberg regions; XII: Flexible Regions-Karplus-Schulz; XIII: Antigenic-Jameson-Wolf index; and XIV: Surface Probability Graph-Emini. Figure 8 shows the nucleotide sequence (SEQ ID NO: 31) and the deduced amino acid sequence (SEQ ID NO: 32) of an IL-22. The locations of conserved I-IV and VI-VII Domains are underlined and marked as such. The locations of two N-glycosylation sites . JíL-J? ... -t tai. *. *. . «Flfc. lhasJ ». -,. »Í = &ÍÉfe! Fa, ¿S **« '^ =' ~ - «.» ^ Ii ^ i. - ~ a¿A ?? aa.jlt ¿k¿? mg '. . j'itoilit. The linked, potentials are identified by a bold asparagine symbol (N) accompanied by a bold number sign (#) located above the initial nucleotide of the codon coding for the corresponding asparagine. The two potential N-linked glycosylation sites are located at Asn-39 (N-39, A-40, S-41) and Asn-152 (N-152, S-153, S-154) of SEQ ID NO. : 32. Figure 9 shows an analysis of the amino acid sequence of IL-22 provided in Figure 8 and SEQ ID NO: 32. The alpha, beta, back and helix regions are shown; hydrophilicity and hydrophobicity; the unfriendly regions; the flexible regions; the antigenic index and the superficial probability. In the "Antigenic Index" or "Jameson-Wolf" graph, the positive peaks indicate the locations of the highly antigenic regions of a full length IL-22 protein, ie, the regions from which they can be determined. peptides of the invention that possess epitope. The polypeptides and polynucleotides that code for the polypeptides comprising the domains defined by these graphs are contemplated by the present invention.

The data presented in Figure 9 are also esented in tabular form in Table II. The columns are marked with the headings "Res", "Position" and Roman Numerals I-XIV. The column headings refer to the following characteristics of the amino acid sequence presented in Figure 9 and Table III: "Res": amino acid residue of SEQ ID NO: 32 or Figure 8; "Position": position of the corresponding residue within SEQ ID NO: 32 or Figure 8; I: Alfa-Garnier-Robson regions; II: Alpha-Chou-Fasman regions; III: Beta-Garnier-Robson Regions; IV: Beta-Chou-Fasman Regions; V: Regions of Vuelta-Garnier-Robson; VI: Regions of Vuelta-Chou-Fasman; VII: Regions of Helix-Garnier-Robson; VIII: Hydrophilicity Chart-Kyte-Doolittle; IX: Hopp-Woods Hydrophobicity Chart; X: Alfa-Eisenberg Antipatic Regions; XI: Antipática Beta-Eisenberg regions; XII: Flexible Regions-Karplus-Schulz; XIII: Antigenic-Jameson-Wolf index; and XIV: Surface Probability Graph-Emini.

DETAILED DESCRIPTION Definitions The following definitions are provided to facilitate the understanding of certain terms used throughout this specification. In the present invention, "isolated" refers to the material removed from its original environment (for example, the natural environment if it is of natural origin), and in this way is altered "by the hand of man" from its natural state . For example, an isolated polynucleotide could be part of a vector or a composition of matter, or it could be contained within a cell, and still be "isolated" because that particular vector, composition of matter, or cell is not the original environment of the polynucleotide. However, a nucleic acid contained in a clone that is a member of a library (e.g., a genomic or cDNA library) that has not been isolated from other members of the library (e.g., in the form of a homogeneous solution). containing the clone and other members of the library) or that is contained in a chromosomal preparation (eg, a chromosome spread), is not "isolated" for the purposes of this invention. In the present invention, a "secreted" IL-21 or IL-22 protein refers to a protein capable of being directed towards the ER, towards the secretory vesicles or towards the extracellular space as a result of a signal sequence, as well as a 11-21 or IL-22 protein released into the extracellular space without necessarily containing a signal sequence. If the secreted protein IL-21 or IL-22 is released into the extracellular space, the secreted protein IL-21 or IL-22 can undergo extracellular processing to produce a "mature" IL-21 or IL-22 protein. Release to the extracellular space can occur through many mechanisms, including exocytosis and proteolytic cleavage. As used herein a "polynucleotide" IL-21 or IL-22 refers to a molecule having an amino acid sequence contained in SEQ ID NO: 1 or SEQ ID NO: 3, respectively, or the cDNA contained within the respective clones deposited with the ATCC. For example, the polynucleotide IL-21 or IL-22 can contain the nucleotide sequence of the full length cDNA sequence, including the 5 'and 3' untranslated sequence, the coding region, with or without the signal sequence , the coding region of secreted protein, as well as fragments, epitopes, domains, and variants of the nucleic acid sequence. In addition, as used herein, a "polypeptide" IL-21 or IL-22 refers to a molecule having the translated amino acid sequence generated from the polynucleotide as broadly defined. As used herein, a "polynucleotide" IL-21 refers to a molecule having a nucleic acid sequence contained in SEQ ID NO: 1 or SEQ ID NO: 28, or the cDNA contained within the respective clones deposited with the ATCC. For example, the IL-21 polynucleotide may contain the nucleotide sequence of the full-length cDNA sequence, including the 5 'and 3' untranslated sequences, the coding region, with or without the signal sequence, the region of encoding secreted protein, as well as fragments, epitopes, domains, and variants of the nucleic acid sequence. In addition, as used herein, a "polypeptide" IL-21 refers to a molecule having the translated amino acid sequence generated from the polynucleotide as broadly defined. As used herein, a "polynucleotide" IL-22 refers to a molecule having a nucleic acid sequence contained in SEQ ID NO: 3 or SEQ ID NO: 31, or the cDNA contained within the respective clones deposited with the ATCC. For example, the IL-22 polynucleotide can contain the nucleotide sequence of the full-length cDNA sequence, including the 5 'and 3' untranslated sequences, the coding region, with or without the signal sequence, the coding region of the secreted protein, as well as the fragments, epitopes, domains, and variants of the nucleic acid sequence. In addition, as used herein, a "polypeptide" IL-22 refers to a molecule having the translated amino acid sequence generated from the polynucleotide as broadly defined. A representative clone containing all or most of the sequence for SEQ ID NO: 1 (designated HTGED19) was deposited with the Noteamerican Species Crop ("ATCC") collection on March 5, 1998, and he was given the Deposit Number 209666 of the ATCC. In addition, a representative clone containing all or most of the sequence for SEQ ID NO: 3 (designated HFPBX96) was also deposited with the ATCC on March 5, 1998, and was given Deposit Number 209665 of the ATCC. The ATCC is located at 10801 University Blvd., Manassas, VA 20110-2209, USA. The deposit with the ATCC was made in accordance with the terms of the Budapest Treaty on the international recognition of the deposit of microorganisms for patent procedure purposes. A "polynucleotide" IL-21 also includes those polynucleotides capable of hybridizing, under stringent hybridization conditions, to the sequences contained in the S ^ EQ ID NO: 1 or in the SEQ ID NO: 28, the complements thereof, or the cDNA inside the deposited clone. In addition, a "polynucleotide" IL-22 also includes those polynucleotides capable of hybridizing, under stringent hybridization conditions, to the sequences contained in SEQ ID NO: 3 or SEQ ID NO: 31, the components thereof, or the cDNA inside the deposited clone. "Strict hybridization conditions" refers to an incubation ^^^^^^^^^^^ «^^^^^^^^^^^^^^ g ^ fa ^^^^^^^^^^^^^^ * of all night to 42 ° C in a solution comprising 50% formamide, 5X SSC (750 mM sodium chloride, 75 mM sodium citrate), 50 mM sodium phosphate (pH 7.6), 5x Denhardt's solution, 10% dextran sulfate , and 20 μg / ml of salmon sperm DNA, cut, denatured, followed by washing the filters in SSC O.lx at approximately 65 ° C. Also contemplated are nucleic acid molecules that hybridize to IL-21 and IL-22 polynucleotides at less stringent hybridization conditions. The changes in the requirement for hybridization and in the detection of the signal are mainly achieved through the manipulation of the concentration of formamide (lower percentages of formamide result in a decreased requirement); the salt conditions, or the temperature. For example, the lowest requirement conditions include an overnight incubation at 37 ° C in a solution comprising 6X SSPE (20X SSPE = 3M sodium chloride, 0.2M NaH2P04, 0.02M EDTA, pH 7.4), 0.5% of SDS, 30% formamide, 100 μg / ml of salmon sperm DNA; followed by washes at 50 ° C with IX SSPE, 0.1% SDS. In addition, to achieve even lower demands, the washings carried out following the demanding hybridization can be carried out at higher salt concentrations (for example 5X SSC). Note that variations in the above conditions can be achieved through the inclusion and / or substitution of the alternative blocking reagents used to suppress the background in the hybridization experiments. Typical blocking reagents include Denhardt reagent, BLOTTO, heparin, salmon sperm DNA, denatured, and commercially available formulations. The inclusion of specific blocking reagents may require modification of the hybridization conditions described above, due to problems with compatibility. Of course, a polynucleotide that hybridizes only to the polyA + sequences (such as any polyA + 3 '-terminal portion of a cDNA shown in the sequence listing), or to a complementary stretch of T (or U) residues, might not be included in the definition of "polynucleotide", since such a polynucleotide could hybridize to any nucleic acid molecule that contains a polyA + stretch or the complement thereof (e.g. & & ggSs¿éa3 .- & fcfc Mt *? ^ u £ 1ji? FitíMi¡MfflmM «*? && MM @ kñ» * aá. && amp; & amp; & practically any clone of double-stranded cDNA). The polynucleotides IL-21 and IL-22 can be composed of any polyribonucleotide or polydeoxyribonucleotide, which can be unmodified RNA or DNA or modified RNA or DNA. For example, the polynucleotides IL-21 and IL-22 can be composed of single stranded and double stranded DNA, DNA which is a mixture of single and double stranded regions, single and double stranded RNA, and RNA which is a mixture of single and double stranded regions, hybrid molecules comprising DNA and RNA which can be single-stranded or, more typically, double-stranded or a mixture of single-stranded and double-stranded regions. In addition, IL-21 polynucleotides may be composed of triple-stranded regions comprising RNA or DNA or RNA and DNA. The IL-21 polynucleotides may also contain one or more modified bases or backbones of DNA or RNA modified for stability or for other reasons. "Modified" bases include, for example, tritylated bases and unusual bases such as inosine. A variety of modifications can be made to DNA and RNA; thus, "polynucleotide" encompasses chemically, enzymatically or metabolically modified forms. The IL-21 and IL-22 polypeptides may be composed of amino acids linked to one another by peptide bonds or modified peptide bonds, for example, peptide isosteres, and may contain amino acids other than the 20 amino acids encoded by genes. The IL-21 and IL-22 polypeptides can be modified by natural processes such as post-translational processing, or through clinical modification techniques which are well known in the art. Such modifications are well described in the basic texts and in more detailed monographs, as well as in literature and voluminous research. Modifications can occur anywhere in the IL-21 and IL-22 polypeptides, including the peptide backbone, the amino acid side chains and the amino or carboxyl termini. It will be appreciated that the same type of modification may be present in the same or varying degrees at various sites in a given IL-21 or IL-22 polypeptide. Also, a given IL-21 or IL-22 polypeptide may contain many types of modifications. The IL-21 or IL-22 polypeptides may be branched, for example, as a result of ubiquitination, and these may be cyclic, with or without branching. The cyclic, branched and branched cyclic IL-21 and IL-22 polypeptides can result from natural post-translational processes or can be made by synthetic methods. Modifications include acetylation, acylation, ADP-ribosylation, amidation, covalent coupling of flavin, covalent coupling of a heme moiety, covalent coupling of a nucleotide or nucleotide derivative, covalent coupling of a lipid or lipid derivative, covalent coupling of phosphotidylinositol , crosslinking, cyclization, disulfide bond formation, demethylation, formation of covalent crosslinks, cysteine formation, pyroglutamate formation, formylation, gamma-carboxylation, glycosylation, GP1 anchor formation, hydroxylation, iodination, methylation, myristoylation, oxidation, pegylation, proteolytic processing, phosphorylation, prenylation, racemization, selenoylation, sulfation, addition of amino acids mediated by transference RNA, to proteins such as arginylation, and ubiquitination. (See, for example, PROTEINS-STRUCTURE AND MOLECULAR PROPERTIES, 2nd Edition, T.

E. Creighton, W. H. Freeman and Company, New York (1993); POSTTRANSLATIONAL COVALENT MODIFICATION OF PROTEINS, B. C. Johnson, Ed., Academic Press, New York, pp. 1-12 (1983); Seifter, et al., Me th. Enzymol. 182: 626-646 (1990); Rattan, et al., Ann. NY Aca d. Sci. 663: 48-62 (1992)). "SEQ ID NO: 1" and "SEQ ID NO: 28" refer to a polynucleotide sequence of IL-21 while "SEQ ID NO: 2" and SEQ ID NO: 29 refer to an IL polypeptide sequence. -twenty-one. Similarly, SEQ ID NO: 3 and SEQ ID NO: 31 refer to a polynucleotide sequence of IL-22 while SEQ ID NO: 4 and SEQ ID NO: 32 refer to a polypeptide sequence of IL-22. An IL-21 polypeptide "having biological activity" refers to polypeptides that exhibit activity similar, but not necessarily identical to, an activity of an IL-21 polypeptide, including mature forms, as measured in a particular biological assay, with or without dose dependence. In addition, an IL-22 polypeptide "having biological activity" refers to polypeptides that exhibit similar activity, but not necessarily identical to, an activity of an IL-22 polypeptide, including mature forms, - ^ 8fa. »« J¡a »i. -j,: .--. ^. , .. < ,. ^ ^^ as measured in a particular biological assay, with or without dose dependence. In the case where the dose dependency does exist, it does not need to be identical to that of the IL-21 or IL-22 polypeptide, but rather substantially similar to the dose dependence at a given activity compared to IL polypeptides. -21 or IL-22 (e.g., the candidate polypeptide will show greater activity or no more than about 25 times less, and preferably no more than about ten times less activity, and more preferably, no more than about three times less activity relative to to IL-21 polypeptide).

Polynucleotides and Polypeptides IL-21 and IL-22 Clone HTGED19, which codes for IL-21, was isolated from a cDNA library derived from apoptotic T cells. This clone contains the entire coding region identified as SEQ ID NO: 2. The deposited clone contains a cDNA that has a total of 705 nucleotides, which codes for a predicted, partial, open reading structure of 87 amino acid residues (see Figure 1). The partial open reading structure begins at a point in the entire IL-21 ORF such that the "G" at position 1 of SEQ ID NO: 1 is effectively in position 3 of a coding triplet. As such, the predicted partial polypeptide sequence of IL-21 is shown starting infrastructurally with an alanine residue at position 1 of SEQ ID NO: 2. The alanine residue at position 1 of SEQ ID NO: 2 is encoded by nucleotides 2-4 of the nucleotide sequence shown in SEQ ID NO: 1. The ORF shown as SEQ ID NO: 2 terminates at a stop codon at the position of nucleotides 263-265 of the nucleotide sequence shown as SEQ ID NO: 1. The predicted molecular weight of the partial IL-21 protein should be approximately 9.558 Daltones. An initial BLAST analysis of the expression of the IL-21 cDNA sequence against the HGS EST database has also revealed a highly specific expression of this cDNA clone. In such an analysis, the HTGED19 cDNA sequence seems to be found only in apoptotic T cells. Thus, 11-21 appears to be expressed in a highly restricted pattern limited to apoptotic T cells, and, for example, other populations of lymphocytes or other cells in a state of activation or at rest. Clone HTGED19, which codes for IL-21, was used to select a panel of bacterial artificial chromosomes containing various segments of human genomic DNA (Research Genetics, Inc.). A positive clone was sequenced to identify the potential splice donor and acceptor sites. The analysis of several sites revealed a partial ORF upstream (5 ') that, when it was placed immediately 5 'and within the structure with the existing IL-21 DNA sequence, it generated a complete ORF which codes for a polypeptide with additional sequence identity to the IL-17 family (see Figures 3A, 3B and 3C). A clone of the full length IL-21 ORF was constructed by combining the IL-21 exons amplified by PCR from the HTGED19 genomic clone. The clone has been deposited with the ATCC as Deposit No. PTA-69 of the ATCC of May 14, 1999. The nucleotide sequence of the full-length IL-21 clone contains the entire coding region identified as SEQ ID NO: * M £ gá¡ & i £ 29. The resulting clone contains an insert that has a total of 1067 nucleotides, which codes for a predicted open reading structure of 197 amino acid residues (see Figures 6A and 6B). The open reading frame begins at nucleotide position 34 in the complete IL-21 polynucleotide shown as SEQ ID NO: 28 (Figures 6A and 6B). The ORF terminates at a stop codon at the position of nucleotides 625-627 of the nucleotide sequence shown as SEQ ID NO: 28 (Figures 6A and 6B). The predicted molecular weight of the IL-21 polypeptide shown in Figures 6A and 6B and as SEQ ID NO: 29 should be approximately 21,764 Daltones. Further BLAST analysis of the expression of the full-length IL-21 cDNA sequence against the HGS EST database has also revealed a highly specific expression of this cDNA clone. In such an analysis, the full-length HTGED19 cDNA sequence appears to be found only in apoptotic T cells. Thus, IL-21 appears to be expressed in a highly restricted pattern limited to apoptotic T cells, and, for example, other subpopulations of lymphocytes or other cells in a state of activation or rest. A PCR product comprising exons 1 and 2 (based on the genomic organization predicted 5 above) has been amplified using an early stage, 12-week-old cDNA library as template DNA. This PCF product confirms that at least exons 1 and 2 of the genomic organization predicted above, exists as messenger RNA in early stage human embryos of at least 12 weeks of age. The clone HFPBX96, which codes for IL-22, was isolated from a cDNA library derived from the frontal cortex of epileptics. East The clone contains the entire coding region identified as SEQ ID NO: 4. The deposited clone contains a cDNA having a total of 1,642 nucleotides, which codes for a predicted, partial reading structure of 160 residues of amino acids (see Figures 2A and 2B). The partial open reading structure begins at a point in the entire IL-22 ORF, such that the "G" in position 1 of SEQ ID NO: 3 is effectively in position 2 of a coding triplet. As such, the polypeptide sequence of IL-22 predicted, g & a * a ^ w r- > partial, is shown beginning intrastructurally with an asparagine residue at position 1 of SEQ ID NO: 4. The asparagine residue at position 1 of SEQ ID NO: 4 is encoded by nucleotides 3-5 of the nucleotide sequence shown as SEQ ID NO: 3. The ORF shown as SEQ ID NO: 4 terminates at a stop codon at the position of nucleotides 483-485 of the nucleotide sequence shown as SEQ ID NO: 3. The predicted molecular weight of the Partial IL-22 protein should be approximately 17,436 Daltones. Clone HFPBX96, which codes for IL-22, was used to select a human fetal brain cDNA library containing approximately one million cDNA clones (Genome Systems, INc.). A positive clone was sequenced to identify 59 nucleotides of the additional 5 'sequence. The cDNA clone has been deposited with the ATCC as Deposit No. PTA-70 of ATCC on May 14, 1999. The extended IL-22 ORF analysis reveals a polypeptide with additional sequential identity to the IL-17 family ( see Figures 3A, 3B and 3C). The nucleotide sequence of the IL-22 clone of apparently still partial length contains the region of The entire coding identified as SEQ ID NO: 31. The resulting clone contains an insert having a total of 522 nucleotides, which codes for a predicted open reading structure of 5 174 amino acid residues (see Figure). 8). The open reading structure begins at the position of nucleotide 1 in the complete IL-22 polynucleotide shown in SEQ ID NO: 31 (Figure 8). The ORF ends at a stop codon in the nucleotide position 520-522 of the nucleotide sequence shown as SEQ ID NO: 31 (Figure 8). The predicted molecular weight of polypeptide II-22 shown in Figure 8 and as SEQ ID NO: 31 is about 19,636 Daltons. 15 Using the BLAST and MegAlign analyzes, SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 29, and SEQ ID NO: 32 each was found to be highly homologous to several members of the Interleukin family. Particularly, SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 29 and SEQ ID NO: 32 contain at least four domains homologous to the translation products of human mRNA for Interleukin (IL) -20 (Provisional Patent Application North American Copending Serial No. 60 / 060,140; filed on September 26, 1997; SEQ ID NO: # & amp; & amp; amp; 8), IL-17 (Genbank Accession No. U32659; SEQ ID NO: 5; see also Figures 3A, 3B and 3C), murine mRNA for Interleukin (IL) -17 (GenBank Accession No. U43088; SEQ ID NO: 6, see also Figures 3A, 3B and 3C), and the human viral mRNA for Interleukin (ID-17 (GenBank Accession No. X64346; SEQ ID NO: 7; see also Figures 3A, 3B and 3C) Specifically, the molecules of the present invention, in particular, SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 29, and SEQ ID NO: 32 share a high degree of sequential identity with IL-20, IL -17, mIL-17 and vIL-17 in the following conserved domains: (a) a predicted NXDPXRYP domain (where X represents any amino acid) located approximately at the amino acids valine-3 to proline-11 of SEQ ID NO: 2, serine-57 to proline-64 of SEQ ID NO: 4, valine-113 to proline-121 of SEQ ID NO: 29, serine-70 to proline-77 of SEQ ID NO: 32, and asparagine-79 to proline -86 of the amino acid sequence of human IL-17 (SEQ ID NO: 5); predicted CLCXGC domain (where X represents any amino acid) located approximately at the amino acids cysteine-19 to cysteine-24 of SEQ ID NO: 2, cysteine-72 to cysteine-77 of SEQ ID NO: 4, cysteine-129 to cysteine- 134 of SEQ ID NO: 29, cysteine-85 to cysteine-90 of SEQ ID NO: 32, and cysteine-94 to cysteine-99 of the amino acid sequence of human IL-17 (SEQ ID NO: 5); (c) a predicted LVLRRXP domain (where X represents any amino acid) located approximately at the amino acids leucine-46 to proline-52 of SEQ ID NO: 2, valine-99 to proline-105 of SEQ ID NO: 4, leucine -156 to proline-162 of SEQ ID NO: 29, valine-112 to proline-118 of SEQ ID NO: 32, and leucine-120 to proline-126 of the amino acid sequence of human IL-17 (SEQ ID. NO: 5); and (d) a predicted VXVGCTCV domain (where X represents any amino acid) located approximately at the valine-75 to valine-82 amino acids of SEQ ID NO: 2, isoleucine-121 to valine-128 of SEQ ID NO: 4, valine-187 to valine-192 of SEQ ID NO: 29, isoleucine-134 to valine-141 of SEQ ID NO: 32, and valine-140 to valine-147 of the amino acid sequence of human IL-17 (SEQ ID NO: 5). In addition, the full-length IL-21 molecule shown in Figures 6A and 6B (SEQ ID NO: 29) and the IL-22 molecule shown in Figure 8 (SEQ ID NO: 32) show several additional conserved domains, when compared to IL-20 and the other members of the IL-17 family as - -, '* f é ft- "f"' sfe ^ r-. shown in Figures 3A, 3B and 3C). These conserved Domains are underlined in Figures 6A and 6B and in Figure 8, and are marked as conserved Domains V, VI and VII. Specifically, the molecules of the present invention, in particular, SEQ ID NO: 29 and SEQ ID NO: 32, share a high degree of sequential identity with IL-20, IL-17, mlL-17 and vIL-17 in the following conserved domains: (a) a predicted PXCXSAE domain (where X represents any amino acid) located approximately at the amino acids proline-34 to glutamic acid-40 of SEQ ID NO: 29; (b) a predicted PXXLVS domain (where X represents any amino acid) located approximately at the amino acids proline-63 to serine-68 of SEQ ID NO: 29 and approximately er. the amino acids alanine-18 to serine-23 of SEQ ID NO: 32; and (c) a predicted RSXSPW domain (where X represents any amino acid) located approximately at the amino acids arginine-104 to tryptophan-109 of SEQ ID NO: 29 and approximately at the amino acids arginine-60 to tryptophan-65 of SEQ ID NO: 32. These polypeptide fragments of IL-21 and IL-22 are specifically contemplated in the present invention. Because each of these molecules of IL-17 and similar to IL-17 are & • * • & > • & > - thinks that they are important immunoregulatory molecules, the homology between these molecules of IL-17 and similar to IL-17 and IL-21 and IL-22 suggest that IL-21 and IL-22 can also be important immunoregulatory molecules. In addition, based on their apparent sequential identities with IL-17 and IL-20 (see Figures 3A, 3B and 3C), the full-length IL-21 and IL-22 polypeptides are each likely possessing a peptide leader sequence. signal, secretory, amino-terminal. Since the present invention appears to be the partial cDNA clones of the molecules of IL-21 (SEQ ID NOs: 1 and 2) and IL-22 (SEQ ID NOs: 3 and 4) (in addition to the IL-21 molecule of full length shown as SEQ ID NOs: 28 and 29 and the IL-22 molecule shown as SEQ ID NOs: 31 and 32), it is also contemplated that the translation products of SEQ ID NOs: 2, 4 and 32 of this invention will be promoted to enter the cellular secretory pathway by virtue of being expressed as fusion proteins comprising several different portions of the N-terminus of the IL-20 molecule of the co-pending US Provisional Application Serial No. 60 / 060,140 fused to the known coding sequence of the IL-21 or IL-22 molecules of the present invention. Such expression constructs will secrete the hybrid molecules IL-20 / IL-21 or IL-20 / IL-22 from the host cell. In one embodiment, the mature IL-21 protein used in these fusion proteins encompasses approximately amino acids 12-87 of SEQ ID NO: 2, while the IL-20/21 fusion protein encompasses approximately 104 or 113 amino acids N-terminals of IL-20, encoded within the structure with approximately amino acids 12-87 of IL-21 of SEQ ID NO: 2. In other embodiments, an IL-20/21 fusion protein encompasses approximately 104 or 113 N-terminal amino acids of IL-20 encoded intrastructurally with approximately amino acids 3-87 of the IL-21 protein of SEQ ID NO: 2. These polypeptide fragments of IL-21 are specifically contemplated in the present invention. In another embodiment, the mature IL-22 proteini used to generate these fusion proteins encompasses approximately amino acids 1-160 of SEQ ID NO: 4, while the IL-20/22 fusion protein encompasses approximately 95, 104 or 113 N-terminal amino acids of IL-20 encoded intrastructurally with approximately amino acids 1-160 of IL-22 of SEQ ID NO: 4. In other embodiments, the IL-22 protein used to generate these fusion proteins encompasses approximately amino acids 47-160 of SEQ ID NO: 4, while the IL-20/22 fusion protein encompasses approximately 95, 104 or 113 N-terminal amino acids of IL-20 encoded intrastructurally with approximately amino acids 1-160 of IL-22 of SEQ ID NO: 4. In still other embodiments, the IL-22 protein used to generate these fusion proteins encompasses approximately amino acids 56-160 of SEQ ID NO: 4, while the protein of fusion IL- 20/22 encompasses approximately 95, 104 or 113 N-terminal amino acids of IL-20 encoded intrastructurally with approximately amino acids 1-160 of IL-22 of SEQ ID NO: 4. In other additional embodiments, the IL protein -22 used to generate these fusion proteins encompasses approximately amino acids 65-160 of SEQ ID NO: 4, while the IL-20/22 fusion protein encompasses approximately 95, 104 or 113 N-terminal amino acids of IL -20, coded intrastructurally with approximately amino acids 1-160 of IL-22 of SEQ ID NO: 4. These polypeptide fragments of IL-22 are specifically contemplated in the present invention. In yet another embodiment, the mature IL-22 protein used to generate these fusion proteins encompasses approximately amino acids 1-173 of SEQ ID NO: 32, while the IL-20/22 fusion protein encompasses approximately 95, 104 or 113 N-terminal amino acids of IL-20 encoded intrastructurally with approximately amino acids 1-173 of IL-22 of SEQ ID NO: 32. These polypeptide fragments of IL-22 are specifically contemplated in the present invention. The nucleotide sequences of IL-21 and IL-22 identified as SEQ ID NO: 1 and SEQ ID NO: 3, respectively, were assembled from partially homologous ("overlapping") sequences obtained from the deposited clones. The nucleotide sequence of IL-21 identified as SEQ ID NO: 28 was assembled from the partially homologous ("overlapping") sequences obtained from the deposited clone and a clone of genomic DNA. The nucleotide sequence of IL-22 identified as SEQ ID NO: 32 was assembled from partially homologous ("overlapping") sequences obtained from the deposited clones (ATCC Deposit No. 209665 and ATCC Deposit No. PTA- 70). The overlapping sequences specific for the IL-21 and IL-22 partial molecules of the invention and the full-length IL-21 molecule of the invention were each assembled into contiguous simple sequences of high redundancy (usually 3 to 5 sequences in overlap at each nucleotide position), resulting in four final sequences identified as SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 28 and SEQ ID NO: 31. Therefore, SEQ ID NO: 1 and SEQ ID NO: 2 translated; SEQ ID NO: 3 and SEQ ID NO: 4 translated; SEQ ID NO: 31 and SEQ ID NO: 32 translated; and SEQ ID NO: 28 and SEQ ID NO: 29 translated, are sufficiently accurate and otherwise suitable for a variety of uses well known in the art and described hereinafter. For example, SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 28 and SEQ ID NO: 31 are useful for designating nucleic acid hybridization probes that will detect nucleic acid sequences _.afc.a ¡.. .. +. - r - ..; _ «T¿blamsßk.A¿ + '? MJß. ... rifofea, ...; < contained in SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 28, and SEQ ID NO: 31, or the cDNA contained in the deposited cDNA clones, respectively. These probes will also hybridize to the nucleic acid molecules in biological samples, thereby making a variety of forensic and diagnostic methods of the invention possible. Similarly, polypeptides identified from SEQ ID NO; 2 and SEQ ID NO: 29 can be used to generate antibodies that bind specifically to IL-21 and polypeptides identified from SEQ ID NOr 4 and SEQ ID NO: 32, can be used to generate antibodies that bind specifically to IL-22. However, DNA sequences generated by sequencing reactions may contain sequencing errors. Errors exist as misidentified nucleotides, or as insertions or deletions of nucleotides in the generated DNA sequence. Erroneously inserted or deleted nucleotides cause structural shifts in the reading structures of the predicted amino acid sequence. In these cases, the predicted amino acid sequence! diverges from the effective amino acid sequence, even though the generated DNA sequence may be more than 99.9% identical to the effective DNA sequence (eg, a base insertion or deletion in an open reading structure of more than 1000 bases) . Accordingly, for those applications that require precision in nucleotide sequence or amino acid sequence, the present invention provides not only the generated nucleotide sequence, identified as SEQ ID NO: 1 and the predicted, translated amino acid sequence identified as SEQ ID. NO: 2, and the generated nucleotide sequence identified as SEQ ID NO: 28 and the predicted translated amino acid sequence, identified as SEQ ID NO: 29, but also a plasmid DNA sample containing a human IL-21 cDNA deposited with the ATCC. In addition, the present invention also provides not only the generated nucleotide sequence identified as SEQ ID NO: 3 and the predicted, translated amino acid sequence identified as SEQ ID NO: 4, and the generated nucleotide sequence identified as SEQ ID NO: 3, and the predicted amino acid sequence, predicted, identified as SEQ ID NO: 4, but also a sample of plasmid DNA containing a human cDNA of IL-22, deposited with the ATCC. Accordingly, the nucleotide sequence of the deposited IL-21 and IL-22 clones can be easily determined by sequencing the deposited clone according to known methods. The predicted amino acid sequences of IL-21 and IL-22 can then be verified from such deposits. In addition, the amino acid sequence of the protein encoded by the deposited clone can also be directly determined by peptide sequencing or by expression of the protein in a suitable host cell containing the deposited human IL-21 or IL-22 cDNAs, protein harvesting, and sequence determination. The present invention also relates to the IL-21 gene corresponding to SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 28, SEQ ID NO: 29 or the deposited clone encoding an IL -21 partial. The present invention also relates to the IL-22 gene corresponding to SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 31, SEQ ID NO: 32 or the deposited clone encoding IL-22. The genes for IL-21 and IL-22 can be isolated according to known methods using sequential information described herein. Such methods include ^ ¡S ^^^^ i ^^^^^^^^^^^^^? ^^^^^^^^^^^^^^^^^^^^^^^^ ^^^^^^^^^^^ ^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^^ preparation of probes or primers from the described sequences and identification or amplification of the 22 from appropriate sources of genomic material. Also homologous to IL-21 and IL-22 are provided in the present invention. The homologous species can be isolated and identified by making suitable probes or primers from the sequences provided herein, and selecting a suitable source of nucleic acid for the desired homologue. The IL-21 and IL-22 polypeptides can be prepared in any appropriate manner. Such polypeptides include isolated polypeptides of natural origin, recombinantly produced polypeptides, synthetically produced polypeptides, or polypeptides produced by a combination of these methods. Means for preparing such polypeptides are well understood in the art. The IL-21 and IL-22 polypeptides may be in the form of the secreted protein, including the mature form, or they may be a part of a larger protein, such as a fusion protein. It is often advantageous to include amino acids *? * ^ - S & sfa ^ a. ^ "> - A "extra" comprising secretory sequences or guides, pro-sequences, sequences that aid in purification, such as multiple histidine residues, or an additional sequence for stability during recombinant production. The IL-21 and IL-22 polypeptides are preferably provided in an isolated form, and are preferably substantially purified. A recombinantly produced version of an IL-21 or IL-22 polypeptide, including the secreted polypeptide, can be substantially purified by the one-step method described in the publication by Smith and Johnson (Gene 67: 31-40). (1988)). The IL-21 and IL-22 polypeptides can also be purified from natural or recombinant sources using antibodies of the invention produced against IL-21 and IL-22 proteins, respectively, in methods that are well known in the art.

Variants of Polynucleotides of Polypeptides "Variant" refers to a polynucleotide or polypeptide that differs from the polynucleotides or polypeptides IL-21 and IL-22, but which retains essential properties thereof. In general, the variants are in general closely similar, and in many regions, identical to the polynucleotide or polypeptide IL-21 and IL-22. For a polynucleotide having a nucleotide sequence at least, for example, 95? "identical" to a reference nucleotide sequence of the present invention, it is intended that the nucleotide sequence of the polynucleotide be identical to the reference sequence, except that the polynucleotide sequence may include up to five point mutations per 100 nucleotides of the nucleotide sequence of reference coding for IL-21 or IL-22 polypeptides. In other words, to obtain a polynucleotide having a nucleotide sequence at least 95% identical to a reference nucleotide sequence, up to 5% of the nucleotides in the reference sequence can be inserted, deleted or substituted with another nucleotide. The search sequence can be a complete sequence shown in SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 28, SEQ ID NO: 31, the ORF (open reading structure) of IL-21 or IL-22, or any fragment specified as described herein. As a matter, practical, if any particular molecule of nucleic acid or polypeptide is at least 90%95%, 96%, 97%, 98% or 99% identical to (or 10%, 5%, 4%, 3%, 2% or 1% different from) a nucleotide sequence of the present invention, this may be determined conventionally using known computer programs. A preferred method for determining the best complete coupling between an inquiry sequence (a sequence of the present invention) and an objective sequence, also referred to as a global sequential alignment, can be determined using the FASTDB computer program based on the algorithm of Brutlag and colleagues (Comp. App. Bi osci. 6: 237-245 (1990)). In sequential alignment, the search and target sequences are both DNA sequences. The RNA sequence can be compared by converting uridine residues (U) to thymidine residues (T). The result of global sequential alignment is a percentage identity. The preferred parameters used in a FASTDB alignment of the DNA sequences to calculate the percent identity: Unitary Matrix, k-tuple = 4, Punishment for Malfunction = 1, Punishment for Union = 30, Length of Randomization Group = 0, Qualification of Cut = 1, Punishment for Empty Space = 5, Punishment for Space Size = 0.05, Window Size = 500 or the length of the target nucleotide sequence, whichever is shorter. If the target sequence is shorter than the inquiry sequence due to 5 'or 3' deletions, but not due to internal deletions, a manual correction to the results must be made. This is because the FASTDB algorithm does not explain the 5 'and 3' truncations of the target sequence when the percent identity is calculated. For target sequences truncated at the 5 'or 3' ends, relative to the search sequence, the percentage identity is corrected by calculating the number of bases of the search sequence that are 5 'and 3' of the target sequence, not are paired / aligned, as a percentage of the total bases of the inquiry sequence. If a nucleotide is paired / aligned this is determined with the results of the sequential FASTDB alignment. This percentage is then subtracted from the percentage identity, calculated by the previous FASTDB program using the specified parameters, to arrive at a final percentage identity score. This corrected rating is that which is used for the purposes of the present invention. Only the bases outside the 5 'and 3' bases of the target sequence, as shown by the FASTDB alignment, which are not paired / aligned with the inquiry sequence, are calculated for the purpose of manually adjusting the percentage identity rating. . For example, a target sequence of 90 bases is aligned to a 100-base search sequence to determine percent identity. Deletions occur at the 5 'end of the target sequence and therefore, the FASTDB alignment does not show a matching / alignment of the first 10 bases at the 5' end. The 10 unpaired bases represent 10% of the sequence ((number of bases at the 5 'and 3' unpaired ends) / (total number of bases in the inquiry sequence)), so that 10% is subtracted from the percentage identity grade calculated using the FASTDB program. If the remaining 90 bases were perfectly matched, the final percentage identity would be 90%. In another example, a target sequence of 90 bases is compared to a 100-base search sequence. This time the deletions are internal deletions so that there are no bases on the 5 'or 3' ends of the target sequence, which are not paired / aligned with the inquiry. In this case, the percentage identity calculated using FASTDB is not manually corrected. Again, only the 5 'and 3' bases of the target sequence that are not matched / aligned with the inquiry sequence are manually corrected. No other manual corrections will be made for the purposes of the present invention. For a polypeptide having an amino acid sequence that is at least, for example, 95% "identical" to (or 5% different from) an amino acid sequence of the present invention, it is intended that the amino acid sequence of the The target polypeptide is identical to the screening sequence, except that the target polypeptide sequence may include up to five amino acid alterations per 100 amino acids of the query amino acid sequence. In other words, to obtain a polypeptide having an amino acid sequence at least 95% identical to an amino acid sequence of inquiry, up to 5% of the amino acid residues in the target sequence can be inserted, deleted (the insertions and deletions are collectively referred to as "indels" in the art) or substituted with another amino acid. These alterations of the reference sequence can occur at the amino- or carboxyl-terminal positions of the reference amino acid sequence or in gold site between those terminal positions, interspersed either individually between the residues in the reference sequence or in one or more contiguous groups within the reference sequence. As a practical matter, if any particular polypeptide is at least 90%, 95%, 96%, 97%, 98% or 99% identical to (or 10%, 5%, 4%, 3%, 2% or 1% different from), for example, the amino acid sequence shown in SEQ ID NO: 2 or SEQ ID NO: 29, or that shown in SEQ ID NO: 4 or SEQ ID NO: 32, or to the encoded amino acid sequence by deposited cDNA clones, you can be determined conventionally using known computer programs. A preferred method for determining the best fit or complete match between an inquiry sequence (a sequence of the present invention) and an objective sequence, also referred to as a global sequence alignment, can be determined using the FASTDB computer program based on the algorithm of Brutla? ry colleagues (Comp.App.Bi osci. 6: 237-245 (1990)). In a sequential alignment, the sequences of. Inquiry and objective are either both nucleotide sequences or both amino acid sequences. The result of the global sequential alignment is in percentage identity. The preferred parameters used in the alignment of FASTDEi amino acids are: Matrix = PAM 0, k-tuple = 2, Punishment for Malfunction = 1, Punishment for Union = 20, Length of Randomization Group = 0, Cutting Rating = 1 , Window Size = length of sequence, Punishment for Empty Space = 5, Punishment for Empty Space Size = 0.05, Window Size = 500 or the length of the target amino acid sequence, whichever is shorter.

If the target sequence is shorter than the inquiry sequence due to N- or C-terminal deletions, not due to internal deletions, one should be performed. Manual correction to the results. This is because the FASTDB program does not explain the N- and C-terminal truncations of the target sequence when calculating the overall percent identity. For target sequences truncated at the N and C ends in relation to the search sequence, the percentage identity is corrected by calculating the number of residues in the search sequence that are N- and C-terminal of the target sequence, which are not paired / aligned with a corresponding target residue, as a percentage of the total bases of the inquiry sequence. If a residue is paired / aligned this is determined by the alignment results of the FASTDB sequence. This percentage is then subtracted from the percentage identity, calculated by the aforementioned FASTDB program, using the specified parameters, to arrive at a final qualification of the percentage identity. This final qualification of percentage identity is that which is used for the purposes of the present invention. Only the residuals towards the N and C ends of the target sequence, which are not paired / aligned with the inquiry sequence, are considered for the purpose of manually adjusting the percentage identity rating. That is, only the positions of the residues outside the N- and C-terminal residues furthest from the target sequence. For example, a target sequence of 90 amino acid residues is aligned with a 100-residue search sequence to determine percent identity. Deletion occurs at the N-terminus of the target sequence and therefore, the FASTDB alignment does not show an alignment / alignment of the first 10 residues at the N-terminus. The 10 unpaired residues represent 10% of the sequence (number of residues at the N and C ends not the matched / total number of residues in the inquiry sequence), so that 10% is subtracted from the percentage identity rating calculated by the FASTDB program. If the remaining 90 residues were perfectly matched, the final percentage identity would be 90%. In yet another example, an objective sequence of 90 residues is compared with j * ^ a 100-residue search sequence. This time the deletions are internal deletions, so that there are no residues at the N or C ends of the target sequence, which are not paired / aligned with the inquiry. In this case, the percentage identity calculated using FASTDB is not manually corrected. Again, only the residue positions outside the terminal ends N and C of the target sequence, as shown in the FASTDB alignment, which are not paired / aligned with the inquiry sequence, are manually corrected. No other manual corrections will be made for the purposes of the present invention. The IL-21 and IL-22 variants may contain alterations in the coding regions, the non-coding regions, or both. Especially preferred are polynucleotide variants that contain alterations that produce substitutions, additions, or deletions, which do not alter the properties or activities of the encoded polypeptide. The nucleotide variants produced by silent substitutions due to the degeneracy of the genetic code are preferred. In addition, the variants in which ^ "^ .A amino acids 5-10, 1-5 or 1-2 are substituted, deleted, or aggregated in any combination, they are also preferred. The polynucleotide variants of IL-21 and IL-22 can be produced for a variety of reasons, for example, to optimize codon expression for a particular host (changing codons in human mRNA to all those preferred by a bacterial host such as E. col i). The variants of IL-21 and IL-22 of natural origin are called "allelic variants", and refer to one of several alternative forms of a gene that occupies a given locus on a chromosome of an organism (Genes II, Lewin, B., ed., John Wiley &Sons, New York (1985)). These allelic variants may vary either at the polinucletidic or polypeptide level. Alternatively, variants of non-natural origin can be produced by mutagenesis techniques or by direct synthesis. Using known methods of protein engineering and recombinant DNA technology, variants can be generated to improve or alter the characteristics of the IL-21 and IL-22 polypeptides. For example, one or more amino acids can be deleted from the N or C end of the protein When secreted, without substantial loss of biological function, Ron and coworkers reported variant KGF proteins that had heparin binding activity even after deleting amino acid residues 3, 8, or 27 amino acids. terminals (J. Biol. Chem. 268: 2984-2988 (1993)) Similarly, gamma interferon showed up to ten times greater activity after deletion of 8 to 10 carboxyl-terminal residues of this protein (Dobeli, et al. , J. Bi ot echnol.1: 199-216 (1988)) In the present case, since the proteins: IL-21 and IL-22 of the invention are highly related to the family of polypeptides similar to Interleukin- 17, deletions of N-terminal amino acids up to cysteine at position 19 of SEQ ID NO: 2 and up to cysteine at position 29 of SEQ ID NO: 4 may retain some biological activity.Polypeptides having deletions Additional N-terminals including the cyste residue na 19 in SEQ ID NO: 2 and the cysteine residue-29 in SEQ ID NO: 4 may not be expected to retain such biological activities because it is likely that these residues are required for forming a disulfide bridge to provide - * & a .3 & 8É & - afe. structural stability that is necessary for receptor binding and signal transduction. However, even if a deletion of one or more amino acids from the N-terminus of a protein results in modification or loss of one or more biological functions of the protein, other biological activities may still be conserved. Thus, the ability of the shortened protein to induce and / or bind to antibodies that recognize complete or mature IL-21 or IL-22 proteins will generally be conserved at least from most of the protein residues. Complete or mature IL-21 or IL-22 are removed from the N-termini of the respective proteins. If a particular polypeptide lacking the N-terminal residues of a complete protein retains such immunological activities, this can be easily determined by routine methods described herein and otherwise known in the art. Accordingly, the present invention further provides polypeptides having one or more residues deleted from the amino terminus of the amino acid sequence of the IL-21 polypeptide shown in SEQ ID NO: 2, to the cysteine residue at position number 19 , and the polynucleotides that code for such polypeptides. In addition, the present invention provides additional polypeptides having one or more residues deleted from the amino terminus of the amino acid sequence of the IL-22 polypeptide shown in SEQ ID NO: 4, up to the cysteine residue at position number 29, and the polynucleotides encoding such polypeptides. In particular, the present invention provides polypeptides comprising the amino acid sequence of residues n1-87 of SEQ ID NO: 2, where n1 is an integer in the range of 1 to 18, and 19 is the position of the first residue from the N-terminus of the complete IL-21 polypeptide (shown in SEQ ID NO: 2) that is believed to be required for binding activity to the IL-21 protein receptor. Likewise, the present invention provides the polypeptides comprising the amino acid sequence of residues n2-160 of SEQ ID NO: 4, where n2 is an integer in the range of 1 to 28, and 29 is the position of the first residue from the N-terminus of the complete IL-22 polypeptide (shown in SEQ ID -4 &ií * JÍ? É £ i i? Xií as & »s, ^!» Ik - «a b y - > .-al-. -üiBfc ».

NO: 4) is believed to be required for binding activity to the IL-22 protein receptor. More particularly, the invention provides the polynucleotides that encode the polypeptides comprising, or consisting alternatively of, the amino acid sequence of residues 1-87, 2-87, 3-87, 4-87, 5-87, 6-87, 7-87, 8-87, 9-87, 10-87, 11-87, 12-87, 13-87, 14-87, 15-87, 16-87, 17-87, 18- 87 and 19-87 of SEQ ID NO: 2. The polypeptides encoded by these polynucleotides are also provided. The present application is also directed to nucleic acid molecules comprising, or alternatively consisting of, a polynucleotide sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to the polynucleotide sequence encoding for the IL-21 polypeptides described above. The present invention also encompasses the above polynucleotide sequences fused to a heterologous polynucleotide sequence. The invention also provides polynucleotides that encode polypeptides comprising, or consisting alternatively of, the amino acid sequence of residues 1-160, 2-160, 3-160, 4-160, 5- Xffb, 6-160. , 7-160, 8-160, 9-160, 10-160, 11-160, 12-160, 13-160, 14-160, 15-160, 16-160, 17-160, 18-160, 19 -160, 20-160, 21-160, 22-160, 23-160, 24-160, 25-160, 26-160, 27-160, 28-160 and 29-160 of SEQ ID NO: 4. The polypeptides encoded by these polynucleotides are also provided. The present application is also directed to nucleic acid molecules comprising, or alternatively consisting of, a polynucleotide sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to the polynucleotide sequence encoding for the IL-21 polypeptides described above. The present invention also encompasses the above polynucleotide sequences fused to a heterologous polynucleotide sequence. In addition, since the IL-21 and IL-22 proteins of the invention are highly related to the family of IL-17-like polypeptides, deletions of C-terminal amino acids to leucine at position 83 of SEQ ID NO: 2 and up to the proline at position 129 of SEQ ID NO: 4, may retain some biological activity. Polypeptides having additional C-terminal deletions, including the leucine residue at position 83 of SEQ ID NO: 2 and the proline at position 129 of SEQ ID NO: 4, could not be expected to retain such biological activities since these residues are at the beginning of the conserved domain required for biological activities. However, even if the deletion of one or more amino acids from the C-terminus of a protein results in the modification of the loss giving one or more biological functions of the protein, other biological activities can still be conserved. Thus, the ability of the shortened protein to induce and / or bind to the antibodies that recognize the complete or mature IL-21 and IL-22 proteins will generally be conserved at least from most of the residues of the proteins. Complete or mature IL-21 and IL-22 proteins are removed from the C-terminus. If a particular polypeptide lacking the C-terminal residues of a complete protein retains such immunological activities or not, this can be easily determined by routine methods described in present and otherwise known in the art. Accordingly, the present invention further provides polypeptides having one or more residues removed from the carboxyl terminus of 1e? amino acid sequence of the IL-21 polypeptide shown in SEQ ID NO: 2, to the leucine residue at position 83 of SEQ ID NO: 2, and the polynucleotides encoding such polypeptides. In addition, the present invention further provides polypeptides having one or more residues removed from the carboxyl terminus of the amino acid sequence of the IL-22 polypeptide shown in SEQ ID NO: 4, up to the proline residues at position 129 of SEQ ID NO: 4. In particular, the present invention provides polypeptides having the amino acid sequence of residues 1-m1 of the amino acid sequence in SEQ ID NO: 2, wherein m1 is any integer in the range of 83 to 87, and residue 82 is at the position of the first residue from the C-terminus of the complete IL-21 polypeptide (shown in SEQ ID NO: 2) that is Cree is required for the activity of the IL-21 protein. In addition, the present invention also provides polypeptides having the amino acid sequence of residues 1-m2 of the amino acid sequence of SEQ ID NO: 4, where m2 is any integer in the range of 129 to 160, and residue 128 is the - < & position of the first residue from the C-terminus of the complete IL-22 polypeptide (shown in SEQ ID NO: 4) that is believed to be required for the activity of the IL-22 protein. More particularly, the invention provides polynucleotides that encode polypeptides comprising, or consisting alternatively of, the amino acid sequence of residues 1-83, 1-84, 1-85, 1-86 and 1-87 of SEQ ID NO: 2. The polypeptides encoded by these polynucleotides are also provided. The present application is also directed to nucleic acid molecules comprising, or alternatively consisting of, a polynucleotide sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to the polynucleotide sequence encoding the IL-21 polypeptides described above. The present invention also encompasses the above polynucleotide sequences fused to a heterologous polynucleotide sequence. The present invention also provides the polynucleotides that encode the polypeptides comprising, or consisting alternatively of, the amino acid sequence of waste 1-129, 1-130, 1-131, 1-132, 1-133, 1- "rifeaA i» * «.« «», _ «», =. & ,; -M. ».« < * «Mf '.? aBBfaa. ,, <. < feJ > ..» «• * # 134, 1-135, 1-136, 1-137., 1-138, 1-139, 1-140, 1- 141, 1-142, 1-143, 1-144, 1-145, 1-146 , 1-147, 1- 148, 1-149, 1-150, 1-151, 1-152, 1-153, 1-154, 1-155, 1-156, 1-157, 1-158, 1 -159 and 1-160 of SEQ ID NO: 4. The polypeptides encoded by these polynucleotides are also provided. The present application is also directed to nucleic acid molecules comprising, or alternatively consisting of, a polynucleotide sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to the polynucleotide sequence encoding for the IL-22 polypeptides described above. The present invention also encompasses the above polynucleotide sequences fused to a heterologous polynucleotide sequence. The present invention also provides polypeptides having one or more amino acids deleted from the amino and carboxyl termini of IL-21, which can be described in general as having residues n ^ 1 of SEQ ID NO: 2, where n1 and m1 are integers as described above. Similarly, the invention also provides polypeptides having one or more amino acids deleted from the amino termini and carboxyl of IL-22, which can be described in general as having the n2-m2 residues of SEQ ID NO: 4, where n2 and 2 are integers as described above. 5 In addition, extensive evidence shows that variants frequently retain a biological activity similar to that of the protein of natural origin. For example, Gayle and colleagues conducted extensive mutational analyzes of the human cytokine IL-la (J. Bi ol. Chem. 268: 22105-22111 (1993)). They used random mutagenesis to generate over 3,500 individual IL-la mutants that averaged 2.5 amino acid changes per variant over full length of the molecule. Multiple mutations were examined at each possible amino acid position. The researchers found that "[most of the molecules could be altered with little effect on either the link or the biological activity]" (see, extract). In fact, only 23 unique amino acid sequences from more than 3,500 nucleotide sequences examined yielded a protein that differed significantly in activity from the wild type.

Furthermore, even if one or more amino acids are deleted from the N-terminus or the C-terminus of a polypeptide, it results in the modification or loss of one or more biological functions, and other biological activities can still be conserved. For example, the ability of a deletion variant to induce and / or bind antibodies that recognize the secreted form, will probably be retained at least you? most of the residues of the secretadat form that are removed from the N-terminus or the C-terminus. If a particular polypeptide lacking the N- and C-terminal residues of a protein preserves such biological activities or not, this can be easily determined by routine methods described herein and otherwise known in the art. As mentioned above, even if the deletion of one or more amino acids from the N-terminus of a protein results in the modification of the loss of one or more biological functions of the protein, other biological activities can still be conserved. In this way, the ability of the shortened protein to induce and / or bind to the antibodies that recognize full or mature IL-21 or IL-22 proteins, it will be generally conserved, at least that the majority of the residues of the complete or mature IL-21 or IL-22 proteins that are removed, from the N termini of the respective proteins. If a particular polypeptide lacking the N-terminal residues of a complete protein, whether or not it retains such immunological activities, this can be easily determined by routine methods described herein and otherwise known in the art. Accordingly, the present invention further provides polypeptides having one or more residues deleted from the amino terminus of the amino acid sequence of the IL-21 polypeptide shown in SEQ ID NO: 2, up to the valine residue at position number 82, and the polynucleotides encoding such polypeptides. In addition, the present invention further provides polypeptides having one or more residues deleted from the amino terminus of the amino acid sequence of IL-22 polypeptide shown in SEQ ID NO: 4, up to the aspartic acid residue at position number 155, and the polynucleotides encoding such polypeptides. In particular, the present invention provides the polypeptides comprising the amino acid sequence of residues n3-87 of SEQ ID NO: 2, where n3 is an integer in the range of 1 to 82, and 83 is the position of the first residue from the N-terminus of the complete IL-21 polypeptide (shown in SEQ ID NO: 2) that is believed to be required for the immunogenic activity of the IL-21 protein. Likewise, the present invention provides the polypeptides comprising the amino acid sequence of residues n -160 of SEQ ID NO: 4, where n4 is an integer in the range of 1 to 155, and 156 is the position of the first residue from the N-terminus of the complete IL-22 polypeptide (shown in SEQ ID NO: 4) that is believed to be required for the immunogenic activity of the IL-22 protein. More particularly, the invention provides polynucleotides that encode polypeptides comprising, or consisting alternatively of, the amino acid sequence at residues R-2 to V-87; V-3 A V-87; D-4 to V-87; T-5 to V-87; D-6 to V-87; E-7 to V-87; D-8 to V-87; R-9 to V-87; Y- 10 to V-87; P-ll to V-87; Q-12 to V-87; K-13 to V-87; L-14 to V-87; A- 15 to V- 87; F-16 to V-87; A- 17 to V- 87 E-18 to V-87 C-19 to V-87 L-20 to V-87 C-21 to V-87 R-22 to V-87 G-23 to V-87 C -24 to V-87 1-25 to V-87 D-26 to V-87 A- 27 to V-87 R-28 to V-87 T-29 A V-87 G-30 A V-87 R- 31 to V-87 E-32 to V-87 T-33 to V-87 A- 34 to V-87 A-35 to V-87 L-36 A V-87 N-37 to V-87 S-38 to V-87 V-39 to V-87 R-40 to V-87 L-41 to V-87 L-42 to V-87 Q-43 to V-87 S-44 to V-87 L-45 to V-87 L-46 to V-87 V-47 to V-87 L-48 to V-87 R-49 A V-87 R-50 to V-87 R-51 to V-87 P-52 to V -87 C-53 to V-87 S-54 to V-87 R-55 to V-87 D-56 to V-87 G-57 to V-87 S-58 to V-87 G-59 to V- 87 L-60 A V-87 P-61 to V-87 T-62 A V-87 P-63 A V-87 G-64 to V-87 A-65 to V-87 F-66 to V-87 A-67 A V-87 F-68 A V-87 H-69 to V-87 T-70 to V-87 E-71 to V-87 F-72 to V-87 1-73 to V-87 H -74 to V-87 V-75 to V-87 P-76 to V-87 V-77 A V-87 G-87 to V-87 C-79 to V-87 T-80 to V-87 C- 81 to V-87 and V-82 to V-87 of SEQ ID NO 2. The polypeptides encoded by these polynucleotides are also provided. The present application is also directed to nucleic acid molecules comprising, or alternatively consisting of, a polynucleotide sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to the polynucleotide sequence encoding for the IL-21 polypeptides described above. The present invention also encompasses the above polynucleotide sequences fused to a heterologous polynucleotide sequence. In addition, the invention provides the polynucleotides that encode the polypeptides comprising, or consisting alternatively of, the amino acid sequence of residues S-2 to P-160; A- 3 to P-160; R-4 to P-160; A- 5 to P-160; R- 6 to P-160; A- 7 to P-160; V-8 to P-160; L-9 to P-160; S-10 to P-160; A-11 to P-160; F-12 to P-160; H-13 to P-160; H-14 to P-160; T-15 to P-160; L-16 to P-160; Q-17 to P-160; L-18 to P-160; G-19 to P-160; P-20 to P-160; R-21 to P-160; E-22 to P-160; Q-23 to P-160; A-24 to P-160; R-25 to P-160; N-26 to P-160; A-27 to P-160; S-28 to P-160; C-29 to P-160; P-30 to P-160; A-31 to P-160; G-32 to P-160; G-33 to P-160; R-34 to P-160; P-35 to P-160; A- 36 to P-160; D-37 to P-160; R-38 to P-160; R-39 to P-160; F-40 to P-160; R-41 to P-160; P-42 to P-160; P-43 to P-160; T-44 to P-160; N-45 to P-160; L-46 to P-160; R-47 to P-160; S-48 to P-160; V-49 to P-160; S-50 to P-160; P-51 to P-160; W-52 to P-160; A-53 to P-160; Y-54 to P-160; R-55 to P-160; 1-56 to P-160; S-57 to P-160; Y-58 to P-160; D-59 to P-160; P-60 to P-160; A-61 to P-160; R-62 to P-160; Y-63 to P-160; P-64 to P-160; R-65 to P-160; Y-66 to P-160; L-67 to P-160; P-68 to P-160; E-69 to P-160; A-70 to P-160; Y-71 to P-160; C-72 to P-160; L-73 to P-160; C-74 to P-160; R-75 to P-160; G-76 to P-160; C-77 to P-160; L-78 to P-160; T-79 to P-160; G-80 to P-160; L-81 to P-160; F-82 to P-160; G-83 to P-160; E-84 to P-160; E-85 to P-160; D-86 to P-160; V-87 to P-160; R-88 to P-160; F-89 to P-160; R-90 to P-160: S-91 to P-160; A-92 to P-160; P-93 to P-160; V-94 to P-160; Y-95 to P-160; M-96 to P-160; P-97 to P-160; T-93 to P-160; V-99 to P-160; V-100 to P-160; L-101 to P-160 R-102 to P-160; R-103 to P-160; T-104 to P-160; P-105 to P-160; A-106 to P-160; C-107 to P-160; A-108 to P-160 G-109 to P-160; G-110 to P-160; R-111 to P-160; S-112 to P-160; V-113 to P-160; Y-114 to P-160; T-115 to P-160 / E-116 to P-160; A-117 to P-160; Y-118 to P-160; V-119 to P-160; T-120 to P-160; 1-121 to P-160; P-122 to P-160 / V-123 to P-160; G-124 to P-160; C-125 to P-160; T-126 to P-160; C-127 to P-160; V-128 to P-160; P-129 to P-160; E-130 to P-160; P-131 to P-160; E-132 to P-160; K-133 to P-160; D-134 to P-160; A-135 to P-160; D-136 to P-160; S-137 to P-160; 1-138 to P-160; N-139 to P-160; S-140 to P-160; S-141 to P-160; 1-142 to P-160; D-143 to P-160; K-144 to P-160; Q-145 to P-160; G-146 to P-160; A-147 to P-160; K-148 to P-160; L-149 to P-160; L-150 to P-160; L-151 to P-160; G-152 to P-160; P-153 to P-160; N-154 to P-160; and D-155 to P-160 of SEQ ID NO: 4. they also provide the polypeptides encoded by these polynucleotides. The present application is also directed to nucleic acid molecules comprising, or alternatively consisting of, a polynucleotide sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to the polynucleotide sequence encoding for the IL-22 polypeptides described above. The present invention also encompasses the above polynucleotide sequences fused to a heterologous polynucleotide sequence. Also as mentioned above, even if the deletion of one or more amino acids from the C-terminus of a protein results in the modification of the loss of one or more biological functions of the protein, other biological activities may still be conserved. Thus, the ability of the shortened protein to induce and / or bind antibodies that recognize complete or mature IL-21 and IL-22 proteins will generally be conserved at least from most protein residues. Complete or mature IL-21 and IL-22 are removed from the C-terminus. If a particular polypeptide lacking the C-terminal residues of a complete protein retains such immunological activities or not, this can be easily determined by routine methods described in FIG. present and otherwise known in the art. Accordingly, the present invention further provides polypeptides having one or more residues removed from the carboxyl terminus of the amino acid sequence of the IL-21 polypeptide shown in SEQ ID NO: 2, up to the residue of aspartic acid in position 6 of SEQ ID NO: 2, and the polynucleotides encoding such polypeptides. In addition, the present invention further provides polypeptides having one or more residues removed from the carboxyl terminus of the amino acid sequence of the IL-22 polypeptide shown in SEQ ID NO: 4, up to the arginine residues at position 6 of SEQ ID NO: 4. In particular, the present invention provides polypeptides having the amino acid sequence of the 1-m3 residues of the amino acid sequence in SEQ ID NO: 2, where m3 is any integer in the range of 6 to 87, and residue 5 is the position of the first residue from the C-terminus of the polypeptide Complete IL-21 (shown in the SEQ ID NO: 2) that is believed to be required for the activity Immunogenicity of the IL-21 protein. In addition, the present invention also provides polypeptides having the amino acid sequence of residues 1-m4 in SEQ ID NO: 4, where m4 is any integer in the range of 6 to 160, and residue 5 is in the position of the first residue from the C-terminus of the complete IL-22 polypeptide (shown in SEQ ID NO: 4) that is believed to be required for the immunogenic activity of the IL-22 protein. More particularly, the invention provides polynucleotides that encode polypeptides that comprise or consist alternatively of the amino acid sequence of residues A-1 through S-86.; A-1 to R-85; A-1 to P-84; A-1 to L-83; A-1 to V-82; A-1 to C-81; A-l to T-80; A-1 to C-79; A-1 to G-78; A-1 to V-77; A-1 to P-76; A-1 to V-75; A-1 to H-74; A-1 to 1-73; A-1 to F-72; A-1 to E-71; A-1 to T-70; A-1 to H-69; A-1 to F-68; A-1 to A-67; A-1 to F-66; A-1 to A-65; A-1 to G-64; A-1 to P-63; A-1 to T-62; A-1 to P-61; A-1 to L-60; A-1 to G-59; A-1 to S-58; A-1 to G-57; A-1 to D-56; A-1 to R-55; A-1 to S-54; A-1 to C-53; A-1 to P-52; A-1 to R-51; A-1 to R-50; A-1 to R-49; A-1 to L-48; A-1 to V-47; A-1 to L-46; A-1 to L-45; A-1 to S-44; A-1 to Q-43; A-1 to L-42; A-1 to L-41; A-1 to R-40; A-1 to V-39; A-1 to S-38; A-1 to N-37; A-1 to L-36; A-1 to A-35; A-1 to A-34; A-1 to T-33; A-1 to E-32; A-1 to R-31; A-1 to G-30; A-1 to T-29; A-1 to R-28; A-1 to A-27; A-1 to D-26; A-1 to 1-25; A-1 to C-24; A-1 to G-23; A-1 to R-22; A-1 to C-21; A-1 to L-20; A-1 to C-19; A-1 to E-18; A-1 to A-17; A-1 to F-16; A-1 to A-15; A-1 to L-14; A-1 to K-13; A-1 to Q-12; A-1 to P-11; A-1 to Y-10; A-1 to R-9; A-1 to D-8; A-l to E-7; and A-1 to D-6 of SEQ ID NO: 2. Polynucleotides encoding these polypeptides are also provided. The present application is also directed to nucleic acid molecules comprising, or alternatively consisting of, a polynucleotide sequence of at least 90%, 95%, 96%, 97%, 98% or 99% identical to the polynucleotide sequence that encodes the IL-21 polypeptides described above. The present invention also encompasses the above polynucleotide sequences fused to a heterologous polynucleotide sequence. In addition, the invention provides the polynucleotides that encode the polypeptides comprising, or consisting alternatively of, the amino acid sequence of residues N-1 to G-159; N-1 to A-158; N-1 to P-157; N-1 to A-156; N-1 to D-155 N-1 to N-1 54; N-1 to N-1 to G-152; N-l to L ~ 151 N-1 to L-150; N-1 to N-1 to K-148; N-1 to A-147 N-1 to G-146; N-1 to Q-145 N-1 to K-144; N-1 to D-143 N-1 to 1-142; N-1 to S-141 N-1 to S-140; N-1 to N-139 N-1 to 1-138; N-1 to S-137 N-1 to D-136; N-1 to A-135 N-1 to D-134; N-1 to K-133 N-1 to E-132; N-1 to P-131 N-1 to E-130; N-1 to P-129 N-1 to V-128; N-1 to C-127 N-1 to T-126; N-1 to C-125 N-1 to G-124; N-1 to V-123 N-1 to P-122; N-1 to 1-121 N-1 to T-120; N-1 to V-119 N-1 to Y-118; N-1 to A-17 N-1 to E-116; N-1 to T-115 N-1 to Y-114; N-1 to V-113 N-1 to S-112; N-1 to R-111 N-1 to G-110; N-1 to G-109 N-1 to A-1 08; N-1 to C-107 N-1 to A-1 06; N-1 to P-105 N-1 to T-104; N-1 to R-103 N-1 to R-102; N-1 to L-101 N-1 to V-100; N-1 to V-99; N-1 to T-98; N-l to P -97; N-1 to M-96; N-1 to Y-95; N-1 to V-94; N-1 to P-93; N-1 to A-92; N-1 to S-91; N-1 to R 90; N-1 to F-89; N-1 to R-88; N-1 to V-87; N-1 to D-86; N-1 to E-85; N-1 to E -84; N-1 to G-83; N-1 to F-82; N-1 to L-81; N-1 to G-80; N-1 to T-79; N-1 to L-78; N-1 to C 77; N-1 to G-76; N-1 to R-75; N-1 to C-74; N-1 to L-73; N-1 to C-72; N-l to Y -71; N-1 to A-70; N-1 to E-69; N-1 to P-68; N-1 to L-67; N-1 to Y-66; N-1 to R-65; N-1 to P 64; N-1 to Y-63; N-1 to R-62; N-1 to A-61; N-1 to P-60; N-1 to D-59; N-1 to Y -58; N-1 to S-57; N-1 to 1-56; N-1 to R-55; N-1 to Y-54; N-1 to A-53; N-1 to W-52; N-l ^ s ^ ^ AB ^ Ma ^ í ^ Jfe. to P-51; N-1 to S-50; N-1 to V-49; N-1 to S-48; N-1 to R-47; N-1 to L-46; N-1 to N-45; N-l to T-44; N-l to P-43; N-1 to P-42; N-1 to R-41; N-1 to F-40; N-1 to R-39; N-1 to R-38; N-1 to D-37; N-1 to A-36; N-1 to P-35; N-1 to R-34; N-1 to G-33; N-1 to G-32; N-1 to A-31; N-1 to P-30; N-1 to C-29; N-1 to S-28; N-1 to A-27; N-1 to N-26; N-1 to R-25; N-1 to A-24; N-1 to Q-23; N-1 to E-22; N-1 to R-21; N-1 to P-20; N-1 to G-19; N-l 'to L-18; N-1 to Q-17; N-1 to L-16; N-1 to T-15; N-1 to H-14; N-1 to H-13; N-1 to F-12; N-l to A-ll; N-1 to S-10; N-1 to L-9; N-1 to V-8; N-1 to A-7; and N-1 to R-6 of SEQ ID NO: 4. Polypeptides encoded by these polynucleotides are also provided. The present application is also directed to nucleic acid molecules comprising, or alternatively consisting of, a polynucleotide sequence at least 90%, 95%, 96%, 97%, 98%, or 99% identical to the polynucleotide sequence encoding for the IL-22 polypeptides described above. The present invention also encompasses the above polynucleotide sequences, fused to a heterologous polynucleotide sequence. The invention also provides polypeptides having one or more amino acids deleted from the amino and carboxyl termini of the IL-21, which can be described in general as having the residues n3-m3 of SEQ ID NO: 2, where n3 and m3 are integers as described above. Similarly, the invention also provides polypeptides having one or more amino acids deleted from the amino and carboxyl termini of IL-22, which can be described in general as having residues n -m4 of SEQ ID NO: 4, where n4 and m4 are integers as described above. In addition, any polypeptide having one or more amino acids deleted from the amino and carboxyl termini of IL-22, specifically described as having the residues n4-m4 of SEQ ID NO: 4 (where n4 and m4 are integers as described above) may be excluded from the invention. In particular, any polypeptide having one or more amino acids deleted from the amino and carboxyl termini of IL-22 and which is defined by residues n4-m4 of SEQ ID NO: 4, where n4 is equal to 21, 22 , 23, 24 or 25 and m4 is equal to 271, 272, 273, 274, 275 or 276 may be excluded from the invention. Also, as mentioned above, even if the deletion of one or more amino acids from the N-terminus of a protein results in the modification or loss of one or more biological functions of the protein, other biological activities may still be conserved . Thus, the ability of the shortened protein to induce and / or bind antibodies that recognize full-length or mature IL-21 polypeptides, will be generally conserved when at least the majority of the residues of the full-length or mature IL-21 polypeptides are removed from the N-terminus. If a particular polypeptide lacks the N-terminal residues of a full or length polypeptide complete, preserve or not such immunological activities, this can be easily determined by routine methods described herein and otherwise known in the art. Accordingly, the present invention further provides polypeptides having one or more residues deleted from the amino terminus of the amino acid sequence of the IL-21 polypeptide shown in SEQ ID NO: 29, up to the valine residue at position number 192 , and the polynucleotides that code for such polypeptides. In particular, the present invention provides the polypeptides comprising the amino acid sequence of residues n5-197 of SEQ ID NO: 29, where n5 is an integer in the range of 1 to 192, and 193 is the position of the first residue from the N terminus of the IL-21 polypeptide in length complete (shown in SEQ ID NO: 29) that is believed to be required for the immunogenic activity of the IL-21 protein. More particularly, the invention provides the polynucleotides that encode the polypeptides comprising, or consisting alternatively of, the amino acid sequence of residues T-2 through V-197; L-3 to V-197; L-4 to V-197; P-5 to V-197; G-6 to V-197; L-7 to V-197; L-8 to V-197; F-9 to V-197; L-10 to V-197; T-ll to V-197; W-12 to V-197; L-13 to V-197; H-14 to V-197; T-15 to V-197; C-16 to V-197; L-17 to V-197; A-18 to V-197; H-19 to V-197; H-20 to V-197; D-21 to V-197; P-22 to V-197; S-23 to V-197; L-24 to V-197; R-25 to V-197; G-26 to V-197; H-27 to V-197; P-28 to V-197; H-29 to V-197; S-30 to V-197; H-31 to V-197; G-32 to V-197; T-33 to V-197; P-34 to V-197; H-35 to V-197; C-36 to V-197; Y- 37 to V-197; S-38 to V-197; A-39 to V-197; E-40 to V-197; E-41 to V-197; L-42 to V-197; P-43 to V-197; L-44 to V-197; G-45 to V-197; Q-46 to V-197; A-47 to V-197; P-48 to V-197; P-49 to V-197; H-50 to V-197; L-51 to V-197; L-52 to V-197; , -fi A-53 to V-197; R-54 to V-197; G-55 to V-197; A-56 to V- 197; K-57 to V-197; W-58 to V-197; G-59 to V-197; Q-60 to V-197; A-61 to V-197; L-62 to V-197; P-63 to V-197; V-64 to V-197; A-65 to V-197; L-66 to V-197; V-67 to V- 197; S-68 to V-197; S-69 to V-197; L-70 to V-197; E-71 to V-197; A-72 to V-197; A-73 to V-197; S-74 to V-197; H-75 to V-197; R-76 to V-197; G-77 to V-197; R-78 to V-197; H-79 to V-197; E-80 to V-197; R-81 to V-197; P-82 to V-197; S-83 to V-197; A-84 to V-197; T-85 to V-197; T-86 to V-197; Q-87 to V-197; C-88 to V-197; P-89 to V- 197; V-90 to V-197; L-91 to V-197; R-92 to V-197; P-93 to V-197; E-94 to V-197; E-95 to V-197; V-96 to V-197; L-97 to V-197; E-98 to V-197; A-99 to V-197; D-100 to V-197; T-101 to V-197; H-102 to V-197; Q-103 to V-197; R-104 to V-197; S-105 to V-197; 1-106 to V-197; S-107 a V-197; P-108 to V-197; W-109 to V-197; R-110 to V-197; Y-III to V-197; R-112 to V-197; V-113 to V-197; D-114 to V-197; T-115 to V-197; D-116 to V-197; E-117 to V-197; D-118 to V-197; R-119 to V-197; Y-120 to V-197; P-121 to V-197; Q-122 to V-197; K-123 to V-197; L-124 to V-197; A-125 to V-197; F-126 to V-197; A-127 to V-197; E-128 to V-197; C-129 to V-197; L-130 to V-197; C-131 to V-197; R-132 to V-197; G-133 to V-197; C-134 to V-197; 1-135 to V-197; D-136 to V-197; A-137 to V-197; R-138 to V-197; T-139 to V-197; G-140 to V-197; R-141 to V-197; E-142 to V-197; T-143 to V-197; A-144 to V-197; A-145 to V-197; L-146 to V-197; N-l 47 to V-197; S-148 to V-197; V-149 to V-197; R-150 to V-197; L-151 to V-197; L-152 to V-197; Q-153 to V-197; S-154 to V-197; L-155 to V-197; L-156 to V-197; V-157 to V-197; L-158 to V-197; R-159 to V-197; R-160 to V-197; R-161 to V-197; P-162 to V-197; C-163 to V-197; S-164 to V-197; R-165 to V-197; D-166 to V-197; G-167 to V-197; S-168 to V-197; G-169 to V-197; L-170 to V-197; P-171 to V-197; T-172 to V-197; P-173 to V-197; G-174 to V-197; A-175 to V-197; F-176 to V-197; A-177 to V-197; F-178 to V-197; H-179 to V-197; T-180 to V-197; E-181 to V-197; F-182 to V-197; 1-183 to V-197; H-184 to V-197; V-185 to V-197; P-186 to V-197; V-187 to V-197; G-188 to V-197; C-189 to V-197; T-190 to V-197; C-191 to V-197; and V-192 to V-197; of SEQ ID NO: 29. Polypeptides encoded by these polynucleotides are also provided. The present application is also directed to nucleic acid molecules comprising, or alternatively consisting of, a polynucleotide sequence at least 90%, 95%, 96%, 97%, 98%, or 99% identical to the polynucleotide sequence encoding for the IL-21 polypeptides described above. The present invention also encompasses the above polynucleotide sequences, fused to a heterologous polynucleotide sequence. Also, as mentioned above, even if the deletion of one or more amino acids from the C-terminus of a protein results in the modification or loss of one or more biological functions of the protein, other biological activities can still be conserved. Thus, the ability of the shortened protein to induce and / or bind antibodies that recognize full-length or mature IL-21 polypeptides will in general be conserved at least from most of the IL-1 polypeptide residues. 21 full-length or mature are removed from the C-terminus. If a particular polypeptide lacking the C-terminal residues of a full-length or full-length polypeptide, retains or not such immunological activities, this can be easily determined by routine methods described in present and otherwise known in the art. Accordingly, the present invention further provides the polypeptides having one or more residues removed from the carboxyl terminus of the amino acid sequence of the IL-21 polypeptide shown in SEQ ID NO: 29, to the glycine residue in the 6-position of the SEQ ID NO: 29, and the polynucleotides that code for such polypeptides. In particular, the present invention provides polypeptides having the amino acid sequence of residues 1-m5 of the amino acid sequence in SEQ ID NO: 29, where m5 is any integer in the range of 6 to 196, and residue 5 is the position of the first residue from the C-terminus of the full-length IL-21 polypeptide (shown in SEQ ID NO: 29) that is believed to be required for the immunogenic activity of the IL-21 protein. More particularly, the invention provides The polynucleotides that encode the polypeptides comprising, or consisting alternatively of, the amino acid sequence of residues M-1 through S-196; M-1 to R-195; M-1 to P-194; M-1 to L-193; M-1 to V-192; M-1 to C-191; M-l to T-190; M-1 to C-189; M-1 to G-20 188; M-1 to V-187; M-1 to P-186; M-1 to V-185; M-1 to H-184; M-1 to 1-183; M-1 to F-182; M-1 to E-181; M-1 to T-180; M-1 to H-179; M-1 to F-178; M-1 to A-1 11; M-1 to F-176; M-1 to A-175; M-1 to G-174; M-1 to P-173; M-l to T-172; M-1 to P-171; M-1 to L-170; M-1 to G-169; M-1 to S-25 168; M-1 to G-167; M-1 to D-166; M-1 to R-165; M-l to S- -ateo "o-» >, "- -" ** £ Vi .. _. V- »-: .- '&. <. • - U ^ Sb * ¿¿- .. .. * 164 M- -1 to C- -163 M- -1 to P- -? S2 M- -1 to R- -161 M-l to R 160 M- -1 to R- -159 M- -1 to v - -158 M- -1 to V- -157 M-l to L 156 M- -1 to L- -155 M- -1 to S- -154 M- -1 to Q- -153 M-l to L 152 M- -1 to L- -151 M- -1 to R- -150 M- -1 to V- -149 M-l to S 148 M- -1 to N- -147 M- -1 to L- -146 M- -1 to A- -145 M-l to A 144 M- -1 to T- -143 M- -1 to E- -142 M- -1 to R- -141 M-l to G 140 M- -1 to T- -139 M- -1 to R- -138 M- -1 to A- -137 M-l to D 136 M- -1 to I- -135 M- -1 to C- -134 M- -1 to G- -133 M-l to R 132 M- -1 to C- -131 M- -1 to L- -130 M- -1 to C- -129 M-l to E 128 M- -1 to A- -127 M- -1 to F- -126 M- -1 to A- -125 M-l to L 124 M- • 1 to K- -123 M- -1 to Q- -122 M- -1 to P- -121 M-l to Y 120 M-1 to R- -119 M-1 to D- -118 M--1 to E- -117 M-l to D 116 M- • 1 to T- -115 M- -1 to D- -114 M- -1 to V- -113 M-l to R 112 M- -1 to Y- -111 M- • 1 to R- -110 M- -1 to W- -109 M-l to P 108 M- -1 to S- -107 M- -1 to I- -106 M- -1 to s- -105 M-l to R 104 M- • 1 to Q- -103 M-l to H-102 M- -1 to T- -101 M-l to D 100 M-l to A-99; M-l to E-98; M-1 to L-97; M-1 to V-96; M-l to E-95; M-l to E-94; M-1 to P-93; M-1 to R-92; M-l to L 91; M-l to V-90; M-1 to P-89; M-1 to C-88; M-1 to Q-87; M-1 to T-86; M-l to T-85; M-1 to A-84; M-1 to S-83; M-1 to P-82; M-1 to R-81; M-l to E-80; M-1 to H-79; M-1 to R 78; M-1 to G-77; M-1 to R-76; M-1 to H-75; M-1 to S-74; M-l to A-73; M-1 to A-72; M-l to E-71; M-1 to L-70; M-1 to S-69; M-1 to S-68; M-1 to V-67; M-1 to L-66; M-l to A 65; M-1 to V-64; M-l to P-63; M-l to L-62; M-l a A- t * s? * "61; Ml to Q-60; Ml to G-59; Ml to W-58; Ml to K-57 Ml to A-56; Ml to G-55; Ml to R-54; Ml to A-53;; Ml to L-52; Ml to L-51; Ml to H-50; Ml to P-49; Ml to P- 48; Ml to A-l; Ml to Q-46; Ml to G-45; Ml to L-44; Ml to P-43; Ml to L-42; Ml to E-41; Ml to E-40; M- :. to A-39; Ml to S-38; Ml to Y-37; Ml to C-36, Ml to H-35, Ml to P-34, Ml to T-33, Ml to G-32, Ml to H-31, Ml to S-30, Ml to H-29, Ml to P-28; Ml to H-27; Ml to G-26; Ml to R-25; Ml to L-24; Ml to S-23; Ml to P-22; Ml to D-21; Ml to H- 20; Ml to H-19; Ml to A-18; Ml to L-17; Ml to C-16; Ml to T-15; Ml to H-14; Ml to L-13; Ml to W-12; Ml to T-ll; Ml to L-10; Ml to F-9; Ml to L-8; Ml to L-7; and Ml to G-6 of SEQ ID NO: 29. Polypeptides are also provided; encoded by these polynucleotides The present invention is also directed to the nucleic acid molecules comprising, or consisting alternatively of a polynucleotide sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical. λ to the polynucleotide sequence encoding the IL-22 polypeptides described above. The present invention also encompasses the above polynucleotide sequences, fused to a heterologous polynucleotide sequence.

The invention also provides polypeptides having one or more amino acids deleted from the amino and carboxyl termini of IL-21, which can be described in general as having residues n5-m5 of SEQ ID NO: 29, where n5 and m5 they are integers as described above. Polynucleotides that code for such polypeptides are also provided. further, any polypeptide having one or more amino acids deleted from the amino and carboxyl termini of IL-21, specifically described as having the residues n5-m5 of SEQ ID NO: 29 (where n5 and m5 are integers as described above) may be excluded from the invention. Also as mentioned above, even if the deletion of one or more amino acids from the N-terminus of a protein results in the modification or loss of one or more biological functions of the protein, other biological activities may still be conserved. Thus, the ability of the shortened protein to induce and / or bind antibodies that recognize full-length, part-length or mature IL-22 polypeptides will in general be retained at least from most of the residues of full-length, part-length or mature IL-22 polypeptides are removed from the N-terminus. If a particular polypeptide lacking the N-thermal residues of a full length or full length polypeptide retains such immunological activities, this may be easily determined by routine methods described herein and otherwise known in the art. Accordingly, the present invention further provides polypeptides having one or more residues deleted from the amino terminus of the amino acid sequence of the IL-22 polypeptide shown in SEQ ID NO: 32, up to the aspartic acid residue at position number 168, and the polynucleotides encoding such polypeptides. In particular, the present invention provides the polypeptides comprising the The amino acid sequence of residues n6-173 of SEQ ID NO: 32, where n6 is an integer in the range of 1 to 168, and 168 is the position of the first residue from the N-terminus of IL-22 polypeptide ( shown in SEQ ID NO: 32) that is believed to be required for the immunogenic activity of the IL-22 protein. More particularly, the invention provides polynucleotides that encode polypeptides comprising, or consisting alternatively of, the amino acid sequence of residues C-2 to P-173; A-3 to P-173; D-4 to P-173; R-5 to P-173; P-6 to P-173; E-7 to P-173; E-8 to P-173; L-9 to P-173; L-10 to P-173; E-ll to P-173; Q-12 to P-173; L-13 to P-173; Y- 14 to P-173; G-15 to P-173; R-16 to P-173; L-17 to P-173; A-18 to P-173; A- 19 to P-173; G-20 to P-173; V-21 to P-173; L-22 to P-173; S-23 to P-173; A-24 to P-173; F-25 to P-173; H-26 to P-173; H-27 to P-173; T-28 to P-173; L-29 to P-173; Q-30 to P-173; L-31 to P-173; G-32 to P-173; P-33 to P-173; R-34 to P-173; E-35 to P-173; Q-36 to P-173; A- 37 to P-173; R-38 to P-173; N-39 to P-173; A- 40 to P-173; S-41 to P-173; C-42 to P-173; P-43 to P-173; A-44 to P-173; G-45 to P-173; G-46 to P-173; R-47 to P-173; P-48 to P-173; A- 49 to P-173; D-50 to P-173; R-51 to P-173; R-52 to P-173; F-53 to P-173; R-54 to P-173; P-55 to P-173; P-56 to P-173; T-57 to P-173; N-58 to P-173; L-59 to P-173; R-60 to P-173; S-61 to P-173; V-62 to P-173; S-63 to P-173; P-64 to P-173; W-65 to P-173; A-66 to P-173; Y-67 to P-173; R-68 to P-173; 1-69 to P-173; S-70 to P-173; Y- 71 to P-173; D-72 to P-173; P-73 to P-173; A-74 to P-173; R-75 to P-173; Y-76 to P-173; P-77 to P-173; R-78 to P-173; Y-79 to P-173; L-80 to P-173; P-81 to P-173; E-82 to P-173; A- 83 to P-173; Y- 84 to P-173; C-85 to P-173; L-86 to P-173; C-87 to P-173; R-88 to P-173; G-89 to P-173; C-90 to P-173; L-91 to P-173; T-92 to P-173; G-9? to P-173; L-94 to P-173; F-95 to P-173; G-96 to P-173; E-97 to P-173; E-98 to P-173; D-99 to P-173; V-100 to P-173; R-101 to P-173; F-102 to P-173; R-103 to P-173; S-104 to P-173; A-105 to P-173; P-106 to P-173; V-107 a P-173; Y-108 to P-173; M-l 09 to P-173; P-110 to P-173; T-111 to P-173; V-112 to P-173; V- 113 to P-173; L-114 to P-173; R-115 to P-173; R-116 to P-173; T-117 to P-173; P-118 to P-173; A- 119 to P-173; C-120 to P-173; A-121 to P-173; G-122 to P-173; G-123 to P-173; R-124 to P-1 3; S-125 to P-173; V-126 to P-173; Y-127 to P-173; T-128 to P-173; E-129 to P-173; A-130 to P-173; Y-131 to P-173; V-132 to P-173; T-133 to P-173; 1-134 to P-173; P-135 to P-173; V-136 to P-173; G-137 to P-173; C-138 to P-173; T-139 to P-173; C-140 to P-173; V-141 to P-173; P-142 a P-173; E-143 to P-173; P-144 to P-173; E-145 to P-173; K-146 to P-173; D-147 to P-173; A-148 to P-173; D-149 to P-173; S-150 to P-173; 1-151 to P-173; N-152 to P-173; S-153 to P-173; S-154 to P-173; 1-155 to P-173; D-156 to P-173; K-157 to P-173; Q-158 to P-173; G-159 to P-173; A-160 to P-173; K-161 to P-173; L-162 to P-173; L-163 to P-173; L-164 to P-173; G-165 to P-173; P-166 to P-173; N-167 to P-173; and D-168 to P-173 of SEQ ID NO: 32. Polypeptides encoded by these polynucleotides are also provided. The present application is also directed to nucleic acid molecules comprising, or alternatively consisting of, a polynucleotide sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to the The polynucleotide sequence encoding the IL-21 polypeptides described above. The present invention also encompasses the above polynucleotide sequences, fused to a heterologous polynucleotide sequence. 15 Also as mentioned above, even if the deletion of one or more amino acids from the C-terminus of a protein results in the modification or loss of one or more biological functions of the protein, other activities biologicals can still be conserved. Thus, the ability of the shortened polypeptide to induce and / or bind antibodies that recognize the full-length, part-length or mature IL-22 polypeptide will be in general retained at least from most of the Sískíar. ? ? »« Y A r ~ »r ¿? £ - f Ü ^ ^ - .-- a ^^ afea & aBa». «Residues of full-length, part-length or mature IL-22 polypeptides are removed from the C-terminus. If a particular polypeptide lacking the C-terminal residues of a complete protein does or does not preserve such immunological activities, this may be particularly determined by routine methods described herein and otherwise known in the art. . Accordingly, the present invention further provides polypeptides having one or more residues removed from the carboxyl terminus of the amino acid sequence of the IL-22 polypeptide shown in SEQ ID NO: 32, up to the residue of proline in position 6 of SEQ ID NO: 32, and the polynucleotides encoding such polypeptides. In particular, the present invention provides polypeptides having the amino acid sequence of residues 1-m6 of the sequence of amino acids in SEQ ID NO: 32, where m6 is any integer in the range of 6 to 173, and residue 6 is the position of the first residue of the C-terminus of the IL-22 polynucleotide (shown in SEQ ID NO. : 32) that is believed to be required for immunogenic activity of the IL-22 protein.

-XA, -A. , fc. ^ «Jt- -. . * • < «• -» • t *? M < *F? > «..

More particularly, the invention provides polynucleotides that encode polypeptides comprising, or consisting alternatively of, the amino acid sequence of residues G-1 to G-172; G-1 to A-171; G-1 to P-170 G-1 to A-169; G-1 to D-168; G-1 to N-167; G-1 to P-166 G-1 to G-165; G-1 to L-164; G-1 to L-163; G-1 to L-162 G-1 to K-161; G-1 to A-l 60; G-1 to G-159; G-1 to Q-158 G-1 to K-157; G-1 to D-156; G-1 to 1-155; G-1 to S-154 G-1 to S-153; G-1 to N-152; G-1 to 1-151; G-1 to S-150 G-1 to D-149; G-1 to A-148; G-1 to D-147; G-1 to K-146 G-1 to E-145; G-1 to P-144; G-1 to E-143; G-1 to P-142 G-1 to V-141; G-1 to C-140; G-1 to T-139; G-1 to C-138 G-1 to G-137; G-1 to V- 136; G-1 to P-135; G-1 to 1-134 G-1 to T-133; G-1 to V-132; G-1 to Y-131; G-1 to A-130 G-1 to E-129; G-1 to T-128; G-1 to Y-127; G-1 to V-126 G-1 to S-125; G-1 to R-124; G-1 to G-123; G-1 to G-122 G-1 to A-121; G-1 to C-120; G-1 to A- 119; G-1 to P-118 G-1 to T-117; G-1 to R-116; G-1 to R-115; G-1 to L-114 G-1 to V-113; G-1 to V-112; G-1 to T-III; G-1 to P-110 G-1 to M-l 09; G-1 to Y- 108; G-1 to V-107; G-1 to P-106 G-1 to A-105; G-1 to S-104; G-1 to R-103; G-1 to F-102 G-1 to R-101; G-1 to V-100; G-1 to D-99; G-1 to E-98; G-1 to E-97; G-1 to G-96; G-1 to F-95; G-1 to L-94; G-1 to G-93; G-1 to T-92; G-1 to L-91; G-1 to C-90; G-1 to G- »* T¡? Ák¿3 89; G-1 to R-88; G-1 to C-87; G-1 to L-86; G-1 to C-85; G-1 to Y-84; G-1 to A-83; G-1 to E-82; G-1 to P-81; G-1 to L-80; G-1 to Y-79; G-1 to R-78; G-1 to P-77; G-1 to Y-76; G-1 to R-75; G-1 to A-74; G-1 to P-73; G-1 to D-72; G-1 to Y-71; G-1 to S-70; G-1 to 1-69; G-1 to R-68; G-1 to Y- 67; G-1 to A-66; G-1 to W-65; G-1 to P-64; G-1 to S-63; G-1 to V-62; G-1 to S-61; G-1 to R-60; G-1 to L-59; G-1 to N-58; G-1 to T-57; G-1 to P-56; G-1 to P-55; G-1 to R-54; G-1 to F-53; G-1 to R-52; G-1 to R-51; G-1 to D-50; G-1 to A-49; G-1 to P-48; G-1 to R-47; G-1 to G-46; G-1 to G-45; G-1 to A-44; G-1 to P-43; G-1 to C-42; G-1 to S-41; G-1 to A-40; G-1 to N-39; G-1 to R-38; G-1 to A-37; G-1 to Q-36; G-1 to E-35; G-1 to R-34; G-1 to P-33; G-1 to G-32; G-1 to L-31; G-1 to Q-30; G-1 to L-29; G-1 to T-28; G-1 to H-27; G-1 to H-26; G-1 to F-25; G-1 to A-24; G-1 to S-23; G-1 to L-22; G-1 to V-21; G-1 to G-20; G-1 to A-l 9; G-1 to A-l 8; G-1 to L-17; G-1 to R-16; G-1 to G-15; G-1 to Y-14; G-1 to L-13; G-1 to Q-12; G-1 to E-11; G-1 to L-10; G-1 to L-9; G-1 to E-8; G-1 to E-7; G-1 a and G-1 to P-6 of SEQ ID NO: 32. Polypeptides encoded by these polynucleotides are also provided. The present application is also directed to the nucleic acid molecules comprising, or alternatively consisting of a polynucleotide sequence at least 90%, 95%, 96%, 97%, 98% or 99% identical to the polynucleotide sequence encoding the IL-22 polypeptides described above. The present invention also encompasses the above polynucleotide sequences, fused to a heterologous polynucleotide sequence. The invention also provides polypeptides having one or more amino acid residues deleted from the amino and carboxyl termini of IL-22, which can be generally described as having residues.; n6-m6 of SEQ ID NO: 32, where n6 and m6 are integers as described above. The polynucleotides encoding these polypeptides are also provided. In addition, any polypeptide having one or more amino acids deleted from the amino and carboxyl termini of IL-22, specifically described as having the residues n6-m6 of SEQ ID NO: 32 (where n6 and m6 are integers such as described above) can be excluded from the invention, as can polynucleotides encoding such polypeptides. The invention further includes variants of IL-21 and IL-22 polypeptides that show substantial biological activity. Such variants include deletions, insertions, inversions, repetitions, and substitutions selected according to the general rules known in the art because they have little effect on the activity. For example, regarding the guide to how to make phenotypically silent amino acid substitutions, it is provided by Bowie collaborators (Sci en ce 247: 1306-1310 (1990)), where the authors indicate that there are two main strategies for studying tolerance of an amino acid sequence to change. The first strategy exploits the tolerance of amino acid substitutions by natural selection during the evolutionary process. "By comparing amino acid sequences in different species, conserved amino acids can be identified.These conserved amino acids are probably important for protein function.In contrast, amino acid positions where substitutions have been tolerated by natural selection, indicate that these positions In this way, positions that tolerate amino acid substitution could be modified while still maintaining the biological activity of the protein.The second strategy uses genetic engineering to introduce amino acid changes in the proteins. specific positions of a cloned gene to identify regions critical to the function of the protein.For example, site-directed mutagenesis or alanine selection mutagenesis (introduction of simple alanine mutations into any residue in the molecule) can be used (Cunningham and Wells, Sci en ce 244: 1081-1085 (1989)). The resulting mutant molecules can then be tested for biological activity. As the authors state, these strategies have revealed that proteins are surprisingly tolerant of amino acid substitutions. The authors also indicate which amino acid changes are probably allowed at certain amino acid positions in the protein. For example, the most hidden amino acid residues (within the tertiary structure of the protein) require non-polar side chains, while few features of the surface side chains are generally conserved.

In addition, conservative amino acid substitutions, tolerated, involve the replacement of an aliphatic or hydrophobic amino acid with another aliphatic or hydrophobic amino acid such as Ala, Val, Leu or lie; the replacement of a hydroxylic residue with another hydroxylic residue such as Ser or Thr; the replacement of an acid residue with another acid residue such as Asp or Glu; the replacement of an amide residue with another amide residue such as Asn or Gln, the replacement of a basic residue with another basic residue such as Lys, Arg or His; the replacement of an aromatic residue with another aromatic residue such as Phe, Tyr, or Trp, and the replacement of a small-sized amino acid with another small-sized residue such as Ala, Ser, Thr, Met or Gly. In addition to the amino acid conservative substitution, variants of IL-21 and IL-22 include (i) substitutions with one or more of the non-conserved amino acid residues, where the substituted amino acid residues may or may not be one encoded by the code genetic, or (ii) substitution with one or more of the amino acid residues having a substituent group, or (iii) fusion of the mature polypeptide with another compound, such as a compound for ^^ increase the stability and / or solubility of the polypeptide (eg, polyethylene glycol), or (iv) fusion of the polypeptide with additional amino acids, such as a peptide from the IgG Fe fusion region, or the leader sequence or secretory, or a sequence that facilitates purification. Such variant polypeptides are considered within the scope of those skilled in the art from the teachings of the present invention. For example, variants of the IL-21 and IL-22 polypeptides containing amino acid substitutions, of amino acids loaded with other charged or neutral amino acids, can produce proteins with improved characteristics, such as less aggregation. Aggregation of pharmaceutical formulations reduces activity and increases clearance due to the immunogenic activity of the aggregate (Pinckard et al., Cl in. Exp. Immunol., 2: 331-340 (1967); Robbins et al., Di abe t is 36: 838-845 (1987); Cleland et al., Cri., Rev. Ther. Drug Carri er Sys t ems 10: 307-377 (1993)).

Fragments of Polynucleotides y_ of Polypeptides The invention provides nucleic acid molecules having nucleotide sequences related to extensive portions of SEQ ID NO: 3 and SEQ ID NO: 31, which have been determined from the following related cDNA clones: HE2CD08R (SEQ. ID NO: 24); HAGBX04R (SEQ ID NO: 25); HCEBA24FB (SEQ ID NO: 26); and HCELE59R (SEQ ID NO: 27). In addition, the invention provides nucleic acid molecules having related nucleotide sequences or extended portions of SEQ ID NO: 28 that has been determined from a designated cDNA clone designated HTGED19RB (SEQ ID NO: 30). Such polynucleotides (for example, SEQ ID NOs: 24, 25, 26 and 30) may be preferably excluded from the present invention. In the present invention, a "polynucleotide fragment" refers to a short polynucleotide having a nucleic acid sequence contained in the deposited clones, or shown in SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 28 or SEQ ID NO: 31. The short nucleotide fragments are preferably at least about 15 nucleotides, and more preferably at least 20 nt, still more preferably at least about 30 nt, and even more preferably, at least about 40 nt of length. A fragment "of at least 20 nt in length", for example, is intended to include 20 or more contiguous bases from the cDNA sequence contained in the deposited clones or the nucleotide sequences shown in SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 28 or SEQ ID NO: 31. These nucleotide fragments are useful as diagnostic probes and primers as discussed herein. Of course, larger fragments are preferred (eg, 50, 150, 500, 600, 2000 nucleotides). In addition, representative examples of the polynucleotide fragment of IL-21 include, for example, fragments having a sequence from about nucleotide number 1-50, 51-100, 101-150, 151-200, 201-250, 251-300. , 301-350, 351-400, 401-450, 451-500, 501-550, 551-600, 651-700, or 701 until the end of SEQ ID NO: 1 or the cDNA contained in the deposited clone. In addition, representative examples of IL-22 polynucleotide fragments include, for example, fragments having a sequence from about polynucleotide number 1-50, 51-100, 101-150, 151-200, 201-250, 251 -300, 301-350, 351-400, 401-450, 451-500, 501-550, 551-600, 601-650, 651-700, 701-750, 751-800, 801-850, 851-900 , 901-950, 951-1000, 1001-1050, 1051-1100, 1101-1150, 1151-1200, 1201-1250, 1251-1300, 1301-1350, 1351-1400, 1401-1450, 1451-1500, 1551 -1600 or 1601 until the end of SEQ ID NO: 3 or the cDNA contained in the deposited clone. In addition, representative examples of full-length IL-21 polynucleotide fragments include, for example, fragments having a sequence from about nucleotide number 1-1025, 50-1025, 100-1025, 150-1025, 200-1025, 250 -1025, 300-1025, 350-1025, 400-1025, 450-1025, 500-1025, 550-1025, 600-1025, 650-1025, 700-1025, 750-1025, 800-1025, 850-1025 , 900-1025, 950-1025, 1000-1025, 1-1000, 50-1000, 100-1000, 150-1000, 200-1000, 250-1000, 300-1000, 350-1000, 400-1000, 450 -1000, 500-1000, 550-1000, 600-1000, 650-1000, 700-1000, 750-1000, 800-1000, 850-1000, 900-1000, 950-1000, 1-950, 50-950 , 100-950, 150-950, 200-950, 250-950, 300-950, 350-950, 400-950, 450-950, 500-950, 550-950, 600-950, 650 -950, 700 -950, 750-950, 800-950, 850-950, 900-950, 1-900, 50-900, 100-900, 150-900, 200-900, 250-900, 300-900, 350-900 , 400-900, 450-900, 500-900, 550 -900, 600-900, 650-900, 700-900, 750-900, 800-900, 850-900, 1-850, 50-850, 100 -850, 150-850, 200-850, 250-850, 300-850, 350-850, 400-850, 450-850, 500 -850, 550-850, 600-850, 650-850, 700-850, 750-850, 800- 850, 1-800, 50-800, 100-800, 150-800, 200-800, 250-800, 300-800, 350-800, 400-800, 450-800, 500 -800, 550-800, 600-800, 650-800, 700-800, 750-800, 1-750, 50-750, 100-750, 150-750, 200-750, 250-750, 300-750, 350-750, 400-750, 450-750, 500-750, 550- 750, 600-750, 650-750, 700-750, 1-700, 50-700, 100-700, 150-700, 200-700, 250-700, 300-700, 350-700, 400-700 , 450-700, 500-700, 550-700, 600-700, 650-700, 1-650, 50-650, 100-650, 150-650, 200-650, 250- -650, 300-650, 350-650, 400-650, 450-650, 500-650, 550-650, 600-650, 1-600, 50-600, 100-600, 150-600, 200-600, 250-600, 300- 600, 350-600, 400-600, 450-600, 500-600, 550-600, 1-550, 50-550, 100- 550, 150- -550, 200-550, 250-550, 300-550 , 350-550, 400-550, 450-550, 500-550, 1-500, 50-500, 100-500, 150-500, 200-500, 250-500, 300-500, 350-500, 400 -500, 450-500, 1-450, 50-450, 100-450, 150-450, 200-450, 250-450, 300-450, 350-450, 400-450, 1-400, 50-400 , 100-400, 150-400, 200-400, 250-400, 300-400, 350-400, 1-350, 50-350, 100-350, 150-350, 200-350, 250-350, 300 -350, 1-300, 50-300, 100-300, 150-300, 200-300, 250-300, 1-250, 50-250, 100-250, 150-250, 200-250, 1-200 , 50-200, 100-20 0, 150-200, 1-150, 50-150, 100-150, 1-100, 50-100 and 1-50 of SEQ ID NO: 28. In this context "approximately" includes the intervals particularly indicated, more large or smaller by several (5, 4, 3, 2 or 1) nucleotides, at either end or at both ends. Preferably, these fragments encode a polypeptide having biological activity. In addition, the invention includes a polynucleotide comprising any portion of at least about 30 nucleotides, preferably at least about 50 nucleotides of SEQ ID NO: 1 from residue 1 to 650, 25 to 650, 50 to 650, 75 to 650 , 100 to 650, 125 to 650, 150 to 650, 200 to 650, 225 to 650, 250 to 650, 275 to 650, 300 to 650, 325 to 650, 350 to 650, 375 to 650, 400 to 650, 425 to 650, 450 to 650, 475 to 650, 500 to 650, 525 to 650, 550 to 650, 575 to 650, 600 to 650, 625 to 650, 1 to 600, 25 to 600, 50 to 600, 75 to 600 , 100 to 600, 125 to 600, 150 to 600, 175 to 600, 200 to 600, 225 to 600, 250 to 600, 275 to 600, 300 to 600, 325 to 600, 350 to 600, 375 to 600, 400 to 600, 425 to 600, 500 to 600, 525 to 600, 550 to 600, 575 to 600, 1 to 550, 25 to 550, 50 to 550, 75 to 550, 100 to 550, 125 to 550, 150 to 550 175 to 550, 200 to 550, 225 to 550, 250 to 550, 275 to 550, 300 to 550, 325 to 550, 350 to 550, 375 to 550, 400 to 550, 425 to 550, 500 to 550, 525 to 550, 1 to 500, 25 to 500, 50 to 500, 75 to 500, 100 to 500, 12 5 to 500, 150 to 500, 175 to 500, 200 to 500, 225 to 500, 250 to 500, 275 to 500, 300 to 500, 325 to 500, 350 to 500, 375 to 500, 400 to 500, 425 to 500, 450 to 500, 475 to 500, 1 to 450, 25 to 450, 50 to 450, 75 to 450, 100 to 450, 125 to 450, 150 to 450, 175 to 450, 200 to 450, 225 to 450, 250 to 450, 275 to 450, 300 to 450, 325 to 450, 350 to 450, 375 to 450, 400 to 450, 425 to 450, 1 to 400, 25 to 400, 50 'to 400, 75 to 400, 100 to 400, 125 to 400, 150 to 400, 175 to 400, 200 to 400, 225 to 400, 250 to 400, 275 to 400, 300 to 400, 325 to 400, 350 to 400, 375 to 400, 1 to 350 , 25 to 350, 50 to 350, 75 to 350, 100 to 350, 125 to 350, 150 to 350, 175 to 350, 200 to 350, 225 to 350, 250 to 350, 275 to 350, 300 to 350, 325 to 350, 1 to 300, 25 to 300, 50 to 300, 75 to 300, 100 to 300, 125 to 300, 150 to 300, 175 to 300, 200 to 300, 225 to 300, 250 to 300, 275 to 300 , 1 to 250, 25 to 250, 50 to 250, 75 to 250, 100 to 250, 125 to 250, 150 to 250, 175 to 250, 200 to 250, 225 to 250, 1 to 200, 25 to 200, 50 to 200, 75 to 200, 100 to 200, 125 to 200, 150 to 200, 175 to 200, 1 to 150, 25 to 150, 50 to 150, 75 to 150, 100 to 150, 125 to 150, 1 to 100, 25 to 100, 50 to 100, 75 to 100, 1 to 50 and 25 to 50. In addition, the invention includes a polynucleotide comprising any portion of at least about 30 nucleotides, preferably at least about 50 nucleotides, of SEQ ID NO: 3 from residue 300 to 850. More preferably, the invention includes a polynucleotide comprising the nucleotide residues 50 to 850, 75 to 850, 100 to 850, 125 to 850, 150 to 850, 175 to 850, 200 to 850, 225 to 850, 250 to 850, 275 to 850, 300 to 850, 325 to 850, 350 to 850, 375 to 850, 400 to 850, 425 to 850, 450 to 850, 475 to 850, 500 to 850, 525 to 850, 550 to 850, 575 to 850, 600 to 850, 625 to 850, 650 to 850, 675 to 850 , 700 to 850, 750 to 850, 775 to 850, 800 to 850, 50 to 800, 75 to 800, 100 to 800, 125 to 800, 150 to 800, 175 to 800, 200 to 800, 225 to 800, 250 to 800, 275 to 800, 300 to 800, 325 to 800, 350 to 800, 375 to 800, 400 to 800, 425 to 800, 450 to 800, 475 to 800, 500 to 800, 525 to 800, 550 to 800 , 575 to 800, 600 to 800, 625 to 800, 650 to 800, 675 to 800, 700 to 800, 750 to 800, 50 to 750, 75 to 750, 100 to 750, 125 to 750, 150 to 750, 175 to 750, 200 to 750, 225 to 750, 250 to 750, 275 to 750, 300 to 750, 325 to 750, 350 to 750, 375 to 750, 400 to 750, 425 to 750, 450 to 750, 475 to 750 , 500 to 750, 525 to 750, 550 to 750, 575 to 750, 600 to 750, 625 to 750, 650 to 750, 675 to 750, 700 to 750, 50 to 700, 75 to 700, 100 to 700, 125 to 700, 150 to 700, 175 to 700, 200 to 700, 225 to 700, 250 to 700, 275 to 700, 300 to 700, 325 to 700, 350 to 700, 375 to 700, 400 to 700, 425 to 700, 450 to 700, 475 to 700, 500 to 700, 525 to 700 , 550 to 700, 575 to 700, 600 to 700, 625 to 700, 650 to 700, 50 to 650, 75 to 650, 100 to 650, 125 to 650, 150 to 650, 175 to 650, 200 to 650, 225 to 650, 250 to 650, 275 to 650, 300 to 650, 325 to 650, 350 to 650, 375 to 650, 400 to 650, 425 to 650, 450 to 650, 475 to 650, 500 to 650, 525 to 650 , 550 to 650, 575 to 650, 600 to 650, 50 to 600, 75 to 600, 100 to 600, 125 to 600, 150 to 600, 175 to 600, 200 to 600, 225 to 600, 250 to 600, 275 to 600, 300 to 600, 325 to 600, 350 to 600, 375 to 600, 400 to 600, 425 to 600, 450 to 600, 475 to 600, 500 to 600, 525 to 600, 550 to 600, 50 to 550 , 75 to 550, 100 to 550, 125 to 550, 150 to 550, 175 to 550, 200 to 550, 225 to 550, 250 to 550, 275 to 550, 300 to 550, 325 to 550, 350 to 550, 375 to 550, 400 to 550, 425 to 550, 450 to 550, 475 to 550, 500 to 550, 50 to 500, 75 to 500, 100 to 500, 125 to 500, 150 to 500, 175 to 500, 200 to 500 225 to 500, 250 to 500, 275 to 500, 300 to 500 , 325 to 500, 350 to 500, 375 to 500, 400 to 500, 425 to 500, 450 to 500, 50 to 450, 75 to 450, 100 to 450, 125 to 450, 150 to 450, 175 to 450, 200 to 450, 225 to 450, 250 to 450, 275 to 450, 300 to 450, 325 to 450, 350 to 450, 375 to 450, 400 to 450, 50 to 400, 75 to 400, 100 to 400, 125 to 400 , 150 to 400, 175 to 400, 200 to 400, 225 to 400, 250 to 400, 275 to 400, 300 to 400, 325 to 400, 350 to 400, 50 to 350, 75 to 350, 100 to 350, 125 to 350, 150 to 350, 175 to 350, 200 to 350, 225 to 350, 250 to 350, 275 to 350, 300 to 350, 50 to 300, 75 to 300, 100 to 300, 125 to 300, 150 to 300 , 175 to 300, 200 to 300, 225 to 300, and 250 to 300. In the present invention, a "polypeptide fragment" refers to a short amino acid sequence contained in SEQ ID NO: 2, SEQ ID NO: 4 , SEQ ID NO: 29, SEQ ID NO: 32 or encoded by the cDNAs contained in the deposited clones. The protein fragments may be "free standing" or comprised within a larger polypeptide of which the fragment forms a part or region, more preferably as a single continuous region. Representative examples of the polypeptide fragments of the partial IL-21 of the invention, include, for example, fragments of approximately amino acid number 1-20, 21-40, 41-60, 61-83 or until the end of the coding region. In addition, the polypeptide fragments of IL-21 may be about 10, 20, 30, 40, 50, 60, 70 or 80 amino acids in length. Representative examples of the polypeptide fragments of IL-22 of the invention, include, for example, fragments from about amino acid numbers 1-20, 21-40, 41-60, 61-80, 81-100, 100- 120, 120-140, 140-160, or until the end of the coding region. In addition, the polypeptide fragments of IL-22 may be about 10, 2, 30, 40, 50, 60, 70, 80, 100, 120, 140 or 150 amino acids in length. Representative examples of the full-length IL-21 polypeptide fragments of the invention, include, for example, fragments from about amino acid number 1-20, 21-40, 41-60, 61-80, 81- 100, 100-120, 120-140, 140-160, 160-180, 180-200 or 180 until the end of the coding region. In addition, polypeptide fragments of full-length IL-21 may be about 10, 20, 30, 40, 50, 60, 70, 80, 100, 120, 140, 150, 160, 170, 180, or 190 amino acids of length. In this context "approximately" includes the intervals particularly indicated, larger or smaller by several (5, 4, 3, 2 or 1) amino acids at either end or both ends. A further embodiment of the invention relates to a peptide or polypeptide comprising the amino acid sequence of an IL-21 or IL-22 polypeptide having an amino acid sequence that contains at least one conserved amino acid substitution, but not more than 50 conserved amino acid substitutions, even more preferably, no more than 40 conserved amino acid substitutions, still more preferably, no more than 30 conserved amino acid substitutions, and even more preferably, no more than 20 conserved amino acid substitutions. Of course, in order to still increase the preference, it is highly preferable that a peptide or polypeptide has an amino acid sequence comprising the amino acid sequence of an IL-21 or IL-22 polypeptide, which contains at least one, but does not more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1 conservative amino acid substitutions. Preferred polypeptide fragments include secreted IL-21 and IL-22 proteins as well as mature forms. Additional preferred polypeptide fragments include secreted IL-21 and IL-22 proteins or mature forms having a continuous series of deleted residues from the amino or carboxyl terminus, or both. For example, any number of amino acids in the range of 1-60 can be deleted from the amino terminus of either the secreted or mature form of the IL-21 and IL-22 polypeptides. Similarly, any number of amino acids, in the range of 1 to 30, can be deleted from the carboxyl terminus of the secreted or mature form of the IL-21 and IL-22 polypeptides. In addition, any combination of deletions at the amino and carboxyl terminus above is preferred. Similarly, polynucleotide fragments encoding these polypeptide fragments of IL-21 or IL-22 are also preferred. Also preferred are polypeptide and polynucleotide fragments IL-21 and IL-22 characterized by structural or functional domains, such as the fragments comprising the alpha helical and alpha helical-forming regions, the beta-sheet and beta-sheet-forming regions, spinning and spin forming regions, helix and helix forming regions, hydrophilic regions, hydrophobic regions, alpha antipatic regions, beta antipatic regions, flexible regions, surface forming regions, binding region to the substrate, and regions of high antigenic index. The polypeptide fragments of SEQ. ID. NO: 2, SEQ. ID. NO: 4, SEQ. ID. NO: 29 or SEQ. ID. NO: 32 that fall within the conserved domains are specifically contemplated by the present invention (Figures 4, 5, 7 and 9). In addition, the polynucleotide fragments encoded by these domains are also contemplated. In additional embodiments, the polynucleotides of the invention encode functional attributes of IL-21 or IL-22. Preferred embodiments of the invention in this regard include the fragments comprising the alpha helix and alpha helix forming regions ("alpha regions"), the beta sheet and beta sheet forming regions ("beta regions"), the regions spinning and spin-forming ("spin regions"), helix and helix-forming regions ("helix regions"), hydrophilic regions, hydrophobic regions, alpha antipatic regions, beta antipatic regions, flexible regions , surface forming regions and regions of high antigenic index IL-21 or IL-22. The data representing the structural or functional attributes of IL-21 described in Figure 7 and / or Table I, as described above, were generated using the various DNA * STAR algorithms and modules set in the default parameters. The data representing the structural or functional attributes of IL-22 described in Figure 5 and / or Table II, Figure 9 and / or Table III, as described above, were generated using the various modules and DNA * STAR algorithms set to the default parameters. In a preferred embodiment, the data presented in columns VIII, IX, XIII and XIV of Table I can be used to determine the regions of IL-21 that show a high degree of potential for antigenicity. In a preferred additional embodiment, the data presented in columns VIII, IX, XIII and XIV of Tables II and / or III can be used to determine regions of IL-22 that show a high degree of potential for antigenicity. The regions of high antigenicity are determined from the data presented in columns VIII, IX, XIII and / or IV by choosing the values representing the polypeptide regions that are likely to be exposed on the surface of the polypeptide in a environment in which the recognition of the antigen can occur in the process of initiation of an immune response. Certain preferred regions in this respect are described in Figure 7, but can, as shown in Table I, be represented or identified by the use of tabular representations of the data presented in Figure 7. The DNA * STAR computer algorithm used to generate Figure 7 (set in the original default parameters) was used to present the data in Figure 7 in a tabular format (see Table I). The tabular format of the data in Figure 7 can be used to easily determine the specific limits of a preferred region. Certain preferred regions in this respect are described in Figures 5 and 8, but can, as shown in Tables II and III, respectively, be represented or identified by the use of tabular representations of the data presented in Figures 5 and 8. , respectively. The DNA * STAR computer algorithm used to generate Figures 5 and 8 (set in the original default parameters) was used to display the data in Figures 5 and 8 in a tabular format (See Tables II and III, respectively). The tabular format of the data in Figures 5 and 8 can be used to easily determine the specific limits of a preferred region. The aforementioned preferred regions described in Figures 5, 7 and 9, and in Tables II, I and III, respectively, include, but are not limited to, the regions of the aforementioned types identified by the sequence analysis. amino acids described in Figures 2A-B, 6A-B, and 8, respectively. As described in Figure 7, and in Table I, and in Figure 5 and Table II, and in Figure 8 and Table III, such preferred regions include the alpha regions, the beta regions, the turn regions, and the Garnier-Robson helix regions, the alpha regions, the beta regions, and the helix regions of Chou-Fasman, the hydrophilic regions of Kyte-Doolittle and the hydrophobic regions, the alpha and beta antiseptic regions of Eisenberg, the flexible regions of Karplus-Schulz, the surface formation regions of Emini and the Jameson-Wolf regions of high antigenic index.

TABLE I Res Position i p m iv v vi vp vip rx xi xp xm xrv -0.80 0.76 -0.40 0.36 -0.76 0.76 -0.40 0.44 Leu -1.18 0.76 -0.40 0.34 -1.60 1.01 -0.20 0.28 1.09 -0.05 0.16 Gly -2.12 1.39 -0.20 0.17 Leu -2.12 1.39 -0.20 0.17 -1.60 1.19 -0.60 0.16 Phe -1.60 1.67 -0.60 0.17 10 -1.42 1.93 -0.60 0.17 Thr 11 -1.39 1.74 -0.60 0.28 Trp 12 -1.24 1.54 -0.60 0.46 -1.24 1.33 -0.60 0.30 -1.13 1.33 -0.60 0.17 Thr 15 -0.36 1.34 -0.60 0.17 Cys 16 -0.08 0.93 -0.60 0.27 TABLE I (continued) Res Position i p ffl rv v vi vp vip rx X XI X? xm xrv 17 0.21 0.74 -0.60 0.27 Wing 18 0.81 0.24 0.10 0.32 His 19 0.54 0.19 0.44 0.91 0.04 -0.00 1.33 1.46 Asp 21 0.82 -0.00 2.22 1.21 Pro 22 1.29 -0.50 2.76 1.74 23 1.84 -0.57 F 3.40 1.27 Leu 24 T T 1.67 -0.57 F 3.06 1.03 Arg 25 1.67 -0.14 F 2.35 1.03 Gly 26 1.37 -0.07 F 2.14 1.05 27 1.54 -0.07 1.78 1.70 1.50 -0.26 1.57 1.18 His 29 2.00 0.17 1.30 1.18 1.17 1.26 His 31 1.99 0.16 0.84 1.26 Gly 1.36 0.23 F 0.86 1.26 Thr 33 1.32 0.30 F 0.58 0.50 Pro 34 1.06 0.67 F 0.15 0.58 His 0.77 0.56 0.20 0.78 Cys 36 0.80 0.20 0.55 Tyr 37 0.14 0.10 0.61 Ser 0.64 -0.29 0.50 0.78 TABLE I ( continuation) Res Position i p m rv v vi vp vm K x xi xp xm xrv 39 0.64 -0.10 0.45 1.20 Glu -0.13 -0.24 F 0.60 1.19 -0.31 0.30 0.73 Leu 42 0.43 -0.27 0.70 0.72 43 -. 43 -0.37 0.70 0.72 44 0.52 0.13 0.50 0.42 Gly 45 0.31 0.56 F 0.35 0.78 Gln 0.28 F 0.05 0.78 Wing 0.28 0.37 F 0.40 1.29 48 .0.32 0.37 F 0.60 1.08 -0.10 0.63 F -0.05 0.51 His 0.36 0.73 -0.20 0.51 51 0.01 0.23 0.10 0.65 Leu 52 0.01 0.23 -0.30 0.42 Wing 53 0.27 0.30 -0.30 0.31 Arg 54 AA 0.19 -0.20 0.30 0.75 Gly 55 -0.12 0.03 F -0.15 0.95 Wing 56 0.69 -0.23 * F 0.45 0.93 Lys 0.91 -0.33 * 0.85 0.83 Trp 58 0.69 0.17 * 0.25 0.84 Gly 59 C 0.37 0.43 -0.25 0.69 Gln 60 -0.14 0.36 * -0.30 0.53 TABLE I (continued) Res Position i p m rv v vi vp vm rx x xi xp xm xrv Wing 61 -0.14 1.00 -0.40 0.38 62 -. 62 -1.00 0.59 -0.60 0.38 63 -. 63 -1.57 0.84 -0.60 0.18 Val 64 -1.52 1.09 -0.60 0.13 Wing 65 -1.82 0.97 -0.60 0.22 66 -. 66 -2.04 0.67 -0.60 0.19 Val 67 -1.23 0.93 -0.60 0.21 -1.61 0.29 -0.30 0.36 69 -. 69 -1.34 0.29 -0.30 0.44 70 -. 70 -1.06 0.10 -0.30 0.60 Glu 71 -0.28 -0.16 0.30 0.60 Wing 72 0.69 0.61 Wing 0.64 -0.43 0.79 1045 74 1.06 1.48 0.83 75 A 1.83 -0.69 2.17 1.60 Arg 76 1.83 -0.69 2.66 2.16 Gly -1.19 3.40 Arg 78 2.91 -1.57 4.02 79 2.91 -1.64 2.66 3.17 -1.26 2.66 Arg 1.93 2.86 2.21 82 1.97 -0.70 3.06 2.35 TABLE I (continued) Res Posici n i p m rv v vi vp vm K x xi xp x xrv 83 1.86 -0.71 3.40 1.96 Wing 84 1.22 -0.31 2.76 1.73 1.01 0.26 1.27 0.60 Thr 86 0.04 0.26 0.93 0.69 Gln 87 -0.56 0.51 0.29 0.51 Cys -0.14 0.70 -0.60 0.29 Pro 89 0.23 0.21 -0.30 0.39 90 0.54 0.16 -0.10 0.35 0.86 0.65 A g 1.10 1.27 -0.14 -0.60 0.90 1.27 94 0.07 -0.56 * 0.90 1.27 Glu 95 0.33 -1.24 0.90 1.13 1.14 -0.74 0.60 0.74 Leu 97 0.72 -1.17 0.60 0.71 98 0.90 -0.69 0.60 0.59 99 0.90 -0.19 0.60 Asp 1.01 1.00 2.28 Thr 101 -1.11 1.30 2.58 His 102 1.49 -0.73 * 1.30 3.42 Gln 103 T T 1.19 -0.54 * 1.91 1.43 Arg 104 1.57 -0.16 * 1.42 1.33 TABLE I (continued) Res Position i p m rv v vi vp vm K xi xp xm xrv Ser 105 1.28 -0.21 > F 1.63 1.51 106 B 1.70 0.20 * F 0.89 0.92 107 1.49 -0.20 F 2.10 0.92 Pro 108 1.60 0.56 * * 1.34 1.07 Trp 109 0.63 0.17 1.28 3.00 Arg 110 0.93 0.13 0.87 1.66 Tyr 111 1.51 -0.26 1.40 1.80 Arg 112 1.81 -0.20 1.53 2.46 Val 113 2.02 -1.11 1.97 2.10 Asp 114 2.31 -1.11 3.06 2.32 Thr 115 2.31 -1.87 3.40 1.98 Asp 116 2.31 -1.87 3.06 5.23 Glu 117 1.99 -1.76 2.72 4.90 Asp 118 -1.33 2.38 5.25 Arg 119 2.89 -1.41 1.64 5.45 Tyr 120 2.39 1.30 6.29 121 1.80 -0.73 1.30 3.11 Gln 122 1.10 -0.23 0.60 1.60 Lys 123 0.51 0.56 * -0.45 0.89 124 0.40 0.30 * -0.30 0.58 Wing 125 -0.02 -0.13 0.30 0.58 Phe 126 A A -0.62 0.04 -0.30 0.16 TABLE I (continued) en Position i p m rv v vi vp vm K x xi xp xm xrv Wing 127 -1.29 0.73 *. . -0.60 0.16 Glu 128 -1.22 0.61 -0.60 0.08 Cys 129 -0.76 0.11 -0.30 0.19 130 -. 130 -0.83 -0.24 0.30 Cys 131 -1.02 -0.17 1.10 0.06 Arg 132 -0.43 0.51 0.20 Gly 133 -1.02 -0.06 1.10 0.15 Cys 134 -0.24 -0.24 0.28 lie 135 0.26 -0.81 1.40 0.28 Asp 136 0.58 1.80 0.41 Wing 137 2.10 0.76 Arg 138 T C 0.92 -0.90 3.00 2.12 Thr 139 1.28 -1.59 * 2.70 2.20 Gly 140 -1.10 2.40 3.14 Arg 141 A 0.99 -1.10 1.90 1.62 Glu 142 A A 0.77 -0.60 1.20 1.13 Thr 143 A A 0.45 0.94 Wing 144 A A 0.67 -0.43 0.30 0.7T Wing 145 A A -0.04 0.30 0.60 Leu 146 A 0.16 0.60 -0.60 0.31 147 -. 147 -0.66 0.11 -0.30 0.60 -1.16 0.30 * -0.30 0.49 TABLE I (continued) Res Position i p m rv v vi vp vm rx XI xp xm xrv Val -0.57 0.49 * -0.60 0.49 Arg 150 -0.28 0.20 * -0.30 0.53 151 -. 151 -0.28 0.19 -0.30 0.53 Leu 152 -1.09 0.49 -0.60 0.58 Gln 153 -1.64 0.53 -0.60 0.25 154 -. 154 -1.60 1.17 -0.60 0.22 Leu 155 -1.60 1.17 -0.60 0.22 156 -. 156 -0.68 0.49 -0.60 0.25 157 B B 0.09 -0.30 0.37 Leu 158 0.30 0.87 Arg -0.33 -0.56 1.30 1.63 Arg B T 0.18 -0.67 1.30 1.18 Arg 161 C 1.10 -0.93 1.10 1.91 Pro 162 1.96 -1.61 1.84 1.91 Cys 163 2.42 -1.16 2.18 1.63 164 2.01 -1.19 2.57 0.82 Arg 165 1.56 -0.80 * 2.91 0.71 Asp 166 0.63 -0.80 * 3.40 1.32 Gly 167 0.63 -0.69 2.91 0.81 168 0.99 -0.64 2.37 0.64 Gly 169 C 1.08 -0.16 1.53 0.55 170 C 0.62 0.27 0.59 0.87 TABLE I (continued) Res Position I H O IV V VI VII Vffl DC X XI XII Xffl XIV 171 0.03 0.27 0.25 0.64 Thr 172 -0.32 0.39 0.45 0.65 173 -. 173 -0.61 0.74 0.15 0.68 Gly 174 -0.97 0.56 0.15 0.45 Wing 175 -0.19 0.91 -0.20 0.27 176 -. 176 -0.29 0.93 -0.60 0.24 Wing 177 0.02 0.99 -0.60 0.34 Phe 178 -0.47 0.56 -0.60 0.59 179 -. 179 -1.01 0.84 -0.60 0.59 180 -. 180 -0.46 0.74 -0.60 0.41 Glu 181 -0.61 0.74 -0.60 0.64 -0.23 0.60 -0.60 0.35 lie 183 -0.39 0.53 -0.20 0.38 His 184 -0.70 0.69 -0.20 0.16 Val C -1.06 1.11 -0.40 0.18 Pro 186 -1.37 0.90 0.20 0.14 187 -. 187 -1.33 0.70 0.20 0.15 Gly T T -1.30 0.77 0.20 0.11 Cys -2.08 0.77 0.20 0.05 190 -. 190 -1.43 1.03 -0.60 0.06 Cys 191 B B -1.11 0.81 -0.60 0.09 -0.56 0.39 * -0.30 0.33 TABLE I (continued) Res Position I H m IV V VI VD Vffl DC X XI XII Xffl XIV Leu 193 -1.07 0.20 * 0.28 0.31 Pro 194 -0.79 0.36 * 0.61 0.42 Arg 195 T T -0.87 0.21 1.04 0.73 196 T T -0.59 -0.00 * 1.97 1.13 Val 197 -0.12 -0.26 1.80 0.93 TABLE II Res Position I II III IV V VI VII VIII IX XI XII XIII 0.85 1.60 0.65 1.26 A B 0.88 0.75 1.93 Arg A B 0.41 0.75 1.22 Wing A B 0.30 0.67 Arg AB -0.31 0.30 0.55 AB -0.60 0.30 0.38 -0.71 -0.30 0.38 AB -0.86 * -0.60 0.17 AB -0.30 * -0.60 0.22 AB -0.72 * -0.60 0.41 12 -0.94 -0.60 0.72 -0.09 * -0.60 0.44 TABLE II (continued) Res Position I II III IV V VI VII VII IX X XI XII XIII His 14. A B. . . . -0.09 *. . -0.60 0.76 Thr 15 A B -0.13 *. . -0.60 0.72 Leu 16 TO . . . . C 0.24 * *. -0.10 0.52 Gln 17 TO . . T. . 1.06 * *. 0.40 0.60 Leu 18 A. . . . C 1.09. * F 0.80 0.81 Gly 19 T C 1.12. * F 2.40 1.70 Pro 20. . . . . T C 0.84 * * F 3.00 1.70 Arg 21 T C 1.77 * * F 2.70 2.08 Glu 22 B. . T 1.77 * * F 2.20 4.12 Gln 23 B 1.99 * * F 1.70 4.28 Wing 24 . . . T. . 2.03 * * F 1.80 2..21 Arg 25 T 1.58 * * F 1.50 1.71 Asn 26 . . . T. . 1.26 * * F 1.05 0.53 Wing 27 . . . T. . 0.67 *. . 0.90 0.81 Ser 28 . B. . . . 0.32. . . 0.78 0.42 Cys 29 B. . T 0.57. *. 0.66 0.26 Pro 30. . . . T T 0.57. *. 1.34 0.25 Wing 31 . . . T T 0.36. * F 2.37 0.37 Gly 32 . . T T 0.36. . F 2.80 1.06 Gly 33 . . . . . C 0.66 * * F 1.97 0.69 Arg 34 B. 1.43 *. F 1.94 1.15 Pro 35 B. . T 1.76 *. F 1.86 2.27 TABLE I I (continued) Res Position I II III IV V VI VII VII IX X XI XII XIII Wing 36 . B. . T. 1.64 * * F 1.58 4.49 Asp 37 . B T 2.10 * * F 1.30 1.99 Arg 38 . B. . T. 2.23 * * F 1.30 2.52 Arg 39 B 1.91. * * F 1.10 3.85 Phe 40 B 1.81 * * F 1.44 3.57 Arg 41 B 2.40 * * F 1.78 2.63 Pro 42 T C 1.59. * F 2.22 2.16 Pro 43 T T 1.59. * F 2.16 2.05 Thr 44 . . . T T 1.18. * F 3.40 2.05 Asn 45 T C 1.02 * * F 2.56 1.78 Leu 46 B B 0.61 * * F 0.87 0.85 Arg 47 B B 0.61 *. F 1.13 0.79 Ser 48 B B 0.53 *. F 0.79 0.76 Val 49 . B B . . 0.26 *. F -0.45 0.97 Be 50 B . T 0.01 * * F 0.25 0.50 Pro 51 . B. . T. 0.93 * *. -0.20 0.59 Trp 52 . B. . T. -0.07 * *. -0.05 1.55 Wing 53 . B. . T. -0.07 * *. -0.20 0.81 Tyr 54 . B B . . 0.54 * *. -0.60 0.70 Arg 55 . B B . . 0.84. *. -0.45 1.05 I'm 56 B B.. 0.84 * *. 0.13 1.73 Be 57. . B. . . . 0.54 * *. 0.61 1.71 TABLE I I (continued) Res Position I II III IV V VI VII VII IX X XI XII XIII Tyr 58 . . . T. . 1.24 * *. 1.74 0.88 Asp 59 T C 1.24 * * F 2.32 2.46 Pro 60 . . . T T 0.92 *. F 2.80 2.88 Wing 61 . . . T T 1.92 *. F 2.52 2.84 Arg 62 . B. . T. 1.98 *. F 2.14 3.33 Tyr 63 B. . T 1.41 *. . 1.41 3.37 Pro 64 B. . T 1.20 *. . 0.53 2.75 Arg 65 . . . T T 1.41 *. . 0.65 2.17 Tyr 66 . B. . T. 1.41 *. F 0.40 2.40 Leu 67 . B. . . . 1.06 *. F 0.80 1.57 Pro 68. . B. . . . 0.63 *. . 0.05 1.26 Glu 69 . . . T. . 0.03 *. . 0.00 0.43 Wing 70 . B B . . -0.74 *. . -0.60 0.43 Tyr 71 . B B . . -0.39 *. . -0.60 0.15 Cys 72 B B 0.08 *. . -0.30 0.17 Leu 73 B B -0.38. *. -0.60 0.16 Cys 74 . B. . T. -1.19. *. -0.20 0.06 Arg 75 . B. . T. -0.91 * *. -0.20 0.09 Sly 76 B. . T -1.01 *. . -0.20 0.15 Cys 77 . B. T. -1.16. *. 0.10 0.28 Leu 78 . B B . . -1.04. . . -0.30 0.12 Thr 79 . B B . . -0.72. *. -0.60 0.10 TABLE I I (continued) Res Position I II III IV V VI VII VIII IX XI XII XIII Gly 80 C -0.83 -0.40 0.19 C -0.49 -0.40 0.40 Phe B B 0.18 0.45 0.48 Gly C 0.13 0.95 0.81 Glu A B 0.56 0.45 0.73 Glu 85 0.20 0.90 1.65 Asp 86 A B B 1.12 0.90 1.45 Val 87 A B B 1.52 0.90 1.63 Arg 1.15 1.26 1.07 1.00 0.77 Arg 90 B T 0.21 0.85 1.59 C -0.03 0.50 0.60 Wing 92 C 0.22 -0.25 1.09 93 C -0.10 -0.10 0.55 Val 94 0.29 * -0.20 0.64 Tyr -0.68 * -0.60 0.91 96 B B. . -0.60 0.44 Pro 97 B ß -1.46 -0.60 0.44 98 -. 98 -1.13 * -0.60 0.23 -0.17 * -0.60 0.46 Val -0.23 0.30 0.58 Leu 101 0.30 0.58 TABLE I I (continued) Res Position I II III IV V VI VII VIII IX XI XII XIII Arg 102 B B 0.60 1.20 Arg 103 B B -0.58 0.60 1.63 Thr 104 B B -0.31 0.60 1.06 Pro 105 B B 0.20 1.00 0.55 Wing 106 0.67 1.00 0.28 Cys 107 0.67 0.85 0.19 Wing 108 0.26 * 2.10 0.24 Gly 109 T T -0.29 * 2.50 0.32 Gly 110 T T -0.32 * 2.25 0.44 Arg B B -0.04 * 0.60 0.69 Ser 112 0.62 * 0.35 1.00 113 B B 0.62 0.70 1.75 Tyr 114 0.72 0.50 0.90 Thr 115 0.21 -0.25 1.05 Glu 116 -0.21 -0.45 1.05 Wing 117 B B -0.80 -0.60 0.97 Tyr -0.16 -0.60 0.47 Val 119 -0.77 -0.60 0.42 Thr 120 B B -0.80 -0.60 0.31 121 B B -1.47 -0.60 0.20 Pro 122 -1.19 -0.20 0.14 -1.61 0.20 0.14 TABLE I I (continued) Res Position I II III IV V VI VII VII IX X XI XII XIII Gly 124 T T -1.61. . . 0.20 0.11 Cys 125 B. . T -1.51. . . -0.20 0.05 Thr 126 . B. . . . -0.62. . . -0.40 0.11 Cys 127 B -0.62. . . -0.10 0.19 Val 128 B. . T 0.23. . . 0.40 0.55 Pro 129. . B. . T. 0.62. . F 1.45 0.65 Glu 130 . B. . T. 1.29 *. F 2.20 2.44 Pro 131. . B. . T. 1.01 *. f 2.50 5.49 Glu 132 1.68 *. F 3.00 3.59 Lys 133 A. . T. . 2.23 *. F 2.30 3.46 Asp 134 A . . . T. 1.56 *. F 2.20 3.00 Ala 135 A. . . T. 1.56 *. F 1.90 1.21 Asp 136 A. . . T. 1.47 *. F 1.45 0.98 Ser 137 . B. . T. 1.17 *. F 1.15 0.78 lie 138. . B. . . . 0.23 *. F 0.80 1.04 Asn 139 . B. . T. 0.23 *. F 0.85 0.44 Ser 140 . B. . T. 0.87 *. F 1.16 1.54 Ser 141 . B. . T. 0.87 *. F 1.62 1.55 I have 142. . B. . T. 0.82. * F 2.23 1.67 Asp 143. B. . T. 1.12 * * F 2.54 1.23 Lys 144 . . . T T 1.17 *. . F 3.10 0.93 Gln 145 . B. . T. 0.66 *. F 2.54 2.65 TABLE I I (continued) Res Position I II III IV V VI VII VII IX X XI XII XIII Gly 146 B. . T 0.14 *. F 2.23 1.31 Wing 147 A B. . . . 0.22 *. F 1.07 0.54 Lys 148 A B. . . -0 12. F 0.16 0.26 Leu 149 A B. . . . -0.38 *. . -0.60 0.26 Leu 150 A B. . . . -0.38. . . -0.60 0.39 Leu 151 A B. . . . -0.03. . . -0.06 0.32 Gly 152 . B. . T. -0.03. . F 0.73 0.64 Pro 153. . . . . T C -0.29. . F 1.17 0.78 Asn 154 . . . T T -0.07. . F 2.36 1.47 Asp 155 . . . T C 0.40. . F 2.40 1.50 Wing 156 . . . . . C 1.00. . F 1.81 0.96 Pro 157. . . . . T C 0.96. . F 1.77 0.921 Wing 158 . . . . T C 0.78. . . 1.38 0.71 Gly 159 . . . . T C 0.39. . . 0.54 0.90 Pro 160 . B. . T. 0.00. . . 0.10 0.74 TABLE I I I Res Position I II III IV V VI VII VII IX X XI XII Gly 0.46 -0.21. 0.70 0.34 Cys 0.63 -0.64. 1.00 0.51 Wing 1.02 -0.64. 0.80 0.62 Asp 1.41 -1.07. 0.85 1.09 Arg 0.99 -1.50 * 0.90 3.51 Pro 0.52 -1.39 »0.90 2.87 Glu 1.19 -1.20 * 0.90 1.42 Glu 1.78 -1.20 * 0.90 1.25 0.97 -0.80 * 0.90 1.40 0.61 -0.54 * 0.75 0.67 11 A A 0.48 0.21 * -0.30 0.60 Gln 12 0.59 -0.60 0.73 Leu 13 A A -0.22 -0.04 * 0.45 1.72 Tyr 14 -0.00 -0.04 * 0.30 0.82 Gly 15 0.22 0.46 * -0.60 0.48 Arg -0.12 0.56 * -0.60 0.59 Leu 17 A A -0.98 0.30 * -0.30 0.37 Wing -0.98 0.19 * -0.30 0.28 Wing -1.03 0.44 * -0.60 0.12 Gly 20 A A -1.28 0.83 * -0.60 0.19 -2.09 0.64 * -0.60 0.19 22 A A -1.31 0.93. -0.60 0.16 TABLE I I I (continued) II III IV V VI VII VII IX X XI XII 23 -0.76 0.93. -0.60 0.22 24 -. 24 -0.48 1.00. -0.60 0.41 -. 25 -0.94 0.84 * -0.60 0.72 His 26 -0.09 0.84 * -O.í 0.44 -0.09 0.86 * -0.60 0.76 28 -. 28 -0.13 1.04. -0.60 0.72 29 0.24 0.69 * -0.10 0.52 Gln 1.06 0.61 * 0.40 0.60 31 C 1.09 0.11. 0.80 0.81 Gly 32 -0.37. 2.40 1.70 Pro 0.84 -0.66 * 3.00 1.70 Arg 1.77 -0.56 * 2.70 2.08 Glu 35 1.77 -1.24 * 2.20 4.12 36 1.99 -1.27 * 1.70 4.28 37 2.03 -1.20 * 1.80 2.21 Arg 38 1.58 -0.81 * 1.50 1.71 39 1.26 F 1.05 0.53 Wing 40 -0.21 * 0.90 0.81 -0.21. 0.78 0.42 Cys 0.57 0.21. 0.66 0.26 43 0.57 0.24. 1.34 0.25 T T 0.36 -0.26. F 2.37 0.37 TABLE I I I (continued) Res Position I II III IV V VI VII VII IX X XI XII XIII Gly 45 T T 0.36 -0.21. . F 2.80 1.06 Gly 46 . . . . . C 0.66 -0.29 * * F 1.97 0.69 Arg 47 B 1.43 -0.71 *. F 1.94 1.15 Pro 48 B. . T 1.76 -1.21 *. F 1.86 2.27 Wing 49 . B. . T. 1.64 -1.64 * * F 1.58 4.49 Asp 50 . B. . T. 2.10 -1.29 * * F 1.30 1.99 Arg 51 B. . T 2.23 -1.29 * * F 1.30 2.52 Arg 52 B 1.91 -1.29 * * F 1.10 3.85 Phe 53 . B. . . . 1.81 -1.36 * * F 1.44 3.57 Arg 54 B 2.40 -0.87 * * F 1.78 2.63 Pro 55. . . . . T C 1.59 -0.47. * F 2.22 2.16 Pro 56 T T 1.59 0.21. * F 2.16 2.05 Thr 57 . T T 1.18 -0.57. * F 3.40 2.05 Asn 58 . . . T C 1.02 -0.19 * * F 2.56 1.78 Leu 59 . B B . . 0.61 0.03 * F 0.87 0.85 Arg 60 B B 0.61 -0.01 * * F 1.13 0.79 Ser 61 B B. 0.53 -0.07 *. F 0.79 0.76 Val 62 . B 3. . . 0.26 0.44 *. F -0.45 0.97 Ser 63 . B. . T. 0.01 0.26 * * F 0.25 0.50 Pro 64 . B. . T. 0.93 1.01 * *. -0.20 0.59 Trp 65 . B. . T. -0.07 0.63 * *. -0.05 1.55 Wing 66 . B. . T. -0.07 0.67 * *. -0.20 0.81 TABLE I I I (continued) Res Position I II III IV V VI VII VII IX X XI XII XIII Tyr 67 . B B . . 0.54 0.67 * *. -0.60 0.70 Arg 68 B B. 0.84 1.00 *. -0.45 1.05 lie 69. . B B . . 0.84 0.09 *. 0.13 1.73 Be 70 . B. . . . 0.54 0.01 *. 0.61 1.71 Tyr 71 . . . T. . 1.24 -0.24 * *. 1.74 0.88 Asp 72 T C 1.24 -0.24 * * F 2.32 2.46 Pro 73 T T 0.92 -0.17 * * F 2.80 2.88 Wing 74 . . . T T 1.92 -0.13 *. F 2.52 2.84 Arg 75 B. . T 1.98 -0.89 *. F 2.14 3.33 Tyr 76 . B. . T. 1.41 -0.13 *. . 1.41 3.37 Pro 77. . B. . T. 1.20 0.13 *. . 0.53 2.75 Arg 78 . . . T T 1.41 0.06 *. . 0.65 2.17 Tyr 79 B. . T 1.41 0.06 *. F 0.40 2.40 Leu 80 B 1.06 -0.20 *. F 0.80 1.57 Pro 81 B 0.63 0.13 *. . 0.05 1.26 Glu 82 . T. 0.03 0.70 *. . 0.00 0.43 Wing 83 B B -0.74 0.63 *. . -0.60 0.43 Tyr 84 . B B . . -0.39 0.51. . . -0.60 0.15 Cys 85 B B 0.08 0.09 *. . -0.30 0.17 Leu 86 B B -0.38 0.51. *. -0.60 0.16 Cys 87 . B. . . . -1.19 0.59. *. -0.20 0.06 Arg 88 . B. . T. -0.91 0.51 * *. -0.20 0.09 TABLE I I I (continued) Res Position I II III IV V VI VII VII IX X XI XII XIII Gly 89 B. . T -1.01 0.43 *. . -0.20 0.15 Cys 90 B. . T -1.16 0.17. *. 0.10 0.28 Leu 91. . B B T. -1.04 0.29. . . -0.30 0.12 Thr 92 . B B . . -0.72 1.07. *. -0.60 0.10 Gly 93 . . B. . C -0.83 1.07. *. -0.40 0.19 Leu 94 . . B. . C -0.49 0.50. . . -0.40 0.40 Phe 95 . B B . . 0.18 -0.19. . F 0.45 0.48 Gly 96 A. . B 0.13 -0.67. * F 0.75 0.81 Glu 97 A A 0.56 -0.46. * F 0.45 0.73 Glu 98 A A. . . . . 0.20 -1.14. * F 0.90 1.65 Asp 99 A A. B 1.12 -1.14. * F 0.90 1.45 Val 100 A A. B. . . 1.52 -1.57. * F 0.90 1.63 Arg 101 A A. B. 1.28. -1.19. *. 0.75 1.26 Phe 102 A A. B. 1.07 -0.69. *. 0.60 0.77 Arg 103 A A. B. . . 0.21 -0.26. *. 0.45 1.59 Ser 104 TO . B. . C -0.03 -0.26. *. 0.50 0.60 Ala 105 . . B. . C 0.22 0.50. *. -0.25 1.09 Pro 106. . . B. . C -0.10 0.33. *. -0.10 0.55 Val 107. . B T . 0.29 0.76 *. . -0.20 0.64 Tyr 108 . B B . . -0.68 0.86 *. . -0.60 0.91 Met 109 B B -1.23 1.00. . . -0.60 0.44 Pro 110 . B B . . -1.46 1.21. *. -0.60 0.44 TABLE I I I (continued) Res Position I II III IV V VI VII VII IX X XI XII XIII Thr 111 B B -1.13 1.26 *. . -0.60 0.23 Val 112 . B B . . -0.17 0.50 *. . 0.30 0.46 Val 113 . B B . . -0.23 -0.11. . . 0.30 0.58 Leu 114. . B B . . 0.16 -0.06. . . 0.60 0.58 Arg 115 . B B . . -0.22 -0.11. . F 0.60 1.20 Arg 116 . B B . . -0.58 -0.26. . F 0.60 1.63 Thr 117 . B B . . -0.31 -0.33. . F 0.60 1.06 Pro 118 B B 0.20 -0.51 *. F 1.00 0.55 Wing 119 . B. . . . 0.67 -0.09. *. 1.00 0.28 Cys 120 . B. . T. 0.67 0.34. *. 0.85 0.19 Wing 121 . . T T 0.26 -0.14 * *. 2.10 0.24 Gly 122 . . . T T -0.29 -0.19 *. F 2.50 0.32 Gly 123 . . . T T -0.32 -0.04 *. F 2.25 0.44 Arg 124 B B -0.04 0.14. . F 0.60 0.69 Ser 125 . B B . . 0.62 0.13. . F 0.35 1.00 Val 126 . B B . . 0.62 -0.30. . . 0.70 1.75 Tyr 127 B 0.72 -0.23. . . 0.50 0.90 Thr 128 B 0.21 0.53. . . -0.25 1.05 Glu 129 . B B . . -0.21 0.79. *. -0.45 1.05 Wing 130 . B B . . -0.80 0.63. *. -0.60 0.97 Tyr 131 B B -0.16 0.56. *. -0.60 0.47 Val 132 B B -0.77 0.50. *. -0.60 0.42 TABLE I I I (continued) Position I II III IV V VI VII VII IX X XI XII XIII 133 -. 133 -0.80 1.14. -0.60 0.31 134 -. 134 -1.47 1.07. -0.60 0.20 135 -. 135 -1.19 0.89. -0.20 0.14 Val 136 T T -1.61 0.73. 0.20 0.14 Gly 137 -1.61 0.81. 0.20 0.11 Cys 138 -1.51 0.77. -0.20 0.05 Thr 139 -0.62 0.77. -0.40 0.11 Cys 140 -0.62 0.13. -0.10 0.19 Val 141 0.23 0.13. 0.10 0.55 Pro 142 0.62 F 0.85 0.65 Glu 143 1.29 -0.93. F 1.30 2.44 Pro 144 1.01 -1.50 * 1.30 5.49 Glu 145 -1.64 * 1.10 3.59 Lys 146 2.23 -2.07 * F 1.10 3.46 Asp 147 1.56 -1.69. F 1.30 3.00 Wing 1.56 -1.43 * F 1.30 1.21 Asp 149 1.47 -1.03 * F 1.15 0.98 150 1.17 -0.64 * F 1.15 0.78 lie 151 0.23 -0.26 * * F 0.80 1.04 152 0.23 -0.07 * F 0.85 0.44 Ser 153 0.87 -0.07 * F 0.85 0.54 Ser 154 0.87 -0.46 * * F 1.00 1.55 TABLE III (continued) Res Position I II III IV V VI VII VIII IX X XI XII XIII lie 155 0.82 -0.74. 1.30 1.67 Asp 156 1.12 -0.71 * 1.30 1.23 Lys 157 1.17 -0.60 * 1.15 0.93 158 0.66 -0.99 * 1.30 2.65 Gly 159 0.14 -0.99 * 1.30 1.31 Wing 160 0.22 -0.30 * 0.45 0.54 Lys 161 -0.12 0.39. -015 0.26 162 -0.38 0.41. -060 0.26 Leu 163 -0.38 0.41. -0.60 0.39 164 A B -0.03 0.31. -0.06 0.32 Gly 165 -0.03 0.31. F 0.73 0.64 Pro 166 -0.29 0.13. F 1.17 0.78 -0.07 -0.13. F 2.36 1.47 Asp 168 -0.31. F 2.40 1.50 Wing 169 1.00 -0.31. F 1.81 0.96 170 0.96 -0.31. F 1.77 0.92 Wing 171 0.78 -0.29. 1.38 0.71 Gly 172 0.39 0.54 0.90 173 0.07. 0.10 0.74 Among the fragments highly preferred to this respect is that of those who understand the . isfessafe-,.

IL-21 or IL-22 regions that combine several structural features, such as several of the features described above. Other preferred fragments are fragments of biologically active IL-21 and IL-22. The biologically active fragments are those that show similar activity, but not necessarily identical, to an activity of the IL-21 and IL-22 polypeptides. The biological activity of the fragments may include a desired, improved activity, or an undesirable, decreased activity.

Transgenic and "inactivated" The polypeptides of the invention can also be expressed in transgenic animals. Animals of any species, including, but not limited to, mice, rats, rabbits, hamsters, guinea pigs, pigs, micro-nerds, goats, sheep, cows and non-human primates, for example, baboons, monkeys and chimpanzees, can be used to generate transgenic animals. In a specific embodiment, techniques described herein or otherwise known in the art, are used to express polypeptides of the invention in humans, as part of a gene therapy protocol. Any technique known in the art can be used to introduce the transgene (e.g., the polynucleotides of the invention) into animals to produce the founder lines of transgenic animals. Such techniques include, but are not limited to, pronuclear microinjection (Paterson et al., Appl Microbiol Biotechnol 40: 691-698 (1994); Carver et al., Biotechnology (NY) 11: 1263-1270 (1993); Wright et al., Biotechnology (NY) 9: 830-834 (1991); and Hoppe et al., U.S. Patent No. 4,873,191 (1989)); gene transfer mediated by retroviruses within germ lines (Van der Putten et al., Proc. Nati, Acad. Sci., USA 82: 6148-6152 (1985)), blasts or embryos; direction to the gene in totipotential embryonic cells (Thompson et al., Cell 56: 313-321 (1989)); electroporation of cells or embryos (Lo, 1983, Mol Cell, Biol. 3: 1803-1814 (1983)); introduction of the polynucleotides of the invention using a gene gun (see for example Ulmer et al., Science 259: 1745 (1993); the introduction of nucleic acid constructs into totipotent embryonic pleuripotent cells and transfer of totipotent cells again in blasts, and sperm-mediated gene transfer (Lavitrano et al., Cell 57: 717-723 (1989), etc. For a review of such techniques, see Gordon, "Transgenic Animáis". Intl. Rev. Cytol. 115: 171-229 (1989), which is incorporated by reference herein in its entirety. Any technique known in the art can be used to produce transgenic clones containing polynucleotides of the invention, for example, nuclear transfer within enucleated oocytes of nuclei from embryonic, fetal, or adult cells, cultured, induced at rest (Campell). et al., Nature 380: 64-66 (1996), Wil ut et al., Nature 385: 810-813 (1997) The present invention provides transgenic animals that carry the transgene in their cells, as well as animals that carry the transgen in some, but not in all of their cells, for example, mosaic or chimeric animals, the transgene can be integrated as a single transgene or as multiple copies such as concatamers, for example, tandems head to head or head to tail tandems.

The transgene can also be selectively introduced and activated in a particular cell type following, for example, the teaching of Lasko et al. (Lasko et al., Proc. Nati, Acad. Sci. USA 89: 6232-6236 (1992)). The regulatory sequences required for such cell-type specific activation will depend on the particular cell type of interest, and will be apparent to those skilled in the art. When it is desired that the polynucleotide transgene be integrated into the chromosomal site of the endogenous gene, the direction to the gene is preferred. In summary, when such a technique is to be used, vectors containing some nucleotide sequences homologous to the endogenous gene are designed for integration purposes, via homologous recombination with chromosomal sequences, within and interrupting the function of the nucleotide sequence of the endogenous gene. The transgene can also be selectively introduced into a particular cell type, thereby inactivating the endogenous gene only in that cell type, following, for example, the teaching of Gu et al. (What's up., Science 265: 103-106 (1994)). The regulatory sequences required for such specific inactivation of the cell type will depend on the particular cell type of interest, and will be apparent to those skilled in the art. In specific preferred embodiments, the IL-21 or IL-22 polynucleotides of the invention can be expressed under the direction of a murine transferrin receptor-promoting construct, thereby restricting expression to the liver of the transgenic animals. In other specific preferred embodiments, the IL-21 or IL-22 polynucleotides of the invention are expressed under the direction of a murine beta-actin promoter construct, thereby effecting ubiquitous expression of the IL-21 or IL-21 polynucleotide. 22 Once the transgenic animals have been generated, the expression of the recombinant gene can be evaluated using standard techniques. The initial selection can be carried out by spotting or Southern blot analysis or PCR techniques to analyze the animal tissues to verify that the integration of the transgene has taken place. The level of mRNA expression of the transgene in the tissues of the transgenic animals can also be evaluated using techniques that include, but are not limited to, Northern blot analysis of tissue samples obtained from the animal, hybridization analysis in itself tu, and reverse transcriptase PCR (rt-PCR) and real-time PCR "TaqMAN". Transgenic tissue samples that express the genes can also be evaluated immunocytochemically or immunohistochemically using antibodies specific for the transgenic product. Once the founding animals are produced, they can be multiplied, multiplied inbreeding, multiplied by mixing with other races or multiplied by crosses to produce colonies of the particular animal. Examples of such multiplication strategies include, but are not limited to: multiplication by crosses with other breeds of founder animals, with more than one integration site in order to establish separate lines; the inbreeding crossing of separate lines in order to produce transgenic compounds that express the transgene at higher levels due to the effects of the additive expression of each transgene; the crossing of heterozygous transgenic animals to produce animals homozygous for a given integration site, in order to increase expression and eliminate the need for animal selection by DNA analysis; crosses it with separate homozygous lines to produce the heterozygous compound or the homozygous lines; and multiplication to place the transgene on a different antecedent that is appropriate for an experimental model of interest. The transgenic and "inactivated" animals of the invention have uses that include, but are not limited to, systems in animal models useful in the elaboration of the biological function of the IL-21 and / or IL-22 polypeptides, the study of the conditions and / or disorders associated with the aberrant expression of IL-21 and / or IL-22, and in the selection for effective compounds in the amelioration of such conditions and / or disorders. In further embodiments of the invention, cells that are genetically engineered to express the polypeptides of the invention, or alternatively, which are genetically engineered to not express the polypeptides of the invention (e.g., inactivations) are administered to a patient in vi. Such cells may be obtained from the patient (e.g., animal, including humans) or a compatible donor in MHC, and may include, but are not limited to, fibroblasts, bone marrow cells, blood cells (e.g., lymphocytes) , adipocytes, muscle cells, endothelial cells, etc. The cells are engineered by in vitro genetic engineering, using recombinant DNA techniques to introduce the coding sequence of the polypeptides of the invention into the cells, or alternatively, to interrupt the coding sequence and / or the associated endogenous regulatory sequence. with the polypeptides of the invention, for example, by transduction (using viral vectors, and preferably vectors that integrate the transgene within the genome of the cells), or transfection methods, including, but not limited to, the use of plasmids, cosmids , YACs, naked DNA, electroporation, liposomes, etc. The coding sequence of the polypeptides of the invention can be placed under the control of a strong or inducible constitutive promoter or a promoter / enhancer to achieve expression, and preferably secretion, of the polypeptides of the invention. Genetically engineered cells that express and preferably secrete the polypeptides of the invention can be introduced into the patient systemically, for example, in the circulation, or intraperitoneally. Alternatively, the cells can be incorporated into a matrix and implanted in the body, for example, engineered fibroblasts can be implanted as part of a skin graft; Endothelial cells manipulated by genetic engineering can be implanted as part of a lymphatic or vascular graft. (See, for example, Anderson, et al., U.S. Patent No. 5,399,349, and Mulligan &Wilson, U.S. Patent No. 5,460,959 each of which is incorporated by reference herein in its entirety) . When the cells to be administered are non-autologous or cells not compatible with MHC, they can be administered using well-known techniques that prevent the development of a host immune response against the introduced cells. For example, the cells can be introduced in an encapsulated form which, while allowing an exchange of the components with the immediate extracellular environment, does not allow the introduced cells to be recognized by the host's immune system.

Epitopes and Antibodies In the present invention "epitopes" refers to fragments of IL-21 and IL-22 polypeptides having antigenic or immunogenic activity in an animal, especially in a human. A preferred embodiment of the present invention relates to an IL-21 or IL-22 polypeptide fragment comprising an epitope, as well as the polynucleotide encoding this fragment. A region of a protein molecule to which an antibody can be linked is defined as an "antigenic epitope". In contrast, an "immunogenic epitope" is defined as a part of a protein that promotes an antibody response (see, for example, Geysen, et al., Proc. Na ti. Acad. Sci USA 81: 3998-4002 ( 1983)). Fragments that function as epitopes can be produced by any conventional means (see, for example, Houghten, RA, Proc. Na ti, Acad. Sci. USA 82: 5131-5135 (1985), further described in the U.S. Pat. No. 4, 631, 211). In the present invention, the antigenic epitopes preferably contain a sequence of at least seven, more preferably at least nine, and most preferably between about 15 to about 30 amino acids. Antigenic epitopes are useful for producing antibodies, including monoclonal antibodies, which specifically bind to the epitope (see, for example, Wilson, et al., CeJl 37: 767-778 (1984); Sutcliffe, J: G. Et al. , Sci en 219: 660-666 (1983)). Similarly, immunogenic epitopes can be used to induce antibodies according to methods well known in the art (see, for example, Sutcliffe, et al., Supra; Wilson, et al., Supra; Chow, M., et. al., Proc Na ti, Acad. Sci. USA 82: 910-914, and Bittle, FJ et al., J. Gen. Vi.R.:66: 2347-2354 (1985)). A preferred immunogenic epitope includes the secreted protein. Immunogenic epitopes can be presented together with a carrier protein, such as an albumin, to an animal system (such as rabbit or mouse), or if it is long enough (at least about 25 amino acids), without a carrier. However, immunogenic epitopes comprising as few as 8 to 10 amino acids have been shown to be sufficient to produce antibodies capable of binding to at least linear epitopes on a denatured polypeptide (eg, staining or Western blotting). Using DNAstar analysis, it was found that SEQ. ID. NO: 2 is immunogenic in the amino acids: from about Arg-2 to about Pro-11, from about Cys-24 to about Glu-22, and from about Arg-51 to about Gly-59. Thus, these regions can be used as epitopes to produce antibodies against the protein encoded by HTGED19. Again, using the ADNstar analysis, it was found that the SEQ. ID. NO: 4 is immunogenic at the amino acids: from about Gly-19 to about Ala-27, from about Pro-30 to about Arg-38, from about Phe-40 to about Ser-48, from about Tyr-58 to about Leu -67, from about Pro-105 to about Val-113, from about Pro-129 to about Ser-137, from about Asn-139 to about Ala-147, and from about Leu-151 to about Gly-159. In this way, these regions can be used as epitopes to produce antibodies against the protein encoded by HFPBX96. As used herein, the term "antibody" (Ab) or "monoclonal antibody" (Mab) is understood to include the intact molecules as well as the antibody fragments (such as, for example, the Fab and F fragments). ') 2) which are capable of binding specifically to the protein. The Fab and F (ab ') 2 fragments lack the Fe fragment of the intact antibody, clear more quickly from the circulation, and may have less non-specific tissue binding than an intact antibody (Wahl, et al., J. Nucí. Med. 24: 316-325 (1983)). Thus, these fragments are preferred, as well as the products of a FAB or other immunoglobulin expression library. In addition, the antibodies of the present invention include chimeric, single chain and humanized antibodies. The present invention further relates to antibodies to T cell antigen (TCR) receptors that specifically bind to the polypeptides of the present invention. Antibodies of the present invention include IgG (including IgG1, IgG2, IgG3 and IgG4), IgA (including IgA1 and IgA2), IgD, IgE, or IgM, and IgY. As used herein, the term "antibody" (Ab) is understood to include whole antibodies, including single chain full antibodies, and antigen binding fragments thereof. More preferably, antibodies that are antibody fragments that bind to human antigens, of the present invention include, but are not limited to, Fab, Fab 'and F (ab') 2, Fd, single chain Fvs (scFv) , single chain antibodies, disulfide-linked Fvs (sdFv) and fragments comprising either a VL or a VH domain. The antibodies can be of any animal origin including birds and mammals. Preferably, the antibodies are human, murine, rabbit, goat, guinea pig, camel, horse, or chicken. Antibody fragments that bind to the antigen, including single chain antibodies, may comprise the variable regions or regions alone or in combination with the full or partial of the following: the hinge region, the CH1, CH2, and CH3 domains . Also included in the invention are any combinations of one or more variable regions and the hinge region, the CH1, CH2, and CH3 domains. The present invention further includes chimeric, humanized, and human monoclonal and polyclonal antibodies, which specifically bind to the polypeptides of the present invention. The present invention further includes antibodies that are anti-idiotypic to the antibodies of the present invention. The antibodies of the present invention may be monospecific, biscope, wild, trichispecific, or of greater or specificity. The multispecific antibodies may be specific for different epitopes of a polypeptide of the present invention or may be specific for a polypeptide of the present invention as well as for heterologous compositions, such as a heterologous polypeptide or the solid support material. See, for example WO 93/17715; WO 92/08802; WO 91/00360; WO 92/05793; Tutt, A. Et al. (1991) J. Iimmuno1. 147: 60-69; U.S. Patents Nos. 5,573,920, 4,474,893, 5,601,819, 4,714,681, 4,925,648; Kostelny, S.A. et al. (1992) J. Im unol. 148: 1547-1553. The antibodies of the present invention can be described or specified in terms of the epitope or portions of a polypeptide of the present invention, which are recognized or specifically bound by the antibody. The epitope (s) or the polypeptide portion (s) can be specified as described herein, for example, by the N-terminal and C-terminal positions, by size in the contiguous amino acid residues, or listed in the Tables and in the figures. Antibodies that bind specifically to any epitope or polypeptide of the present invention may also be excluded. Therefore, the present invention includes antibodies that specifically bind to the polypeptides of the present invention, and allow the exclusion thereof. The antibodies of the present invention can also be described or specified in terms of their cross-reactivity. Antibodies that do not bind to any other analog, ortholog, or homologue of the polypeptides of the present invention are included. Antibodies that do not bind to polypeptides with less than 95%, less than 90%, less than 85%, less than 80%, less than 75%, less than 70%, less than 65%, less than 60%, less than 55%, and less than 50% identity (as calculated using methods known in the art and described herein) to a polypeptide of the present invention, are also included in the present invention. Also included in the present invention are antibodies that only bind to polypeptides encoded by polynucleotides that hybridize to a polynucleotide of the present invention, under stringent hybridization conditions (as described herein). The antibodies of the present invention may also be described or specified in terms of their binding affinity. Preferred binding affinities include those with a dissociation constant or Kd less than 5X10_6M, 10"6M, 5X10" 7M, 10"7M, 5X10_8M, 10" 8M, 5X10"9M, 10" 9M, 5X10"10M, 10 ~ 10M, 5X10_11M, 10_11M, 5X10"12M, 10" 12M, 5X10"13M, 10" 13M, 5X10"14M, 10'14M, 5X10_15M and 10_15M. The antibodies of the present invention have uses that include, but are not limited to, methods known in the art for purifying, detecting, and directing the polypeptides of the present invention, including diagnostic and therapeutic methods in vi and in vi tro. For example, antibodies have use in immunoassays to qualitatively and quantitatively measure the levels of the polypeptides of the present invention in biological samples. See, for example, Harlow et al., ANTIBODIES: A LABORATORY MANUAL (Cold Spring Harbor Laboratory Press, 2nd ed. 1988) (incorporated by reference in its entirety). The antibodies of the present invention can be used either alone or in combination with other compositions. The antibodies can also be recombinantly fused to a heterologous polypeptide at the N or C terminus or chemically conjugated (including covalent and non-covalent conjugations) to the polypeptides or to other compositions. For example, the antibodies of the present invention may be recombinantly fused or conjugated to molecules useful as labels in detection assays and effector molecules such as heterologous polypeptides, drugs, or toxins. See, for example, WO 92/08495; WO 91/14438; WO 89/12624; U.S. Patent No. 5,314,995; and EP-0, 396, 387. The antibodies of the present invention can be prepared by any suitable method known in the art. For example, a polypeptide of the present invention or an antigenic fragment thereof can be administered to an animal for the purpose of inducing the production of sera containing polyclonal antibodies. Monoclonal antibodies can be prepared using a range of techniques known in the art, including the use of hybridoma technology and recombinant technology. See, for example, Harlow et al., ANTIBODIES: A LABORATORY MANUAL, (Cold Spring Harbor Laboratory Press, 2nd ed. 1988); Hammerling et al., In: MONOCLONAL ANTIBODIES AND T-CELL HYBRIDOMAS 563-681 (Elsevier, N.Y., 1981) (these references are incorporated by reference in their entirety). The antibodies of the present invention can be prepared by any of a variety of standard methods. For example, cells expressing the IL-21 and / or IL-22 polypeptide or an antigenic fragment thereof can be administered to an animal for the purpose of inducing the production of sera containing polyclonal antibodies. In a preferred method, a preparation of IL-21 and / or IL-22 polypeptide is carried out and purified to render it substantially free of natural contaminants. Such preparation is then introduced into an animal in order to produce polyclonal antisera of higher specific activity. In the most preferred method, the antibodies of the present invention are monoclonal antibodies (or fragments thereof that bind to the IL-21 and / or IL-22 polypeptide). Such monoclonal antibodies can be prepared using a hybridoma technology (Kohler et al., Na ture 256: 495 (1975), Kohler et al., Eur. J. Immunol. 6: 511 (1976), Kohler et al., Eur. J. Immunol., 6: 292 (1976), Hammerling et al., In: Monoclonal Anal tibodi and T-Cell Hybri domas, Elsevier, NY (1981) pp 563-681). In general, such methods involve immunizing an animal (preferably a mouse) with an IL-21 and / or IL-22 polypeptide antigen or, more preferably, with a cell expressing the IL-21 and / or IL- 21 polypeptide. 22 Suitable cells can be recognized for their ability to bind to the anti-IL-21 polypeptide antibody and / or anti-IL-22 polypeptide. Such cells can be cultured in any suitable medium for tissue culture; however, it is preferable to culture the cells in Earle's modified Eagle's medium supplemented with 10% fetal bovine serum (inactivated at approximately 56 ° C) and supplemented with approximately 10 g / liter of non-essential amino acids, approximately 1000 units / ml of penicillin, and approximately 100 μg / ml of streptomycin. The splenocytes of such mice are extracted and fused with a suitable myeloma cell line. Any suitable myeloma cell line can be employed in accordance with the present invention. However, it is preferable to use the progenitor myeloma cell line (SP20), available from ATCC, Manassas, Virginia. After fusion, the resulting hybridoma cells are selectively maintained in HAT medium, and then cloned by limiting dilution as described by Wands, et al. (Ga s troen t erol ogy 80: 225-232 (1981)). Hybridoma cells obtained through such selection are then evaluated to identify the clones that secrete the antibodies capable of binding to the IL-21 and / or IL-22 antigen. Alternatively, additional antibodies, capable of binding to the IL-21 and / or IL-22 polypeptide antigen, can be produced in a two-step procedure through the use of anti-idiotypic antibodies. Such a method makes use of the fact that the antibodies are themselves antigens, and that, therefore, it is possible to obtain an antibody that binds to a second antibody. According to this method, antibodies specific for IL-21 and / or IL-22 polypeptides are used to immunize an animal, preferably a mouse. The splenocytes of such an animal are then used to produce hybridoma cells, and the hybridoma cells are selected to identify the clones that produce an antibody whose ability to bind to the specific antibody of the IL-21 and / or IL-22 polypeptide can be blocked by the antibody. antigen IL-21 and / or IL-22. Such antibodies comprise anti-idotypic antibodies to the IL-21 and / or IL-22 polypeptide-specific antibody, and can be used to immunize an animal to induce the formation of additional antibodies specific for IL-21 and / or IL-21 polypeptides. 22 Fab and F (ab ') 2 fragments can be produced by proteolytic cleavage, using enzymes such as papain (to produce Fab fragments) or pepsin (to produce fragments F (ab ') 2). Alternatively, the antibodies of the present invention can be produced through the application of recombinant DNA technology or through synthetic chemistry using methods known in the art. For example, antibodies of the present invention can be prepared using various methods of phage display known in the art. In phage display methods, functional antibody domains are displayed on the surface of a phage particle that possesses the polynucleotide sequences that encode it. The phage with a desired binding property is selected from a repertoire or a library of antibodies in combination (eg, human or murine) by selection directly with antigen, typically antigen bound or captured to a solid surface or sphere. The phages used in these experiments are typically filamentous phage including fd and M13 with fab, Fv or disulfide stabilized Fv antibody domains, recombinantly fused to either the phage III gene or the gene VIII protein. Examples of phage display methods that can be used to make the antibodies of the present invention, include those described in Brinkman U. et al (1995) J. Immunol. Methods 182: 41-50; Ames, R.S. et al (1995) J. Immuno. Methods 185: 177-186; Kettleborough, C.A. et al. (1994) Eur. J. Im unol. 24: 952-958; Persic. L. et al. (1995.}. 7) Gene 187 9-18; Burton, D.R. et al. (1994) Advances in Immunology 57: 191-280; PCT / GB91 / 01134; WO 90/02809; WO 91/10737; WO 92/01047; WO 92/18619; WO 93/11236; WO 95/15982; WO 95/20401; and U.S. Patents Nos. 5,698,426, 5,223,409, 5,403,484, 5,580,717, 5,427,908, 5,750,753, 5,821,047, 5,571,698, 5,427,908, 5,516,637, 5,780,225, 5,658,727 and 5,733,743 (these references are incorporated herein by reference in their entirety). As described in the above references, after phage selection, the antibody coding regions from the phage can be isolated and used to generate the complete antibodies, including human antibodies, or any other desired antigen binding fragment, and expressed in any desired host including mammalian cells, insect cells, plant cells, yeast, and bacteria. For example, techniques for recombinantly producing Fab, Fab 'and F (ab') 2 fragments can also be employed using methods known in the art such as those described in WO 92/22324; Mullinax, R.L. et al. BioTechniques 12 (6): 864-869 (1992); and Sawai, H. et al. AJRI 34: 26-34 (1995); and Better, M. et al. Science 240: 1041-1043 (1988) (said references are incorporated by reference in their entirety). Examples of techniques that can be used to produce single chain Fvs (scFvs) and antibodies, include those described in U.S. Patent Nos. 4,946,778 and 5,258,498; Huston et al. Methods in Enzymology 203: 46-88 (1991); Shu L. et al. PNAS 90: 7995-7999 (1993); and Skerra, A. Et al. Science 240: 1038-1040 (1988). For some uses, including the in vivo use of antibodies in humans and in vitro detection assays, it may be preferable to use chimeric, humanized, or human antibodies. Methods for producing chimeric antibodies are known in the art. See, for example, Morrison, Science 229: 1202 (1985); Oi et al, BioTechniques 4: 214 (1986); Gillies, S.D. et al., J. Immunol. Methods 125: 191-202 (1989); and U.S. Patent No. 5,807,715. The antibodies can be humanized using a variety of techniques including CDR grafts (EP-0, 239, 400; WO 91/09967; US Patent No. 5,530,101 and 5,585,089), surface coating or refinishing (European Patent EP -0, 592, 106; EP-0, 519, 596; Padlan, EA, Molecular Immunology 28 (4/5): 489-498 (1991); Studnicka G.M. et al., Protein Engineering 7 (6): 805-814 (1994); Roguska M.A. et al., PNAS 91: 969-973) (1994), and interspersed with chains (U.S. Patent No. 5,565,332). Human antibodies can be made by a variety of methods known in the art including the phage display methods described above. See also U.S. Patent Nos. 4,444,887, 4,716,111, 5,545,806, and 5,814,318; and WO 98/46645 (said references are incorporated by reference in their entirety). Also included in the present invention are recombinantly fused or chemically conjugated antibodies (including covalent and non-covalent conjugations) to a polypeptide of the present invention. The antibodies may be specific for antigens other than the polypeptides of the present invention. For example, the antibodies can be used to direct the polypeptides of the present invention to particular cell types, either in vi tro or in vi ve by fusion or conjugation of the polypeptides of the present invention to the specific antibodies for particular receptors of the cell surface. Antibodies fused or conjugated to the polypeptides of the present invention can also be used in immunoassays and in methods of purification using methods known in the art. See, for example, Harbor et al. Supra and WO 93/21232; EP-0, 439, 095; Naramura, M. et al., Immunol. Lett. 39: 91-99 (1994); U.S. Patent No. 5,474,981; Gillies, S.O. et al. PNAS 89: 1428-1432 (1992); Fell, H.P. et al., J.

Immunol. 146: 2446-2452 (1991) (said references are incorporated by reference in their entirety). The present invention further includes compositions comprising the polypeptides of the present invention fused or conjugated to the antibody domains other than the variable regions. For example, the polypeptides of the present invention can be fused or conjugated to an Fe region of the antibody, or a portion thereof. The antibody portion fused to a polypeptide of the present invention may comprise the hinge region, the CH1 domain, the CH2 domain and the CH3 domain or any combination of the entire domains or portions thereof. The polypeptides of the present invention can be fused or conjugated to the above antibody portions, to increase the half-life of the polypeptides or to be used in immunoassays using methods known in the art. The polypeptides can also be fused or conjugated to the aforementioned antibody portions, to form multimers. For example, Fe portions fused to the polypeptides of the present invention can form dimers through disulfide bonds between Fe portions. Higher multimer forms can be made by fusing the polypeptides to IgA and IgM portions. Methods for fusing or conjugating the polypeptides of the present invention to the antibody portions are known in the art. See, for example, U.S. Patent Nos. 5,336,603, 5,622,929, 5,359,046, 5,349,053, 5,447,851, 5,112,946; EP-0, 307, 434, EP-0, 367, 166; WO 96/04388, WO 91/06570; Ashkenazi, A. et al., PNAS 88: 10535-10539 (1991); Zheng, X.X. et al., J. Immunol. 154: 5590-5600 (1995); and Vil, H. Et al., PNAS 89: 11337-11341 (1992) (said references are incorporated by reference in their entirety). The invention also relates to antibodies that act as agonists or antagonists of the polypeptides of the present invention. For example, the present invention includes antibodies that disrupt receptor / ligand interactions with the polypeptides of the invention either partially or totally. These include receptor-specific antibodies and ligand-specific antibodies. They include receptor-specific antibodies which do not prevent binding to the ligand but prevent the activation of the receptor. Activation of the receptor (eg, signaling) can be determined by techniques described herein or otherwise known in the art. Also included are receptor-specific antibodies which prevent ligand binding and receptor activation. Likewise, neutralizing antibodies that bind to the ligand and prevent ligand binding to the receptor are included, as well as antibodies that bind to the ligand, thereby preventing receptor activation, but the ligand is not prevented. It is linked to the receiver. The antibodies that activate the receptor are also included. These antibodies can act as agonists for all or less than all the biological activities affected by receptor activation mediated by the ligand. The antibodies can be specified as agonists or antagonists for the biological activities comprising specific activities described herein. The above antibody agonists can be made using methods known in the art. See, for example, WO 96/40281; U.S. Patent No. 5,811,097; Deng, B. et al., Blood 92 (6): 1981-1988 (1998); Chen, Z. Et al., Cancer REs. 58 (16): 3668-3678 (1998); Harrop, J.A. et al., J. Immunol. 161 (4): 1786-1794 (1988); Zhu, Z. Et al., Cancer Res. 58 (15): 3209-3214 (1998); Ion, D.Y. et al., J. Immunol. 160 (7): 3170-3179 (1998); Prat, M. et al., J. Cell. Sci. 111 (Pt2): 237-247 (1998); Pitard, V. et al., J. Iimmuno1. Methods 205 (2): 177-190 (1997); Liautard, J. et al., Cytokinde 9 (4): 233-241 (1997); Carlson, N.G. et al., J. Biol. Chem. 272 (17): 11295-11301 (1997); Taryman, R.E. et al., Neuron 14 (4): 755-762 (1995); Muller, Y. A. et al., Structure 6 (9): 1153-1167 (1998); Bartunek, P. et al., Cytokine 8 (1): 14-20 (1996) (said references are incorporated by reference in their entirety). As discussed above, antibodies to the IL-21 and / or IL-22 polypeptides can, in turn, be used to generate anti-idiotype antibodies that "mimic" IL-21 and / or IL-22, using techniques well known to those skilled in the art. (See, for example, Greenspan &Bona, FASEB J. 7 (5): 437-444 (1989), and Nissinoff, J. Immunol. 147 (8): 2429-2438 (1991)). For example, antibodies that bind to IL-21 and / or IL-22 and competitively inhibit the binding of IL-21 and / or IL-22 to the receptor, can be used to generate anti-idiotypes that "mimic" the linkage domain to IL-21 and / or IL-22 and, as a consequence, bind to and neutralize IL-21 and / or IL-22 and / or its receptor. Such neutralizing anti-idiotypes or Fab fragments of such anti-idiotypes can be used in therapeutic regimens to neutralize IL-21 and / or IL-22 ligands.

Fusion proteins Any IL-21 or IL-22 polypeptide can be used to generate fusion proteins. For example, IL-21 or IL-22 polypeptides, when fused to a second protein, can be used as an antigenic tag. Antibodies raised against the IL-21 or IL-22 polypeptides can be used to indirectly detect a second protein by binding to IL-21 or IL-22, respectively. In addition, because the secreted proteins are directed to cell sites based on the traffic signals, the IL-21 and IL-22 polypeptides can be used as targeting molecules once fused to other proteins.

Examples of domains that can be fused to the IL-21 and IL-22 polypeptides include not only heterologous signal sequs, but also other heterologous functional regions. The fusion does not necessarily need to be direct, but can occur through linker sequs. In addition, the fusion proteins can also be engineered to improve the characteristics of the IL-21 and IL-22 polypeptides. For example, a region of additional amino acids, particularly charged amino acids, can be added to the N-terminus of the IL-21 and IL-22 polypeptides to improve stability and persist during purification from the host cell or during handling and subsequent storage. Also, peptide portions can be added to the IL-21 and IL-22 polypeptides to facilitate purification. Such regions can be removed before the final preparation of polypeptides 11-21 and IL-22. The addition of peptide portions to facilitate the handling of the polypeptides are familiar and routine techniques in the art. In addition, IL-21 and IL-22 polypeptides, including fragments, and specifically epitopes, can be combined with constant domain portions of immunoglobulins (IgG), resulting in chimeric polypeptides. These fusion proteins facilitate purification and show an increased half-life. One reported example describes chimeric proteins consisting of the first two domains of the human CD-4 polypeptide and various domains of the constant regions of the heavy or light chains of mammalian immunoglobulins (European Patent EP-A 394,827; Traunecker et al. , Na t ure 331: 84-86 (1988)). Fusion proteins that have dimeric structures linked by disulfide bridges (due to IgG) may also be more efficient in binding and neutralizing other molecules than the secreted monomeric protein or the protein fragment alone (Fountoulakis et al., J. Bi. Echem 270: 3958-3964 (1995)). Similarly, European Patent EP-A-0,464,533 (Canadian counterpart 2045869) describes the fusion proteins comprising various portions of the constant region of the immunoglobulin molecules together with another human protein or part thereof. In many cases, the Fe part in a fusion protein is beneficial in * therapy and in diagnosis, and can thus result, for example, in improved pharmacokinetic properties (EP-A-0, 232, 262). Alternatively, deletion of the Fe part after the fusion protein has been expressed, detected and purified could be desired. For example, the Fe moiety can prevent therapy and diagnosis if the fusion protein is used as an antigen for immunizations. In drug discovery, for example, human proteins, such as hIL-5, have been fused with Fe portions for purposes of high throughput screening assays to identify hIL-5 antagonists (see, Bennett, D. et al. , J. Mol. Recog. 8: 52-58 (1995); Johanson, K., Et al., J. Biol. Ch., 270: 9459-9471 (1995)). In addition, IL-21 and IL-22 polypeptides can be fused to marker sequs, such as a peptide that facilitates the purification of IL-21 and IL-22, respectively. In preferred embodiments, the marker amino acid sequ is a hexa-histidine peptide, such as the label provided in a pQE vector (QIAGEN, Inc., 9259 Eton Avenue, Chatsworth, CA, 91311), among others, many of the which are commercially available.

As described by Gentz et al. (Proc.Nat.Acad.Sci. USA 86: 821-824 (1989)), for example, hexahistidine provides convenient purification of the fusion protein. Another useful peptide tag for purification, the "HA" tag, corresponds to an epitope derived from the influenza hemagglutinin protein (Wilson et al., Cell 37: 767 (1984)). In further preferred embodiments, the IL-21 or IL-22 polynucleotides of the invention are fused to a polynucleotide that encodes a "FLAG" polypeptide. Thus, an IL-21-FLAG or IL-22-FLAG fusion protein is encompassed by the present invention. The FLAG antigenic polypeptide can be fused to an IL-21 or IL-22 polypeptide of the invention at either the amino or carboxyl terminus. In preferred embodiments, an IL-21-FLAG or IL-22-FLAG fusion protein is expressed from an expression vector pFLAG-CMV-5a or pFLAG-CMV-1 (available from Sigma, Saint Louis Missouri, USA) . See, Anderson, S., et al., J. Bi ol. . Ch em. 264: 8222-29 (1989); Tho sen, D.R. et al., Proc. Na ti. Aca d. Sci. USA, 81: 659-63 (1984); and Kozak M., Na t ure 308: 241 (1984) (each of which is incorporated by reference herein).

In additional preferred embodiments, an IL-21-FLAG or IL-22-FLAG fusion protein is detectable by anti-FLAG monoclonal antibodies (also available from Sigma). Thus, any of the above fusion proteins can be engineered using the IL-21 and / or IL-22 polynucleotides or the polypeptides of the invention.

Vectors, Host Cells, and Production Proteins The present invention also relates to the vectors containing the IL-21 and IL-22 polynucleotides, the host cells, and the production of polypeptides by recombinant techniques. The vector can be, for example, a phage, plasmid, viral or retroviral vector. Retroviral vectors may be replication competent or replication defective. In the latter case, viral propagation will generally occur only in the complement of host cells. The polynucleotides IL-21 and IL-22 can be linked to a vector that contains a selectable marker for propagation in a host. In general, a plasmid vector is introduced into a precipitate, such as a calcium phosphate precipitate, or into a complex with a charged lipid. If the vector is a virus, it can be packaged using a suitable packaging cell line and then transduced into the host cells. The polynucleotide inserts IL-21 and IL-22 must be operably linked to an appropriate promoter, such as the PL promoter of lambda phage, the E promoters. coli lac, trp, phoA and tac, the early and late promoters of SV40 and the promoters of the retroviral LTRs, to name a few. Other suitable promoters will be known to the person skilled in the art. The expression constructs will also contain sites for the initiation, termination of transcription, and, in the transcribed region, a binding site to the ribosome for translation. The coding portion of the transcripts expressed by the constructs will preferably include a start codon of translation at the beginning, and a stop codon (UAA, UGA or UAG) appropriately located at the end of the polypeptide to be translated.

As indicated, the expression vectors will preferably include at least one selectable marker. Such markers include dihydrofolate reductase, G418 or neomycin resistance for the culture of eukaryotic cells and the tetracycline, kanamycin or ampicillin resistance genes for E culture. col i and other bacteria. Representative examples of suitable hosts include, but are not limited to, bacterial cells, such as E cells. col i, from Streptomyomyces, and from Salmon el l a typhimuri um; fungal cells, such as yeast cells; insect cells such as Drosophila S2 and Spodoptera Sf9 cells; animal cells such as CHO, COS, 293, and Bowes melanoma cells; and plant cells. Suitable culture media and conditions for the host cells described above are known in the art. Preferred vectors for use in bacteria include pHE4-5 and other vectors similar to pHE; pQE70; pQE60 and pQUE-9 available from QIAGEN, Inc .; pBluescript vectors, Phagescript vectors, pNH8A, pNH16a, pNH18A, available from Stratagene Cloning Systems, Inc .; and ptrc99a, pKK223-3, pKK233-3, pDR540, pRIT5 available from Pharmacia Biotech, Inc. Among the preferred eukaryotic vectors are pWLNEO, pSV2CAT, pOG44, pXT1 and pSDG available from Stratagene; and pSVK3, pBPV, pMSG and pSVL available from Pharmacia. Other suitable vectors will be readily apparent to those skilled in the art. The introduction of the construct into the host cell can be effected by calcium phosphate transfection, DEAE-dextran mediated transfection, cationic lipid-mediated transfection, electroporation, transduction, infection, or other methods. Such methods are described in many standard laboratory manuals (eg Davis et al., Ba si c Me thods In Mol ec ul ar Bi olgy (1986)). It is specifically contemplated that IL-21 and IL-22 polypeptides can, in fact, be expressed by a host cell lacking a recombinant vector. The IL-21 and IL-22 polypeptides can be recovered and purified from recombinant cell cultures by well-known methods including ammonium sulfate or ethanol precipitation, acid extraction, anionic or cation exchange chromatography, phosphocellulose chromatography, chromatography of hydrophobic interaction, affinity chromatography, hydroxyapatite chromatography and lectin chromatography. More preferably, high performance liquid chromatography ("HPLC") is used for the purification. The IL-21 and IL-22 polypeptides, and preferably the secreted forms thereof, can also be recovered from: the products purified from natural sources, including body fluids, tissues and cells, whether directly isolated or cultured; products of chemical synthesis procedures; and products produced by recombinant techniques from prokaryotic or eukaryotic hosts, including, for example, bacterial, yeast, higher plant, insect, and mammalian cells. Depending on the host employed in a recombinant production process, polypeptides 11-21 and IL-22 can be glycosylated or can be non-glycosylated. In addition, polypeptides 11-21 and IL-22 may also include a modified, initial methionine residue, in some cases as a result of the host-mediated processes. Thus, it is well known in the art that the N-terminal methionine encoded by the start codon of translation will generally be removed with high efficiency from any protein after translation in all eukaryotic cells. While N-terminal methionine in most proteins is also effectively removed in most prokaryotes, for some proteins, this process of prokaryotic elimination is inefficient, depending on the nature of the amino acid to which the N-terminal methionine is covalently linked.

Use of IL-21 and IL-22 Polynucleotides The IL-21 and IL-22 polynucleotides identified herein can be used in numerous ways as reagents. The following description should be considered plary and use known techniques. There is a need to come to identify new chromosomal markers, since few chromosome labeling reagents are currently available, based on data from the effective sequence (repeating polymorphisms). The HTGED19 clone and the HFPBX96 clone can each be mapped to a specific chromosome. Thus, the IL-21 and IL-22 polynucleotides can then be used in linkage analysis as a marker for those specific chromosomes. In summary, the sequences can be mapped or mapped to the chromosomes by preparation of PCR primers (preferably 15 to 25 base pairs) from the sequences shown in SEQ. ID. NO: 1, SEQ. ID. NO: 3, SEQ. ID. NO: 28 and SEQ. ID. NO: 31. The primers can be selected using computer analysis so that the primers do not span more than one predicted exon in the genomic DNA. These primers are then used for the selection by PCR of somatic cell hybrids containing individual human chromosomes. Only those hybrids containing the human IL-21 or IL-22 genes corresponding to SEQ. ID. NO: 1, SEQ. ID. NO: 3, SEQ. ID. NO: 28 or SEQ. ID. NO: 31, respectively, will produce a fragment to pli ficado. Similarly, somatic hybrids provide a rapid method of PCR mapping of polynucleotides to particular chromosomes. Three or more clones can be assigned per day using a simple thermal cycler. In addition, the sub-localization of polynucleotides IL-21 and IL-22 can be achieved with panels of specific chromosomal fragments. Other gene map mapping strategies that can be used include i n si t u hybridization, pre-selection with flow-labeled chromosomes, and pre-selection by hybridization to construct chromosome-specific cDNA libraries. The precise chromosomal location of the polynucleotides IL-21 and IL-22 can also be achieved using fluorescence hybridization i n si t u (FISH) or chromosomal multiplication in metaphase. This technique uses polynucleotides as short as 500 or 600 bases; however, polynucleotides of 2,000 to 4,000 base pairs are preferred (Fara a review, see Verma et al., "Human Chromosomes: a Manual or Basic Techniques", Pergamon Press, New York (1988)). For the chromosomal mapping, polynucleotides IL-21 and IL-22 can be used individually (to mark a simple chromosome or a simple site on that chromosome) or in panels (for the marking of multiple sites and / or multiple chromosomes) . The preferred polynucleotides correspond to the non-coding regions of the cDNAs, because the coding sequences are more likely to be conserved within gene families, thereby increasing the opportunity for cross-hybridization during chromosome mapping. In a preferred embodiment, the gene encoding IL-22 of the present invention has been mapped using FISH technology to a site on human chromosome 13 at position 13qll. In addition, the gene coding for IL-21 of the present invention has been mapped to a site on human chromosome 7. See also, Example 4 infra. Once a polynucleotide has been mapped to an accurate chromosomal site, the physical position of the polynucleotide can be used in linkage analysis. The linkage analysis establishes the coherence between a chromosomal location and the presentation of a particular disease (the data of the map map of the disease are found, for example, in McKusick, V., Mendelian Inheritance in Man (available online through of Johns Hopkins University Welch Medical Library)). Assuming a mapped resolution of 1 megabase and one gene per 20 kb, a cDNA located precisely to a chromosomal region associated with the disease, could be one of the 50-500 potential causal genes. Thus, once coherence is established, differences in IL-21 and IL-22 polynucleotides and corresponding genes between affected and unaffected individuals can be examined. First, visible structural alterations in chromosomes, such as deletions or translocations, are examined in chromosome disseminations by PCR. If there are no structural alterations, the presence of point mutations is evaluated. The mutations observed in some or all affected individuals, but not in normal individuals, indicate that the mutation may cause the disease. However, complete sequencing of IL-21 and IL-22 polypeptides and corresponding genes from several normal individuals is required to distinguish the mutation from a polymorphism. If a new polymorphism is identified, this polymorphic polypeptide can be used for the analysis of additional binding. In addition, increased or decreased expression of the gene in affected individuals compared to unaffected individuals can be evaluated using polynucleotides IL-21 and IL-22. Any of these alterations (altered expression, chromosomal rearrangement, or mutation) can be used as a diagnostic or prognostic marker. In addition to the above, the IL-21 or IL-22 polynucleotide can be used for the control of gene expression through triple helix formation or antisense DNA or RNA. Both methods rely on the binding of the polynucleotide to DNA or RNA. For these techniques, the preferred polynucleotides are usually 20 to 40 bases in length and complementary to either the region of the gene involved in the transcription (triple helix - see Lee et al., Nucí. Aci ds Res. 6: 3073 (1979); Cooney et al., Sci in ce 241: 456 (1988); and Dervan et al., Sci in ce 251: 1360 (1991)) or to the mRNA itself (antisense - Okano, J. Ne uroch em 56: 560 (1991); Oligodeoxy-nucleotides as Antisense Inhibitors of Gene Expression, CRC Press, Boca 1 ~ t ^ SH & FaR Raton, Florida (1988)). The formation of the triple helix optimally results in a quenching of the transcription of RNA from the DNA, while the hybridization of the antisense RNA blocks the translation of a mRNA molecule in the polypeptide. Both techniques are effective in model systems, and the information described herein can be used to design the antisense or triple helix polynucleotides in an effort to treat the disease. The polynucleotides IL-21 and IL-22 are also useful in gene therapy. One goal of gene therapy is to insert a normal gene into an organism that has a defective gene, in an effort to correct the genetic defect. IL-21 and IL-22 offer means to address such genetic defects in a highly accurate manner. Another goal is to insert a new gene that was not present in the host's genome, thereby producing a new trait in the host cell. Polynucleotides IL-21 and IL-22 are also useful for identifying individuals from minute biological samples. The United States military, for example, is considering the use of restriction fragment length polymorphism (RFLP) for the identification of its personnel. In this technique, an individual genomic DNA is digested with one or more restriction enzymes, and probed in a Southern stain to produce unique bands for personnel identification. This method does not suffer from the current limitations of "Dog Tags" ("Dog Tags") that can be lost, changed or stolen, making positive identification difficult. The polynucleotides IL-21 and IL-22 can be used as additional DNA markers for RFLP. The polynucleotides IL-21 and IL-22 can also be used as an alternative for RFLP, by determining the base DNA sequence by base, effective from the selected portions of a genome of an individual. These sequences can be used to prepare PCR primers for the amplification and isolation of such selected DNA, which can then be sequenced. Using this technique, individuals can be identified because each individual will have a unique group of DNA sequences. Once a unique identity database is established for an individual, positive identification of that individual, living or dead, can be made from extremely small tissue samples. Forensic biology also benefits from the use of DNA-based identification techniques as described herein. DNA sequences taken from very small biological samples such as tissues, for example, hair or skin, or body fluids, for example, blood, saliva, semen, etc., can be amplified using PCR. In a prior art method, gene sequences amplified from polymorphic loci, such as the HLA DQa gene of class II, are used in forensic biology to identify individuals (Erlich, H., PCR Technol ogy, Freeman and Co. (1992)). Once these specific polymorphic loci are amplified, they are digested with one or more restriction enzymes, producing a group of identification bands in a spot or Southern blot probed with the DNA corresponding to the HLA gene of DQa of class II. Similarly, polynucleotides IL-21 and IL-22 can be used as polymorphic markers for forensic purposes.

There is also a need for reagents capable of identifying the source of a particular tissue. Such a need arises, for example, in forensic practice when presented with tissue of unknown origin. Suitable reagents may comprise, for example, DNA probes or primers specific for particular tissue prepared from the sequences of IL-21 and IL-22. The panels of such reagents can identify tissue by species and / or by type of organ. In a similar way, these reagents can be used to select tissue cultures for contamination. Because IL-21 is found to be expressed almost exclusively in apoptotic T cells, the IL-21 polynucleotides are useful as hybridization probes for the differential identification of the or tissues and / or the type or types of cells present in a biological sample. Similarly, polypeptides and antibodies directed to the IL-21 polypeptides are useful for providing immunological probes for the differential identification of the or of the tissues or of the cell types. In addition, for a number of disorders of the aforementioned tissues or cells, particularly of the immune system, significantly higher or lower levels of IL-21 gene expression can be detected in certain tissues (eg, cancerous and injured tissues) or in bodily fluids (e.g., in serum, plasma, urine, synovial fluid or spinal fluid) taken from an individual having such a disorder, relative to a level of expression of the "standard" IL-21 gene, e.g. expression level of 11-21 in healthy tissue from an individual who does not have the immune system disorder. Similarly, since IL-21 is found expressed in the bone marrow, in skeletal muscle and in the brain, IL-22 polynucleotides are useful as hybridization probes for the differential identification of the or tissues or of the the cell types present in a biological sample. Similarly, polypeptides and antibodies directed to IL-22 polypeptides are useful for providing immunological probes for the differential identification of the or tissues or of the cell types. In addition, for a number of disorders of the aforementioned tissues or cells, particularly of the immune system, significantly higher or lower levels of IL-22 gene expression can be detected in certain tissues (e.g., cancerous and injured tissues) or in bodily fluids (e.g., in serum, plasma, urine, synovial fluid or spinal fluid), taken from an individual having such a disorder, relative to a level of expression of the "standard" IL-22 gene, for example, the level of expression of IL-22 in the healthy tissue of an individual who does not have the immune system disorder. Thus, the invention provides a method of diagnosing a disorder, which involves: (a) evaluating the level of expression of the IL-21 or IL-22 gene in cells or in the body fluid of an individual; (b) comparing the expression level of the IL-21 or IL1-22 gene with an expression level of the standard IL-21 or IL-22 gene, respectively, thereby increasing or decreasing the level of expression of the IL-21 or IL-22 gene evaluated, compared to the level of standard expression, is an indicator of the immune system disorder. At least, IL-21 and IL-22 polynucleotides can be used as molecular weight markers on Southern gels, as diagnostic probes for the presence of a specific mRNA in a particular cell type, as a probe to "subtract" sequences known in the process of discovering novel polynucleotides, for the selection and elaboration of oligomers for coupling to a "piece or gene chip" or other support, to produce anti-DNA antibodies using DNA immunization techniques, and as an antigen to promote an immune response.

Uses of IL-21 and IL-22 Polypeptides Polypeptides 11-21 and IL-22 can be used in numerous ways. The following description should be considered exemplary and use known techniques. IL-21 and IL-22 products can be used to evaluate protein levels in a biological sample using antibody-based techniques. For example, protein expression in tissues can be studied with classical immunohistochemical methods (Jalkanen, M., et al., J. Cell 1. Biol. 101: 976-985 (1985); Jalkanen, M., et. to J. Cell L. Biol. 105: 3087-3096 (1987)). Other methods based on antibodies useful for the detection of protein gene expression include immunoassays, such as enzyme-linked immunosorbent assay (ELISA) and radioimmunoassay (RIA). Suitable antibody assay labels are known in the art and include enzymatic labels, such as, for example, glucose oxidase, and radioisotopes, such as iodine (125 I, 121 I), carbon (14 C), sulfur (35 S), tritium. (3H), indium (112In), and technetium (99mTc), and fluorescent labels such as fluorescein and rhodamine, and biotin. In addition to evaluating protein levels secreted in a biological sample, proteins can also be detected by image formation. Antibody labels or markers for the imaging of the protein include those detectable by X-ray radiography, NMR or ESR. For X-ray radiography, suitable markers include radioisotopes such as barium or cesium, which emit detectable radiation but are not completely harmful to the subject. Suitable markers for NMR and ESR include those with a detectable characteristic spin, such as deuterium, which can be incorporated into the antibody by labeling the nutrients for the relevant hybridoma. An antibody or antibody fragment specific for the protein, which has been labeled with a detectable, appropriate imaging portion, such as a radioisotope (eg, 131I, 112In, 99mTc), a radiopaque substance, or a material detectable by nuclear magnetic resonance, it is introduced (eg, parenterally, subcutaneously or intraperitoneally) into the mammal. It will be understood in the art that the size of the subject and the image-forming system used will determine the amount of image-forming portion necessary to produce diagnostic images. In the case of a radioisotope portion, for a human subject, the amount of radioactivity injected will normally be in the range of about 5 to 20 millicuries of 99mTc. The antibody or labeled antibody fragment will then accumulate preferentially at the site of the cells containing the specific protein. Tumor imaging in vi vo is described by Burchiel et al. ("Immunopharmacokinetics of Radiolabeled Antibodies and Their Fragments" (Chapter 13 in Tumor Imaging: The Radiochemical Detection of Cancer, S. Burchiel and B.A. Rhodes, eds., Masson Publishing Inc. (1982)). Thus, the present invention provides a method of diagnosis of a disorder, which involves (a) the evaluation of the expression of the IL-21 or IL-22 polypeptides in cells or in body fluid of an individual; (b) comparing the expression level of IL-21 or IL-22 with a standard gene expression level, whereby an increase or decrease in the expression level of the IL-21 or IL-22 polypeptide gene evaluated, in comparison to the level of standard expression, it is an indicator of a disorder. In addition, the IL-21 and IL-22 polypeptides can be used to treat the disease. For example, patients can be administered with IL-21 and IL-22 polypeptides in an effort to replace the absent or decreased levels of the IL-21 and IL-22 polypeptides, respectively (e.g., insulin), to supplement absent levels. or decreased of a different polypeptide (e.g., hemoglobin S for hemoglobin B), to inhibit the activity of a polypeptide (e.g., an oncogene), to activate the activity of a polypeptide (e.g., by binding to a receptor), to reduce activity of a membrane-bound receptor by competing with it for the free ligand (eg, the soluble TNF receptors used in reducing inflammation), or to give rise to a desired response (e.g., growth of blood vessels) ). Similarly, antibodies directed to the IL-21 and IL-22 polypeptides can also be used to treat the disease. For example, the administration of an antibody directed to a polypeptide IL-21 and IL-22 can bind and reduce the overproduction of the polypeptide. Similarly, administration of an antibody can activate the polypeptide, such as by binding to a membrane-bound polypeptide (receptor). At least, the IL-21 and IL-22 polypeptides can be used as molecular weight markers on SDS-PAGE gels or on molecular sieve gel filtration columns, using methods well known to those skilled in the art. The IL-21 and IL-22 polypeptides can also be used to produce antibodies, which, in turn, are used to measure the expression of proteins from a recombinant cell, as a way to evaluate the transformation of the host cell. In addition, IL-21 and IL-22 polypeptides can be used to test the following biological activities.

Biological Activities of IL-21 and IL-22 IL-21 polynucleotides and polypeptides and IL-22 can be used in assays to test one or more biological activities. If the IL-21 and IL-22 polynucleotides or polypeptides do show activity in a particular assay, it is likely that IL-21 and IL-22 may be involved in diseases associated with biological activity. Therefore, IL-21 and IL-22 could be used to treat the associated disease. The IL-21 and IL-22 proteins of the present invention modulate the secretion of IL-6 from NIH-3T3 cells. An ELISA in vi tro assay that quantifies the amount of IL-6 secreted in cells, in response to treatment with cytokines or soluble extracellular domains of cytokine receptors, has also been described (Yao, Z., et al. , Immuni ty 3: 811-821 (1995)). In summary, the assay involves plating the target cells at a density of approximately 5 x 10 6 cells / ml in a volume of 500 μl in the wells of a 24-well flat bottom culture plate (Costar). The cultures are then treated with various concentrations of the cytokine or soluble extracellular domain of the cytokine receptor in question. The cells are then cultured for 24 hours at 37 ° C. At this time, 50 μl of supernatants are removed and evaluated for the amount of IL-6 essentially as described by the manufacturer (Genzyme, Boston, MA). The levels of IL-6 are then calculated by reference to a standard curve constructed with the recombinant IL-17 cytokine. Such activity is useful for the determination of the level of secretion of IL-6 mediated by IL-21 or IL-22. The IL-21 and IL-22 protein modulates the proliferation of cells of the immune system and differentiation in a dose-dependent manner in the assay described above. Thus, "a polypeptide having IL-21 or IL-22 protein activity" includes polypeptides that also exhibit any of the same stimulatory activities in the above-described assays in a dose-dependent manner. Although the degree of dose-dependent activity need not be identical to that of the IL-21 or IL-22 proteins, preferably, "a polypeptide having IL-21 or IL-22 protein activity" will show dose dependence substantially similar in a given activity compared to the IL-21 or IL-22 protein (e.g., the candidate polypeptide will show greater activity or no more than about times less, and preferably, no more than about ten times less activity relative to the reference protein IL-21 or IL-22). Lymphocyte proliferation is another in vitro assay which can be performed to determine the activity of IL-21 and IL-22. For example, Yao et al. (Immuni ty 3: 811-821 (1995)) have recently described an in vitro assay for the determination of the effects of various cytokines and soluble cytokine receptors on the proliferation of murine leukocytes. In summary, the lymphoid organs are aseptically harvested, the lymphocytes are isolated from the harvested organs, and the resulting collection of lymphoid cells is suspended in standard culture medium as described by Fanslow et al. (J. Immunol., 147: 535-5540 (1991)). Suspensions of lymphoid cells can then be divided into several different subclasses of lymphoid cells including splenic T cells, B cells of lymph nodes, CD4 + and CD8 + T cells, and mature adult thymocytes. For splenic T cells, the suspensions of spleen cells (200 x 10 6 cells) are incubated with the monoclonal antibody (mAb) CDllb and the MHC class II mAb for 30 minutes at 4 ° C, loaded onto a purification column of T cells (Pierce, Rockford, IL) and T cells are eluted according to the manufacturer's instructions. Using this method, the purity of the resulting T cell populations should be > 95% CD3 + and < 1% of sIgM +. For the purification of subgroups of lymph nodes, B cells are removed by adhesion to tissue culture dishes previously coated with goat anti-mouse IgG (10 μg / ml). The remaining cells were then incubated with anti-CD4 or anti-CD8 for 30 minutes at 4 ° C, and then washed and placed on tissue culture dishes previously coated with a goat anti-rat IgG (20 μg / ml). After 45 minutes, the non-adherent cells are removed and tested for purity by flow cytometry. CD4 cells and depleted in superficial Ig should be > 90% TCR-ab, CD8 +, while CD8 and cells depleted in surface IgG must be more than 95% TCR-ab, CD4 +. Finally, to enrich mature adult titi, cells are suspended at 108 / ml in 10% anti-HSA and 10% rabbit complement of low toxicity (Cedarlane, Ontario, Canada), incubated for 45 minutes at 37 ° C, and the remaining viable cells are isolated on Ficoll-Hypaque (Pharmacia, Piscataway, NJ). This procedure should produce between 90 and 95% of CD3hl cells that are either CD4 + 8"or CD4" 8+.

Immune activity The polypeptides or polynucleotides IL-21 and IL-22 may be useful in the treatment of deficiencies or disorders in the immune system, by activation or inhibition of the proliferation, differentiation, or mobilization (chemotaxis) of immune cells. Immune cells develop through a process called hematopoiesis, producing myeloid cells (platelets, red blood cells, neutrophils and macrophages) and lymphoid cells (B and T lymphocytes) from cells of the pluripotential line. The etiology of these immune deficiencies or immune disorders can be genetic, somatic, such as cancer or some autoimmune disorders, acquired (for example, chemotherapy or toxins), or infectious. In addition, polynucleotides or polypeptides IL-21 and IL-22 can be used as a marker or detector of a particular disease or disorder of the immune system. The polynucleotides or polypeptides IL-21 and IL-22 may be useful in the treatment or detection of deficiencies * or disorders of hematopoietic cells. The IL-21 and IL-22 polypeptides or polynucleotides could be used to increase the differentiation and proliferation of hematopoietic cells, including stem cells, in an effort to treat those disorders associated with a decrease in certain (or many) types of hematopoietic cells. . Examples of syndromes for 'immune deficiency include, but are not limited to: blood protein disorders (eg agammaglobulinemia, disgammaglobulinemia), telangiectasia due to ataxia, common variable immunodeficiency, Digeorge syndrome, HIV infection, HTLV-BLV infection, leukocyte adhesion deficiency syndrome, lymphopenia , bactericidal dysfunction due to phagocytes, severe combined immunodeficiency (SCIDs), Wiskot-Aldrich Disorder, anemia, thrombocytopenia, or hemoglobinuria. In addition, IL-21 and IL-22 polypeptides or polynucleotides can also be used to modulate hemostatic activity (the arrest of bleeding) or thrombolytic activity (clot formation). For example, by increasing hemostatic or thrombolytic activity, IL-21 and IL-22 polynucleotides or polypeptides could be used to treat blood coagulation disorders (e.g., afibrinogenemia, factor deficiencies), platelet disorders. blood (for example, thrombocytopenia), or injuries resulting from trauma, surgery or other causes. Alternatively, IL-21 and IL-22 polynucleotides or polypeptides that can decrease hemostatic or thrombolytic activity could be used to inhibit or dissolve clots, important in the treatment of heart attacks (infarction), strokes or scarring.

The IL-21 and IL-22 polynucleotides or polypeptides may also be useful in the treatment or detection of autoimmune disorders. Many autoimmune disorders result from the inappropriate recognition of own material as foreign by immune cells. This inappropriate recognition results in an immune response that leads to the destruction of host tissue. Therefore, the administration of IL-21 and IL-22 polypeptides or polynucleotides that can inhibit an immune response, particularly the proliferation, differentiation, or chemotaxis of T cells, can be an effective therapy in the prevention of autoimmune disorders. Examples of autoimmune disorders that can be treated or detected by IL-21 and IL-22 include, but are not limited to: Addison's disease, • hemolytic anemia, antiphospholipid syndrome, rheumatoid arthritis, dermatitis, allergic encephalomyelitis, glomerulonephritis, Goodpasture, Graves' Disease, Multiple Sclerosis, Myasthenia Gravis, Neuritis, Ophthalmia, Billerous Pemphigoid, Pemphigoid, Polyendocrinopathies, Purple, Reiter's Disease, Stiff-Man Syndrome, Autoimmune Thyroiditis, Systemic Lupus Erythematosus, Autoimmune Pulmonary Inflammation, Guillain Syndrome Barre, insulin-dependent diabetes mellitus, and autoimmune inflammatory ophthalmic disease. Similarly, allergic reactions, such as asthma (particularly allergic asthma) or other respiratory problems, can also be treated by the polypeptides or polynucleotides IL-21 and IL-22. In addition, IL-21 and IL-22 can be used to treat anaphylaxis, hypersensitivity to an antigenic molecule, or blood group incompatibility. The IL-21 and IL-22 polynucleotides or polypeptides can also be used to treat and / or prevent organ rejection or graft versus host disease (GVHD). Organ rejection occurs by the destruction of host immune cells from the transplanted tissue through an immune response. Similarly, an immune response is also involved in GVHD, but, in this case, foreign, transplanted immune cells destroy host tissues. The administration of polypeptides or polynucleotides IL-21 and IL-22 that inhibit the immune response, particularly the proliferation, differentiation or chemotaxis of T cells, can be an effective therapy in the production of organ rejection or GVHD. Similarly, IL-21 and IL-22 polypeptides or polynucleotides can also be used to modulate inflammation. For example, the polypeptides or polynucleotides IL-21 and IL-22 can inhibit the proliferation and differentiation of cells involved in an inflammatory response. These molecules can be used to treat inflammatory conditions, chronic and acute conditions, including inflammation associated with infection (for example, septic shock, sepsis, or systemic inflammatory response syndrome (SIRS)), ischemia-reperfusion injury, endotoxin lethality , arthritis, complement-mediated hyperacute rejection, nephritis, cytokine-induced or chemokine-induced lung damage, inflammatory bowel disease, Cronhn's disease, or resulting from overproduction of cytokines (eg, TNF or IL-1).

Hyperproliferative disorders The IL-21 and IL-22 polypeptides or polynucleotides can be used to treat or detect hyperproliferative disorders, including neoplasms. The IL-21 and IL-22 polypeptides or polynucleotides can inhibit the proliferation of the disorder through direct or indirect interactions. Alternatively, IL-21 and IL-22 polypeptides or polynucleotides can proliferate other cells that can inhibit hyperproliferative disorder. For example, by increasing an immune response, particularly the increase in the antigenic qualities of the hyperproliferative disorder or by the proliferation, differentiation or mobilization of T cells, hyperproliferative disorders can be treated. This immune response can be increased either by increasing an existing immune response, or by initiating a new immune response. Alternatively, the decrease in an immune response may also be a method for the treatment of hyperproliferative disorders, such as a chemotherapeutic agent.

Examples of hyperproliferative disorders that can be treated or detected by IL-21 and IL-22 polynucleotides or polypeptides include, but are not limited to, neoplasms located in: the abdomen, bone, breast, digestive system, liver , the pancreas, the peritoneum, the endocrine glands (adrenal, parathyroid, pituitary, testes, ovaries, thymus, thyroid), the eye, the head and neck, the nervous system (central and peripheral), the lymphatic system, the pelvis, the skin, the soft tissue, the spleen, the thoracic and urogenital cavities. Similarly, other hyperproliferative disorders can also be treated or detected by IL-21 and IL-22 polynucleotides or polypeptides. Examples of such hyperproliferative disorders include, but are not limited to: hypergammaglobulinemia, lymphoproliferative disorders, paraproteinemias, purpura, sarcoidosis, Sezary's syndrome, Waldenstron's macroglobulinemia, Gaucher's disease, histiocytosis and any other hyperproliferative disease, in addition to neoplasia, localized in an organ system listed above.

Infectious Disease The IL-21 and IL-22 polypeptides or polynucleotides can be used to treat or detect infectious agents. For example, by increasing the immune response, particularly by increasing the proliferation and differentiation of B and / or T cells, infectious diseases can be treated. The immune response can be increased either by increasing an existing immune response, or by initiating a new immune response. Alternatively, the polypeptides or polynucleotides IL-21 and IL-22 can also directly inhibit the infectious agent, without necessarily promoting an immune response. Viruses are an example of an infectious agent that can cause disease or symptoms that can be treated or detected by IL-21 and IL-22 polynucleotides or polypeptides. Examples of viruses include, but are not limited to the following families of DNA- and RNA-viruses: Arboviruses, Adenoviidae, Arenaviridae, Arterivirus, Birnaviridae, Bunyaviridae, Caliciviridae, Circoviridae, Coronaviridae, Flaviviridae, Hepadnaviridae (Hepatitis), Herpesviridae (such such as Cytomegalovirus, Herpes Simplex, Herpes Zoster), Mononegavirus (eg, Parmyxoviridae, Morbillivirus, Rhabdoviridae), Orthomyxoviridae (eg, Influenza), Papovaviridae, Parvoviridae, Picornaviridae, Poxviridae (such as Smallpox or Vaccinia), Reoviridae (eg, Rotavirus), Retroviridae (HTLV-I, HTLV-II, Lentivirus), and Togaviridae (for example, Rubivirus). Viruses that fall within these families can cause a variety of diseases or symptoms, including, but not limited to: arthritis, bronchiolitis, encephalitis, ophthalmic infections (eg, conjunctivitis, keratitis), chronic fatigue syndrome, hepatitis (A, B, C, E, Active Chronic, Delta), meningitis, opportunistic infections (eg, AIDS), pneumonia, Burkitt's lymphoma, chickenpox, hemorrhagic fever, measles, mumps, parainfluenza, rabies, the common cold, polio, leukemia, Rubella, sexually transmitted diseases, skin diseases (eg, Kaposi, warts), and viremia. The IL-21 and IL-22 polypeptides or polynucleotides can be used to treat or detect any of these symptoms or diseases.

Similarly, bacterial or fungal agents that can cause disease or symptoms and that can be treated or detected by IL-21 and IL-22 polynucleotides or polypeptides include, but are not limited to, the following families of Gram-negative and Gram-positive and fungi: Actinomycetales (eg, Corynebacterium, Mycobacterium, Norcardia), Asperillosis, Bacillaceae (eg, Anthrax, Clostridium), Bacteroidaceae, Blasto icosis, Bordetella, Borrelia, Brucellosis, Candidiasis, Campylobacter, Coccidioidomycosis, Cryptococcosis, Dermatocicosis , Enterobacteriaceae (Klebsiella, Salmonella, Serratia, Yersinia), Erysipelothrix, Helicobacter, Legionellosis, Leptospirosis, Listeria, Mycoplasmatales, Neisseriaceae (for example, Acinetobacter, Gonorrhea, Menigococco), Pasteurellacea Infections (for example, Actinobacillus, Heamophilus, Pasteurella), Pseudomonas, Rickettsiaceae, Chlamydiaceae, Syphilis and Staphylococcus. These families of bacteria or fungi can cause the following diseases or symptoms, including, but not limited to: bacteremia, endocarditis, ophthalmic infections (conjunctivitis, tuberculosis, uveitis), gingivitis, opportunistic infections (for example, AIDS-related infections), paronychia , infections related to the prosthesis, Reiter's disease, respiratory tract infections, such as whooping cough or empyema, sepsis, Lyme disease, cat itch disease, dysentery, paratyphoid fever, food poisoning, typhoid, pneumonia, gonorrhea , meningitis, chlamydia, syphilis, diphtheria, leprosy, paratuberculosis, tuberculosis, lupus, botulism, gangrene, tetanus, impetigo, rheumatic fever, scarlet fever, sexually transmitted diseases, skin diseases (eg, cellulitis, dermatocicosis), toxemia, urinary tract infections, wound infections. The IL-21 and IL-22 polypeptides or polynucleotides can be used to treat or detect any of these symptoms or diseases. In addition, parasitic agents that cause disease or symptoms that can be treated or detected by IL-21 polynucleotides or polypeptides include, but are not limited to, the following families: Amibiasis, Babesiosis, Coccidiosis, Cryptosporidiosis, Dientamoebiasis, Durina, Ectoparasitosis, Giardiasis , Helminthiasis, Leish aniasis, Theileriasis, Toxoplasmosis, Trypanosomiasis, and Tricomonia-sis. These parasites can cause a variety of diseases or symptoms, including, but not limited to: scabies, thrombiculiasis, eye infections, intestinal disease (e.g., dysentery, giardiasis), liver disease, lung disease, opportunistic infections (e.g. related to AIDS), Malaria, complications of pregnancy and toxoplasmosis. The IL-21 and IL-22 polypeptides or polynucleotides can be used to treat or detect any of these symptoms or diseases. Preferably, the treatment using polypeptides or polynucleotides IL-21 and IL-22 could be either by administering an effective amount of the IL-21 or IL-22 polypeptide to the patient, or by withdrawing the patient's cells, supplying the cells with polynucleotide IL-21 and IL-22 and returning the cells engineered to the patient (ex vivo therapy). In addition, the IL-21 and IL-22 polypeptide or polynucleotide can be used as an antigen in a vaccine to promote an immune response against the infectious disease.

Regeneration Polynucleotides or polypeptides IL-21 and IL-22 can be used to differentiate, proliferate and attract cells, leading to tissue regeneration (see, Sci ence 276: 59-87 (1997)). The regeneration of tissues could be used to repair, replace or protect tissue damaged by congenital defects, trauma (wounds, burns, incisions, or ulcers), age-related diseases (eg, osteoporosis, osteocarditis, periodontal disease, liver failure), surgery, including cosmetic plastic surgery, fibrosis, reperfusion injury, or systemic cytokine damage. The tissues that could be regenerated using the present invention include organs (eg, pancreas, liver, intestine, kidney, skin, endothelium), muscle tissue (smooth, skeletal or cardiac), vascular tissue (including vascular endothelium), nervous, hematopoietic and skeletal tissue (bone, cartilage, tendon and ligaments) .

Preferably, regeneration occurs with diminished healing. Regeneration may also include angiogenesis. In addition, IL-21 and IL-22 polynucleotides or polypeptides can increase the regeneration of difficult-to-heal tissues. For example, increased regeneration of the tendons / ligaments could accelerate the recovery time after damage. The IL-21 and IL-22 polynucleotides or polypeptides of the present invention could also be used prophylactically in an effort to prevent damage. Specific diseases that could be treated include tendonitis, carpal tunnel syndrome, and other tendon or ligament defects. A further example of tissue regeneration from wounds that do not heal includes pressure ulcers, ulcers associated with vascular insufficiency, surgical or traumatic wounds. Similarly, nerve and brain tissue could also be regenerated by using the polynucleotides or polypeptides IL-21 and IL-22 to proliferate and differentiate nerve cells. Diseases that could be treated using this method include diseases of the central and peripheral nervous system, neuropathies or mechanical and traumatic disorders (for example, disorders of the spine, cranial trauma, cerebrovascular disease and stroke). Specifically, diseases associated with peripheral nerve damage, peripheral neuropathy (eg, resulting from chemotherapy and other medical therapies), localized neuropathies, and diseases of the central nervous system (eg, Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, and Shy-Drager syndrome), they could all be treated using the IL-21 and IL-22 polynucleotides or polypeptides.

Chemotherapy The IL-21 and IL-22 polynucleotides or polypeptides can have chemotaxis activity. A chemotactic molecule attracts or mobilizes cells (e.g., monocytes, fibroblasts, neutrophils, T cells, mast cells, eosinophils, epithelial and / or endothelial cells) to a particular site in the body, such as inflammation, infection or the site of hyperproliferation. The mobilized cells can then fight against and / or heal the particular trauma or particular abnormality. Polynucleotides or polypeptides IL-21 and IL-22 can increase the chemotactic activity of particular cells. These chemotactic molecules can then be used to treat inflammation, infection, hyperproliferative disorders, or any disorder of the immune system by increasing the number of cells directed to a particular site in the body. For example, chemotactic molecules can be used to treat wounds and other trauma to tissues by attracting immune cells to the damaged site. As a chemotactic molecule, IL-21 and IL-22 could also attract fibroblasts, which can be used to treat wounds. It is also contemplated that polynucleotides or polypeptides IL-21 and IL-22 can inhibit chemotactic activity. These molecules could also be used to treat disorders. Thus, IL-21 and IL-22 polynucleotides or polypeptides could be used as an inhibitor of chemotaxis.

Link Activity The IL-21 and IL-22 polypeptides can be used to select the molecules that bind to IL-21 or IL-22 or to the molecules to which IL-21 or IL-22 bind. The binding of IL-21 and IL-22 and the molecule can activate (agonist), increase, inhibit (antagonist), or decrease the activity of IL-21 and IL-22 or the linked molecule. Examples of such molecules include antibodies, oligonucleotides, proteins (e.g., receptors), or small molecules. Preferably, the molecule is closely related to the natural ligand of IL-21 or IL-22, eg, a fragment of the ligand, or a natural substrate, a ligand, a structural or functional mimetic (see, Coligan et al., Current Protocol in Immunology 1 (2): Chapter 5 (1991)). Similarly, the molecule may be closely related to the natural receptor to which IL-21 and IL-22 bind, or at least, a fragment of the receptor capable of being linked by IL-21 or IL-22 (eg, the active site). ). In any case, the molecule can be rationally designed using known techniques.

Preferably, selection for these molecules involves the production of appropriate cells expressing IL-21 and IL-22, either as a secreted protein or on the cell membrane. Preferred cells include cells from mammals, yeasts, Drosophila or E. coli Cells expressing IL-21 and IL-22 (or the cell membrane containing the expressed polypeptide) are then preferably contacted with a test compound that potentially contains the molecule to observe binding, stimulation or inhibition of IL activity. -21 and IL-22 or of the molecule. The assay can simply test for the binding of a candidate compound to IL-21 or IL-22, where the binding is detected by a marker, or in an assay that involves competition with a labeled competitor. In addition, the assay can test whether the candidate compound results in a signal generated by the binding to IL-21 or IL-22. Alternatively, the assay can be carried out using cell-free preparations, polypeptide / molecule attached to a solid support, chemical libraries, or mixtures of natural products. The assay may also simply comprise the steps of mixing a candidate compound with a solution containing IL-21 or IL-22, measuring the IL-21 activity / linkage / molecule or IL-22 / molecule, respectively, and comparing the activity or the binding of IL-21 / molecule or IL-22 / molecule, to a standard. Preferably, an ELISA assay can measure the ls of IL-21 and IL-22 or activities in a sample (eg, the biological sample) using a monoclonal or polyclonal antibody. The antibody can measure the ls of IL-21 and IL-22 or the activities either by binding, directly or indirectly, to IL-21 or IL-22 or by competing with IL-21 or IL-22 for a substrate . All these aforementioned assays can be used as diagnostic or prognostic markers. The molecules discovered using these assays can be used to treat the disease or to give rise to a particular result in a patient (e.g., the dopment of blood vessels) by activating or inhibiting IL-21 or IL-22. In addition, the assays can discover agents that can inhibit or increase the. production of IL-21 or IL-22 from properly manipulated cells or tissues. Therefore, the invention includes a method for identifying compounds that bind to IL-21 and IL-22 comprising the steps of: a) incubating a candidate binding compound with IL-21 or IL-22; and b) the determination of whether the link has occurred. In addition, the invention includes a method for identifying agonists / antagonists, comprising the steps of: a) incubating a candidate compound with IL-21 or IL-22, b) evaluating a biological activity, and c) determining whether a biological activity of IL-21 or IL-22, respectively, has been altered.

Other activities The polypeptides or polynucleotides IL-21 and IL-22 can also increase or decrease the differentiation or proliferation of totipotential embryonic cells, in addition, as discussed above, the hematopoietic line. The polypeptides or polynucleotides IL-21 and IL-22 can also be used to modulate the characteristics of mammals, such as body height and weight, hair color, eye color, skin, percentage of adipose tissue, pigmentation, size and shape (advantageously, cosmetic surgery). Similarly, IL-21 and IL-22 polypeptides or polynucleotides can be used to modulate mammalian metabolism that affects catabolism, anabolism, processing, utilization and energy storage. The polypeptides or polynucleotides IL-21 and IL-22 can be used to change a mental state or physical state of the mammal by influencing biorhythms, heart rate, circadian rhythms, depression (including depressive disorders), the tendency for violence, tolerance for pain (in preferred modalities, analyzed by a pain test in the hyperalgesic foot, rat), reproductive capacities (preferably by activity similar to Activin or Inhibin), hormonal or endocrine levels, appetite, libido disorders, of memory, stress and other cognitive qualities. The IL-21 and IL-22 polypeptides or polynucleotides can also be used as an additive or preservative for foods, such as to increase or decrease the storage capacities, the content of fat, lipids, protein, carbohydrates, vitamins. , of minerals, cofactors or other nutritional components. Having generally described the invention, it will be more readily understood by reference to the following examples, which are provided by way of illustration and are not intended to be limiting.

EXAMPLES In the case where full length IL-21 and partial IL-22 are not specifically mentioned, the specific details of the following examples are given only for the partial length IL-21 molecules of the present invention. However, the examples can be easily performed for full-length IL-21 and the full-length or partial-length IL-22 molecules of the present invention, by using the details provided for partial IL-21 and substituting the nucleotides or appropriate amino acid residues of full-length IL-21, full length or partial length IL-22, and / or any deletion mutations or other variants of IL-21 or IL-22, for example, in the design of suitable PCR primers, and the like. The use or applicability of another IL-21 or IL-22 in place of the IL-21 exemplified below, is thus contemplated in each of the following examples. When provided with the nucleotide and amino acid sequences of IL-21 (SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 28 and SEQ ID NO: 29) and IL -22 (SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 30 and SEQ ID NO: 31) of the present invention, a person of ordinary skill in the art could easily carrying out the following examples with the intention of isolating or characterizing or further manipulating another IL-21 or IL-22 in place of the IL-21 shown in the following Examples.

Ex empl o 1: A i sl ami of the IL-21 and IL-22 cDNA Clones From the Samples Deposited The cDNAs encoding the partial IL-21 and IL-22 molecules are each inserted into the restriction sites co coRI and Xiiol of the multiple cloning site of pBluescript. pBluescript contains a gene for resistance to ampicillin and can be transformed into E. col i strain DH10B, available from Life Technologies (see, for example, Gruber, CE. et al., Focus 15: 59 (1993)). Two procedures can be used to isolate IL-21 from the deposited sample. First, a specific polynucleotide of SEQ. ID. NO: 1 with 30-40 nucleotides is synthesized using an Applied Biosystems DNA synthesizer according to the reported sequence. The oligonucleotide is labeled, for example, with 32p-gamma-ATP using the kinase of the T4 polynucleotide and purified according to routine methods (eg, Maniatis, et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Press, Cold Spring, NY (1982)). The plasmid mixture is transformed into a suitable host (such as XL-1 Blue (Stratagene)) using techniques known to those skilled in the art, such as those provided by the vector provider or in related publications or patents. The transformants are seeded on plates, on 1.5% agar plates (containing the appropriate selection agent, for example ampicillin) at a density of approximately 150 transformants (colonies) per plate. These plates are selected using Nylon membranes according to routine methods for the selection of bacterial colonies (eg, Sambrook, et al., Molecular Cloning: A Laboratory Manual, 2nd Ed., (1989), Cold Spring Harbor Laboratory Press , pages 1.93 or 1.104), or other techniques known to those skilled in the art. Alternatively, two primers of 17-20 nucleotides derived from both ends of SEQ. ID. NO: 1 (for example, within the region of SEQ ID NO: 1 limited by the 5 'and 3' nucleotides of the clone) are synthesized and used to amplify the IL-21 cDNA using the deposited cDNA plasmid , as a template. The polymerase chain reaction is carried out under routine conditions, for example, in 25 microliters of the reaction mixture with 0.5 micrograms of the previous template cDNA. A convenient reaction mixture is 1.5-5 mM magnesium chloride, 0.01% (w / v) gelatin, 20 micromolar each of dATP, dCTP, dGTP, dTTP, 25 pmol of each primer and 0.25 Taq polymerase units. Thirty-five cycles of PCR (denaturation at 94 ° C for 1 minute; annealing at 55 ° C for 1 minute, elongation at 72 ° C for 1 minute) are performed with an Perkin-Elmer Cetus automatic thermal cycler. The amplified product is analyzed by agarose gel electrophoresis and the DNA band with the expected molecular weight is excised and purified. The PCR product is verified to be the selected sequence by subcloning and sequencing the DNA product. Several methods are available for the identification of the 5 'or 3' non-coding portions of the IL-21 gene that may not be present in the deposited clone. These methods include, but are not limited to, filter screening, clone enrichment using the specific probes, and protocols similar or identical to the 5 'and 3' RACE protocols that are well known in the art. For example, a method similar to 5 '-RACE is available for the generation of the missing 5' end of a desired full-length transcript (Fromont-Racine, et al., Nucí Aci s Res. 21 (7): • 1683 -1684 (1993)). Briefly, a specific RNA oligonucleotide is ligated to the 5 'ends of a population of RNA, presumably containing the RNA transcripts of the full-length gene. A group of primers containing a specific primer for the ligated RNA oligonucleotide and a primer specific for a known sequence of the IL-21 gene of interest is used for PCR amplification of the 5 'portion of the IL-21 gene of full length The amplified product can then be sequenced and used to generate the full-length gene. This previous method starts with total DNA isolated from the desired source, although poly-A + RNA can be used. The RNA preparation can then be treated with a phosphatase, if necessary, to remove the 5'-phosphate groups on the degraded or damaged RNA that can interfere with the last step of the RNA-ligase. The phosphatase must then be inactivated and the RNA treated with the tobacco acid phosphatase in order to eliminate the cap structure present at the 5 'ends of the messenger RNA. This reaction leaves a 5 'phosphate group at the 5' end of the RNA cleaved in the cap, which can then be ligated to an RNA oligonucleotide using T4 RNA-ligase. This preparation of the modified RNA is used as a template for the synthesis of the first strand of cDNA using a specific oligonucleotide. The synthesis reaction of the first strand is used as a template for PCR amplification of the desired 5 'end, using a primer specific for the ligated RNA oligonucleotide and a primer specific for the known sequence of the gene of interest. The resulting product is then sequenced and analyzed to confirm that the 5 'end sequence belongs to the IL-21 gene.

Ex empl o 2: A i sl ami on to the Genomic Clusters of IL -21 A human genomic Pl library (Genomic Systems, Inc.) is selected by PCR using the primers selected for the cDNA sequence corresponding to SEQ. ID. NO: 1, according to the method described in Example 1 (see also, Sambrook et al., S upra).

Ex empl o 3: Di s tribuci ón de Tej i dos de IL-21 The tissue distribution of IL-21 mRNA expression is determined using protocols for the Northern blot analysis, described by, among others, Sambrook et al (s upra). For example, an IL-21 probe produced by the method described in Example 1 is labeled with 32P using the Rediprime ™ DNA labeling system (Amersham Life Science), according to the manufacturer's instructions. After labeling, the probe is purified using a CHROMA SPIN-100MR column (Clontech Laboratories, Inc.), according to manufacturer's protocol number. PT1200-1. The labeled, purified probe is then used to screen various human tissues for mRNA expression. Staining or Multiple Tissue Northern blots (MTN), which contain various human tissues (H) or tissues of the human immune system (IM) (Clontech), are examined with the labeled probe using the ExpressHybMR (Clontech) hybridization solution of according to the manufacturer's protocol number PT1190-1. After hybridization and washing, the spots are mounted and exposed to film at -70 ° C overnight, and the films developed according to standard procedures. Using essentially the protocol described above, Northern blot analyzes were used to determine the expression pattern of IL-21 and IL-22. In the case of IL-21, very slight signals of 1.8 and 3.0 kb were detected in the skeletal muscle, and signals of indeterminate sizes were detected in the fetal lung and in the fetal kidney. In the case of IL-22, a message greater than 2.4 kb was detected in conjunction with a larger band in all the brain tissues examined, and was also detected to a lesser extent in the skeletal muscle, in the heart, in the testicles, in the spinal cord, in the bone marrow, in the small intestine, in the kidney and in the lung. A band smaller than 4.4 kb was also detected in the skeletal muscle.

Ex empl o 4: Layout of the Chromosome Map of IL-21 A group of oligonucleotide primers is designed according to the sequence at the 5 'end of the SEQ. ID. NO: 1. This primer preferably comprises about 100 nucleotides. This primer is then used in a polymerase chain reaction under the following group of conditions: 30 seconds, 95 ° C; 1 minute, 56 ° C; 1 minute, 70 ° C. This cycle is repeated 32 times, followed by a 5 minute cycle at 70 ° C. Human, mouse and hamster DNA is used as a template, in addition to a hybrid panel of somatic cells containing individual chromosomes or fragments of chromosomes (Bios, Inc.). The reactions are analyzed either on 8% polyacrylamide genes or on 3.5% agarose gels. The chromosomal mapping is determined by the presence of a PCR fragment of approximately 100 base pairs in the particular somatic cell hybrid.

Example 5: Bacterial expression of IL-21 An IL-21 polynucleotide encoding an IL-21 polypeptide of the invention is amplified using the oligonucleotide PCR primers corresponding to the 5 'and 3' ends of the DNA sequence, as described in Example 1, to synthesize the insertion fragments. The primers used to amplify the cDNA insert should preferably contain restriction sites, such as Bamñl and HindIII, at the 5 'end of the primers in order to clone the amplified product into the expression vector, eg, BamE I and Hi ndIII correspond to the restriction enzyme sites on the bacterial expression vector pQE9 (Qiagen Inc., Chatsworth, CA). This plasmid vector codes for antibiotic resistance (AmpR), a bacterial origin of replication (ori), a promoter / operator regulated by IPTG (P / O), a ribosome binding site (RBS), a label of 6- histidine (6-His) and restriction enzyme cloning sites. Specifically, to clone the mature domain of the IL-21 protein in a bacterial vector, the 5 'primer has the sequence 5'-GAT CGC GGA TCC GAC ACG GAT GAG GAC CGC TAT CCA CAG AAG CTG-3' (SEQ. NO: 9) containing the underlined BamHI restriction site, followed by several nucleotides of the amino-terminal coding sequence of the mature IL-21 sequence in SEQ. ID. NO: 1. A person skilled in the art would appreciate, of course, that the point in the coding sequence of the protein where the 5 'primer starts can be varied to amplify a segment of DNA that codes for any desired portion of the protein. Complete IL-21 protein, shorter or longer than the mature form of the protein. The 3 'primer has the sequence 5' -CCC AAG CTT TCA CAC TGA ACG GGG CAG CAC GCA GGT GCA GC-3 '(SEQ ID NO: 10) containing the underlined HindIII restriction site, followed by a number of nucleotides complementary to the 3 'end of the coding sequence of the DNA sequence of -IL-21 of SEQ. ID. NO: 1. The vector pQE9 is digested with BamHI and Hi ndIII and the amplified fragment is ligated into the pQE9 vector maintaining the reading structure initiated in the bacterial RBS. The ligation mixture is then used to transform E. col i strain M15 / rep4 (Qiagen, Inc.) which contains multiple copies of plasmid pREP4, which expresses the lacl repressor and also confers resistance to kanamycin (KanR). Transformants are identified by their ability to grow on LB plates and colonies that are resistant to ampicillin and kanimycin are selected. The plasmid DNA is isolated and confirmed by restriction analysis. The clones containing the desired constructions are developed overnight (O / N) in liquid culture in LB medium supplemented with Amp. (100 μg / ml) and Kan (25 μg / ml). The cut O / N is used to inoculate a large crop at a ratio of 1: 100 to 1: 250. The cells are developed at an optical density 600 (O.D ^ 60o) of between 0.4 and 0.6. IPTG (isopropyl-β-D-thiogalactopyranoside) is then added to a final concentration of 1 mM. IPTG induces the lacl repressor by inactivation, clearing the promoter / operator, leading to increased expression of the gene. The cells develop for an additional 3 to 4 hours. The cells are then harvested by centrifugation (20 minutes at 6000 x g). The concentrate or cell button is solubilized in the caotxopic agent guanidine-HCl < 5 M to asitax for 3 to 4 hours at 4 ° C. Cell waste is removed by centrifugation, and the supernatant contains the polypeptide loaded onto a nickel-nitri-tri-acetic acid affinity resin column ("Ni-NTA") (QIAGEN, Inc., upra). Proteins with a 6 x His tag bind to Ni-NTA resin with high affinity and can be purified in a simple one step procedure (for details see: The QUIAexprssionist (1995J CUIAGJEN, Inc., s upxa). In summary, the supernatant is loaded onto the column in 6M guanidine-HCJL, pH 8 1 &column is first washed with 10 volumes of 6 M guanidine-HCl, pH 8, then washed with 10 volumes of 6 M guanidine-HCl, pH 6, and finally the polypeptide is eluted with 6 M guanidine-HCl, pH 5. The purified IL-21 protein is then renatured upon dialysis against phosphate buffered saline (PBS) or 50 mM sodium acetate, pH buffer 6 plus 200 mM sodium chloride Alternatively, the IL-21 protein can be successfully refolded while immobilized on the NiNTA column The recommended conditions are as follows: renature using a linear gradient of 6M-1M urea in sodium chloride 500 mM., 20% of glycerol, 20 mM Tris / HCl, pH 7.4, containing protease inhibitors. The renaturation must be done in a period of 1.5 hours or more. After renaturalization, the proteins are fluid by the addition of 250 M imidazole. The imidazole is removed by a final dialysis step against PBS or 50 mM sodium acetate buffer, pH 6, plus sodium chloride .200 ^ The purified JX-_21 proton is stored at 4 ° C or frozen at -80 ° C . In addition to the above expression vector, the present invention further includes an expression vector comprising the phage operator and ros promoter elements operably linked to a polynucleotide 11-21., Called pHE4a (Access ATCC Accession 20965, deposited on February 25, 1998). This vector contains: 1J a neomycin phosphotransferase gene as a selection marker, 2) an origin of replication of j? "Col. , 3) a sequence of the phage promoter T5, 4) two lac operator sequences, a sequence -Shine-Dalgarno and 6) the repressor gene of the lactose operon (laclq).

The origin of replication (oriC) is derived from pUC19 (LT1, Gaithersburjg., JMD). Xa promoter sequence and operator sequence are synthetically elaborated, DNA can be inserted into pHEa by restricting the vector with NdeJ and Xbal BamEI, Xhol or Aspl l S, running the restricted product on a gel and isolating the fragment further. large (the heaviest fragment should also be approximately 310 base pairs). The insert of AD? is generated according to the PCR protocol described in Example 1 and using the PCR primers coding for the restriction sites for Ndel (5 'primer) and Ndel and Xbal., Bam? I ^ Xh olj or Asp718 (3' primer ). The PCR insert is purified in qel and restricted with compatible enzymes. The insert and the vector are ligated according to standard protocols. The genetically engineered vector could easily be substituted in the above protocol to express the protein in a bacterial system.

Example ?; Puxif i cation of the IL-21 Polypeptide From an Inclining Body The following alternative method may be used to purify the IL-21 polypeptide expressed in E, Coli when it is j > resent in the form of inclusion bodies. Unless otherwise specified, all the following steps are conducted at 4-10 ° C. After the completion of the production phase of the E fermentation. col i, the cell culture was cooled to 4-10 ° C and the cells harvested by continuous centrifugation at 15,000 rpm (Heraeus Sepatech). Based on the expected yield of the protein per unit weight of the cell paste, and the amount of purified protein required, an appropriate amount of cell paste by weight is suspended in a buffer solution containing 100 mM Tris, EDTA 50 mM, pH 7.4. The cells are dispersed to a homogeneous suspension using a high shear mixer. The cells are then lysed by passing the solution through a microfluidizer (Microfuidics, Corp. or APV Gaulin, Inc.) twice at 281-421 kg / cm2 (4000-6000 psi). The homogenate is then mixed with the sodium chloride solution to a final concentration of 0.5 M sodium chloride, followed by centrifugation at 7000 x 15 minutes. The resulting button is washed again using 0.5 M sodium chloride, 100 mM Tris, 50 mM EDTA, pH 7.4. The resulting washed inclusion bodies are solubilized with 1.5 M guanidine hydrochloride (GuHCl) for 2-4 hours. After centrifugation at 7000 × g for 15 minutes ^, the button or concentrate is discarded and the supernatant containing the polypeptide is incubated at 4 ° C overnight to allow additional extraction of GuHCl). After high speed centrifugation (30,000 xg) to remove the insoluble particles, the protein solubilized with GuHCl is refolded by rapidly mixing the GuHCl extract with 20 volumes of buffer containing 50 mMv sodium pH 4.5 150 mM sodium chloride. , 2 mM EDTA by vigorous stirring. The diluted, refolded protein solution is kept at 4 ° C without mixing for 12 hours before further purification steps.

To clarify the refolded polypeptide solution, a previously prepared tangential unit or filtration, equipped with a 0.16 micron membrane filter with the appropriate surface area (eg, Filtron), equilibrated with 40 mM sodium acetate, pH 6.0, is employed. Xa filtered sample is loaded onto a cation exchange resin (eg Poros HS-50, Perseptive Biosystems). The column is washed with 40 mM sodium acetate, pB 6.0 and eXuida with 250 mM, 500 mM, 1000 mM, and 1500 mM of sodium chloride in the same buffer, in a stepwise fashion. The absorbance at 280 nm of the effluent It is continuously verified. Xas fractions are collected and subsequently analyzed by SDS-PAGE. The fractions containing the IL-21 pplipeptide are then combined and mixed with 4 volumes of water. Xa diluted sample is then loaded onto a previously prepared team of tandem columns of strong anion exchange (Poros HQ-T0, Perseptive Biosystems) and weak anionic (Poros CM-20, Pjerseptive Biosystems) resins. The columns are equilibrated with 40 mM sodium acetate, pH 6.0. Both columns are washed with 40 mM sodium acetate, pH 6.0, 200 mM sodium chloride. The CM-20 column is then eluted using a linear gradient of 10 volumes in the range of 0.2 M sodium chloride, 50 mM sodium acetate, pH 6.0 to 1.0 M sodium chloride, 50 mM sodium acetate, pH 6.5. The fractions are collected under constant periodic verification A28Q of the effluent. The fractions containing the polypeptide (d terminated, for example, by 16% SDS-PAGE), are then combined. The resulting polypeptide 11-21 should show more than 95 ° -purity after the above refolding and purification steps. No major contaminant bands should be observed from the 16% SDS-PAGE gel, stained with Commassie blue, when 5 micrograms of purified protein are loaded. Protein purified IX-21 can be tested for endotoxin / LPS contamination and typically the LPS content is less than 0.1 ng / ml according to the LAL assays.

Example 7: Cloning and Expression of IL_21 in a Baculovirus Expressing System In this example, the plasmid shuttle vector pA2 is used to insert the IL-21 polynucleotide into a baculovirus, to express IL-21. This expression vector contains the strong polyhedrin promoter of the nuclear polyhedrosis virus Au tigraphic callforni ca (AcMJSfPV) followed by convenient restriction sites such as Bamñl, Xbal and. Asp718. The polyadenylation site of simian virus 40 ("SV40") is used for efficient polyadenylation. For ease of selection of the recombinant virus, the plasmid contains the gene beta beta-galactoside = a from E. col i under the control of a weak Drosophila promoter in the same orientation, followed by the polyadenylation signal of the polyhedrin gene. Xos inserted genes are flanked on both sides by viral sequences for cell-mediated homologous recombination with the wild-type viral DNA, to generate a viable virus expressing the cloned polynucleotide IX-21.

Many other baculovirus vectors may be used in place of the above vector, such as pAc373, pVL941 and pAcIMl, as one skilled in the art would readily appreciate, as long as the construct provides appropriately localized signals for transcription, translation, secretion and similar, including a signal peptide and an intrastructural AUG, as required. Such vectors are described, for example, by Luckow et al. { yolol ogy 170: 31-39 (1989)). Specifically, the cDNA sequence of IL-21 contained in the deposited clone, including the AUG start codon and any naturally associated leader sequence, is amplified using a PCR protocol described in Example 1. If the signal sequence is naturally occurring is used to produce the secreted protein, the pA2 vector does not need a second signal peptide. However, since the signal peptides of natural origin, predichps, IL-21 and IL-22 are not known, the vector can be modified (now designated pA2GP), to include a baculovirus guiding sequence, using the. standard methods described by Summers et al. ("A Manual of Methods for Baculovirus Vectors and Insect Cell Culture Procedures," Texas Agricultural Experimental Station Bulletin No. 1555 (1987)). More specifically, the cDNA sequence encoding the full-length IL-21 protein in the deposited clone is amplified using the oligonucleotide PCR primers corresponding to the 5 'and 3' sequences of the gene. The 5 'primer has the sequence 5' -CGC CGC GGA TCC GCC ATC CGC ACG AGT GGA CAC GG-3 '(SEQ ID NO: 11) containing the BamHI restriction enzyme site, an efficient signal for the start of the translation in eukaryotic cells (shown in the sequence of the primer in italics, Kozak, M., "Mol. Bi ol. 196: 947-950 (1987)), a" C "residue to preserve the reading structure, and 16 nucleotides of the complete IL-21 protein sequence shown in Figure 1. The 3 'primer maintains the 5' sequence CGC GGT ACC CAC TGA ACG GGG CAG CAC GC-3 '(SEQ ID NO: 12) containing the Asp718 restriction site followed by 20 nucleotides complementary to the 3 'non-coding sequence in Figure 1. The amplified fragment is isolated from a 1% agarose gel using commercially available equipment ("Geneclean" BIO 101 Inc., La Jolla, CA) .The fragment is then digested with the appropriate restriction enzymes and again purified on a 1% agarose gel. The plasmid is digested with the corresponding restriction enzymes and can optionally be dephosphorylated using calf intestinal phosphatase, using routine procedures known in the art. The DNA is then isolated from the 1% agarose gene using commercially available equipment ("Geneclean" BIO 101 Inc., La Jolla, CA). The dephosphorylated fragment and plasmid are ligated together with the T4 DNA ligase. E. col i HB101 or other hosts E. Suitable coli, such as XL-1 Blue cells (Stratageme Cloning Systems, La Jolla, CA) are transformed with the ligation mixture and spread on culture plates. The bacteria containing the plasmid are identified by digesting the DNA from individual colonies and analyzing the digestion product by gel electrophoresis. The sequence of the cloned fragment is confirmed by sequencing e- ADK_ Five micrograms of a plasmid containing the polynucleotide are cortansfected with 1.0 μg of commercially available linearized baculovirus DNA ("BaculoGold ™ Baculovirus DNA", Pharmingen, San Diego, CA ), using the lipofection method described by Felgner et al (Proc.Nat.Acid.Sci.USA 84: 7413-7417 (1987)). One μg of the BaculoGold ™ viral DNA and 5 μg of the plasmid are mixed in a sterile well of a microtiter plate containing 50 μl of serum-free Grace medium (Life Technologies Inc., Gaithersburg, MD). After this, 10 μl of Lipofectin plus 90 μl of Grace's medium are added, mixed and incubated for 15 minutes at room temperature. Subsequently, the transfection mixture is added dropwise to the Sf9 insect cells (ATCC CRL 1711) seeded in a 35 mm tissue culture plate with 1 ml Grace's medium without serum. The plate is then incubated for 5 hours at 27 ° C. The transfection solution is then removed from the plate and 1 ml of Grace's insect medium supplemented with 10% fetal calf serum is added. The culture is then continued at 27 ° C for four days.

After four days the supernatant is harvested and a plaque assay is performed, as described by Summers and Smit (s upra). An agarose gel with "Blue Gal" (liie. Technologies Inc., Gaithersburg) is used to allow easy identification and isolation of gal-expressing clones, which produce the blue-stained plates. (A detailed description of such a "plate assay" can also be found in the user guide for the culturing of insect cells and baculovirology distributed by Life Technologies Jnc., Gaitnersburg, pages S-J.0,). After the appropriate incubation, the blue-stained plates are picked up with the tip of a micropipette (e.g., Eppendorf). The agar containing the recombinant viruses is then resuspended in a microcentrifuge tube containing 200 μl of Grace's medium and the suspension containing the recombinant baculovirus is used to infect SE9 cells seeded in 35 mm boxes. Four days later the supernatants of these culture boxes are harvested and then stored at 4 ° C. To verify the expression of the polypeptide, Jas Sf.9 cells are desarralled in Grace's medium supplemented with 10% heat-inactivated FBS. The cells are infected with the recombinant baculovirus containing the polynucleotide at a multiplicity of infection ("MOI") of about 2. If the radiolabeled proteins are desired, 6 hours later the medium is removed and replaced with the SF900 II medium less methionine and cysteine (available from Life Technologies Inc., Rockville, MD). After 42 hours, 5 μCi of 35S-methionine and 5 μCi of 35S-cysteine (available from Amersham) are added. The cells are subsequently incubated for 16 hours and then harvested by centrifugation. The proteins in the supernatant as well as the intracellular proteins are analyzed by SDS-PAGE, which is followed by autoradiography (if radiolabeled). Microsequencing of the amino acid sequence of the amino terminus of the purified protein can be used to determine the amino-terminal sequence of the IL-21 protein produced.

Ex empl o 8: Expression of IL-21 in Mammalian Cells The IL-21 polypeptide can be expressed in a mammalian cell. A typical mammalian expression vector contains a promoter element, which mediates the initiation of mRNA transcription, a protein coding sequence, and the signals required for the termination of transcription and polyadenylation of the transcript. Additional elements include enhancers, Kozak sequences and intervening sequences flanked by donor and acceptor sites for RNA splicing. Highly efficient transcription is achieved with the SV40 late and early promoters, the long terminal repeats (LTRs) of Retroviruses, eg, RSV, HTLV-1, HIV-1 and the cytomegalovirus early promoter (CMV). However, cellular elements can also be used (for example, the human actin promoter). Expression vectors suitable for use in the practice of the present invention include, for example, vectors such as pSVL and pMSG (Pharmacia, Uppsala, Sweden), pRSVcat (ATCC 37152), pSV2dhfr (ATCC 37146), pBC12MI (ATCC 67109 ), pCMVSport 2.0, and pCMVSport 3.0. Mammalian host cells that could be used include, Hela, 293, H9 and Jurkat cells, mouse NIH3T3 and C127 cells, Cos 1, Cos 7 and CVl cells, QC1-3 quail cells, mouse X cells. and Chinese hamster ovary cells (CHO). Alternatively, the IL-21 polypeptide can be expressed in stable cell lines containing the integrated IL-21 polynucleotide within a chromosome. Cotransfection with a selectable marker such as dhfr, gpt, neomycin or hygromycin allows the identification and isolation of the transfected cells. The transfected IL-21 gene can also be amplified to express large amounts of the encoded protein. The DHFR marker (dihydrofolate reductase) is useful in the development of cell lines carrying several hundred or even several thousand copies of the gene of interest (see, for example, Alt, FW, et al., J. Bi ol. Ch em. : 1357-1370 (1978), Hamlin, JL and Ma., C, Bi och em. Et Bi ophys, Acta 1097: 107-143 (1990); Page, MJ and Sydenham, MA, Bi ot echnol ogy 9: 64 -68 (1991)). Another useful selection marker is the enzyme glutamine synthase (GS; Murphy et al., Bi ochem J. 227: 277-279 (1991); Bebbington et al., Bi o / Techn olgy 10: 169-175 ( 1992)). Using the markers, the mammalian cells are developed in selective medium and the cells with the highest resistance are selected. These cell lines contain the amplified gene (s) integrated within a chromosome. Chinese hamster ovary (CHO) and NSO cells are frequently used for protein production. The plasmid derivatives pSV2-dhfr (ATCC Accession No. 37146), the expression vectors pC4 (ATCC Accession No. 209646) and pC6 (Accession Accession No. 209647) contain the strong promoter (LT.R) of the Virus of Sarcoma de Rous (Cullen et al., Mol.Cel. l Bi., 438-447 (March, 1985)) plus a fragment of the CMV enhancer (Boshart, et al., Cel lA l: 521-530 ( 1985)). Multiple cloning sites, for example, with the restriction enzyme cleavage sites, BamHI, Xbal and Asp718, facilitate the cloning of IL-21. The vectors also contain the 3 'intron, the polyadenylation and termination signal of the rat preproinsulin gene, and the mouse DHFR gene under the control of the SV40 early promoter.

Specifically, plasmid pC6, for example, is digested with the appropriate restriction enzymes and then dephosphorylated using intestinal calf phosphatases by methods known in the art. The vector is then isolated from an agarose gel at 19 °. The IL-21 polynucleotide is amplified according to the protocol described in Example 1. If the naturally occurring signal sequence is used to produce the secreted protein, the vector does not need a second signal peptide. Alternatively, if the signal sequence of natural origin is not used, the vector can be modified to include a heterologous signal sequence (see for example, W096 / 34891). The amplified fragment is isolated from a 1% agarose gene using commercially available equipment ("Geneclean" BIO 101 Inc., La Jolla, CA). The fragment is then digested with the appropriate restriction enzymes and again purified on an agarose-1% gel. The amplified fragment is then digested with the same restriction enzyme and purified. on a 1% agarose gel. The isolated fragment and the dephosphorylated vector are then ligated with the T4 DNA ligase. The E cells. col i HB101 or XL-1 Blue are then transformed, and bacteria having the fragment inserted into plasmid pC6 are identified using, for example, restriction enzyme analysis. Chinese hamster ovary cells lacking an active DHFR gene are used for transfection. Five μg of the expression plasmid pC6 are cotransfected with 0.5 μg of the pSVneo plasmid using lipofectin (Felgner et al., Upra). Plasmid pSV-neo contains a dominant selected marker, the neo gene from Tn5 that codes for an enzyme that confers resistance to a group of antibiotics including G418. The cells are seeded in MEM alpha less supplemented with 1 mg / ml of G418. After 2 days, the cells are trypsinized and seeded in hybridoma cloning plates (Greiner, Germany) in MEM alpha less supplemented with 10, 25, or 50 ng / ml of methotrexate plus 1 mg / ml of G418. After approximately 10-14 days, the single clones are trypsinized and then seeded in 6-well petri dishes or in 10-ml flasks using different concentrations of methotrexate. (for example, 50 nM, 100 nM, 200 nM, 400, nM 800 nM). Clones that develop at the highest concentrations of methotrexate are then transferred to new 6-well plates containing even higher concentrations of methotrexate (1 μM, 2 μM, 5 μM, 10 mM, 20 mM). The same procedure is repeated until clones are obtained which develop in a concentration of 100-200 μM. The expression of IL-21 is analyzed, for example, by SDS-PAGE and staining or Western blot or by reverse phase HPLC analysis.

Ex empl o 9: Protein fusions of IL -21 The IL-21 polypeptides are preferably fused to other proteins. These fusion proteins can be used for a variety of applications. For example, fusion of the IL-21 polypeptides to His-tag, HA-tag, protein A, IgG domains, and the maltose binding protein facilitates purification (see Example 5, see also EP-A- 394, 827; Traunecker et al., Na ture 331: 84-86 (1988)). Similarly, fusion to IgG-1, IgG-3 and albumin increases the half-life time in vi. Nuclear localization signals fused to the IL-21 polypeptides can direct the protein to a specific cell site, while the covalent heterodimer or homodimers can increase or decrease the activity of a fusion protein. The fusion proteins can also create chimeric molecules that have more than one function. The fusion proteins can increase the solubility and / or the stability of the fused protein compared to the unfused protein. All types of fusion proteins described above can be elaborated by modifying the following protocol, which describes the fusion of a polynucleotide to an IgG molecule, or the protocol described in Example 5. In summary, the human Fe the IgG molecule can be amplified by PCR, using the primers spanning the 5 'and 3' ends of the sequence described below. These primers should also have convenient restriction enzyme sites, which will facilitate cloning within an expression vector, preferably a mammalian expression vector.

For example, if pC4 (Access No. 209646) is used, the human Fe portion can be ligated into the BamEl cloning site. Note that site 3 'BamHI must be destroyed. Next, the vector containing the human Fe protein is again restricted with BamHI, linearizing the vector, and the IL-21 polynucleotide, isolated by the PCR protocol described in Example 1, is ligated into the BamHI site. Note that the polynucleotide is cloned without a stop codon, otherwise a fusion protein will not be produced. If the signal sequence of natural origin is used to produce the secreted protein, pC4 does not need a second signal peptide. Alternatively, if the naturally occurring signal sequence is not used, the vector can be modified to include a heterologous signal sequence (see, for example, W096 / 34891).

Fe IgG Human Region. { I KNOW THAT. ID. NO: 13): GGGATCCGGAGCCCAAATCTTCTGACAAAACTCACACATGCCCACCGT GCCCAGCACCTGAATTCGAGGGTGCACCGTCAGTCTTCCTCTTCCCCCCAAA ACCCAAGGACACCCTCATGATCTCCCGGACTCCTGAGGTCACATGCGTGGTG GTGGACGTAAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACG GCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAG CACGTACCGTGTGGTCAGCGTCCTCACGTCCTGCACCAGGACTGGCTGAATG GCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAACCCCCATCGA GAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACC CTGCCCCCATCCCGGGATGAGCTGACCAAGAACCAGGTCAGCCTGACCTGCC TGGTCAAAGGCTTCTATCCAAGCGACATCGCCGTGGAGTGGGAGAGCAATGG GCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGC TCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGG GGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACAC GCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGAGTGCGACGGCCGCGACTC TAGAGGAT EXAMPLE 1 0: Producing an An ti body The antibodies of the present invention can be prepared by a variety of methods (see, Current Protocols, Chapter 2). For example, cells expressing IL-21 are administered to an animal to induce the production of sera containing polyclonal antibodies. In a preferred method, an IL-21 protein preparation is prepared and purified to render it substantially free of natural contaminants. Such preparation is then introduced into an animal in order to produce polyclonal antisera of higher specific activity. In the most preferred method, the antibodies of the present invention are monoclonal antibodies (or protein binding fragments thereof). Such monoclonal antibodies can be prepared using the hybridoma technology (Kohler et al., Na ture 256: 495 (1975); Kohler et al., Eur. J. Immunol. 6: 511 (1976); Kohler et al., Eur. J. Immunol. 6: 292 (1976); Hammerling et al., In: Monoclonal Antibodies and T-Cell Hybridomas, Elsevier, N.Y., pp. 563-681 (1981)). In general, such methods involve immunizing an animal (preferably a mouse) with the IL-21 polypeptide or, more preferably, with a cell expressing the secreted IL-21 polypeptide. Such cells can be cultured in any suitable tissue culture medium. However, it is preferable to culture cells in Eagle's modified Eagle's medium supplemented with 10% fetal bovine serum (inactivated at approximately 56 ° C), and supplemented with approximately 10 g / liter of non-essential amino acids, approximately 1,000 U / ml of penicillin, and approximately 100 μg / ml of streptomycin.

The splenocytes of such mice are extracted and fused with an appropriate line of myeloma cells. Any suitable myeloma cell line can be employed in accordance with the present invention. However, it is preferable to use the progenitor myeloma cell line (SP20), available from the ATCC. After fusion, the resulting hybridoma cells are selectively maintained in HAT medium, and then cloned by limiting dilution as described by Wands et al. (Ga s troen t eroi ogy 80: 225-232 (1981)). The hybridoma cells obtained through such selection are then evaluated to identify the clones that secrete the antibodies capable of binding to the IL-21 polypeptide. Alternatively, additional antibodies capable of binding to the IL-21 polypeptide can be produced in a two step procedure using anti-idiotypic antibodies. Such a method makes use of the fact that the antibodies are themselves antigens, and therefore, it is possible to obtain an antibody that binds to a second antibody. According to this method, protein-specific antibodies are used to immunize an animal, preferably a mouse. Splenocytes from such an animal are then used to produce the hybridoma cell, and the hybridoma cells are selected to identify the clones that produce an antibody whose ability to bind to the specific antibody of the IL-21 protein can be blocked by IL-21. . Such antibodies comprise anti-idiotypic antibodies to the IL-21 protein-specific antibody, and can be used to immunize an animal to induce the formation of additional antibodies to the IL-21 protein, additional. It will be appreciated that Fab and F (ab'2) and other fragments of the antibodies of the present invention can be used according to the methods described herein. Such fragments are typically produced by proteolytic cleavage, using enzymes such as papain (to produce Fab fragments) or pepsin (to produce F (ab ') 2 fragments). Alternatively, fragments of binding to the IL-21 protein, secreted, can be produced through the application of recombinant DNA technology or through synthetic chemistry.

For the in vivo use of antibodies in humans, it may be preferable to use "humanized" chimeric monoclonal antibodies. Such antibodies can be produced using the genetic constructs derived from the hybridoma cells that produce the monoclonal antibodies described above. Methods for producing chimeric antibodies are known in the art (for a review see Morrison, Science 229: 1202 (1985); Oi et al., BioTechniques 4: 214 (1986); Cabilly et al., U.S. Patent No. 4,816,567; Taniguchi et al., EP-171496; Morrison et al., EP-173494; Neuberger et al., WO-8601533; Robinson et al., WO-8702671; Boulianne et al., Nature 312: 643 (1984 ); Neuberger et al., Nature 314: 268 (1985)).

Example 11: Production of IL-21 Protein for High Throughput Screening Assays The following protocol produces a supernatant containing the IL-21 polypeptide to be tested. This supernatant can then be used in assays. selection described subsequently in Examples 13-20.

First, the poly-D-lysine buffer solution (644 587 Boehringer-Mannheim) (1 mg / ml in PBS) 1:20 in PBS is diluted (Phosphate-buffered saline solution with / without calcium or magnesium 17-516F Biowhittaker) for a working solution of 50 μg / ml. 200 μl of this solution is added to each well (24-well plates) and incubated at room temperature for 20 minutes. It is ensured that the solution is distributed over each well (note: you can use an automatic 12-channel pipette with tips on each channel). The poly-D-lysine solution is aspirated and rinsed with 1 ml of PBS. The PBS should remain in the well just before plating the cells and the plates can be coated with poly-lysine beforehand for up to two weeks. 293T cells (do not carry cells beyond P + 20) are plated at 2 x 10 5 cells / well in 0.5 ml of DMEM (Dulbecco's Modified Eagle's Medium) supplemented with 4.5 g / l glucose, L-glutamine (12-604F Biowhittaker)), 10% FBS inactivated by heat (14-503F Biowhittaker), and lx Penstrep (17-602E Biowhittaker). The cells are allowed to develop overnight.

After the overnight incubation, they are mixed together in a sterile solution container: 300 μl of Lipofectamine (18324-012 Gibco / BRL) and 5 ml of Optimem I (31985070 Gibco / BRL) in each well of a 96-well plate. wells With a small-volume, multi-channel automatic pipette, the aliquot of approximately 2 μg of an expression vector containing a polynucleotide insert, produced by the methods described in Examples 8 and 9, is placed in a 96-well plate of Round bottom properly marked. With an automatic multi-channel pipette, 50 μl of the Lipofectamine / Optimem I mixture is added to each well. Mix gently with the pipette, aspirating and expelling. It is incubated at room temperature for 15-45 minutes. After approximately 20 minutes, an automatic multi-channel pipette is used to add 150 μl of Optimem I to each well. As a control, a vector DNA plate that lacks an insert must be transfected with each group of transfections. Preferably, the transfection must be performed by performing the following tasks simultaneously in a staggered manner. In this way, the time is reduced to half, and the cells are not excessively incubated in PBS. First, Person A aspirates the medium from four 24-well cell plates, and then Person B rinses each well with 0.5-1 mL of PBS. Person A then aspirates the PBS rinse, and Person B, using an automatic 12-channel pipette coft tips on each channel, adds the 200 μl of the DNA / Lipofectamine / Optimem I complex to the odd wells first, then to the wells pairs, to each row on the 24-well plates. The plates are then incubated at 37 ° C for 6 hours. While the cells are being incubated, the appropriate media are prepared: either 1% BSA in DMEM with lx penstrep. or the HGS CHO-5 medium (116.6 mg / liter of CaCl2 (anhydrous), 0.00130 mg / 1 CuS04-5H20, 0.050 mg / 1 of Fe (N03) 3-9H20, 0.417 mg / 1 of FeS04-7H20, 311.80 mg / 1 of KC1, 28.64 mg / 1 of MgCl2, 48.84 mg / 1 of MgSO4, 6995.50 mg / 1 of NaCl, 2400.0 mg / 1 of NaHCO3, 62.50 mg / 1 of NaH2P04-H20, 71.02 mg / 1 of Na2HP04, 4320 mg / 1 of ZnS04-7H20, 0.002 mg / 1 of araguidonic acid, 1022 mg / 1 of cholesterol, 0.070 mg / 1 of DL-alpha-tocopherol-acetate, 0.0520 mg / ml of linoleic acid, 0.010 mg / 1 linolenic acid; 0.010 mg / 1 of myristic acid, 0.010 mg / 1 of oleic acid, 0.010 mg / 1 of palmic acid, 0.010 mg / 1 of palmitic acid, 100 mg / 1 of Pluronic • F-68, 0.010 mg / 1 of stearic acid 2.20 mg / 1 of Tween 80, 4551 mg / 1 of D-glucose, 130.85 mg / ml of L-alanine, 147.50 mg / ml of L-arginine-HCl, 7.50 mg / ml of L-asparagine-H20, 6.65 mg / ml L-aspartic acid, 29.56 mg / ml L-cystine-2HCl-H20, 31.29 mg / ml L-cystine-2HCl, 7.35 mg / ml L-glutamic acid, 365.0 mg / ml L- glutamine, 18.75 mg / ml glyc ina; 52.48 mg / ml L-histadine-HCl-H20; 106.97 mg / ml of L-isoleucine; 111.45 mg / ml L-leucine; 163.75 mg / ml L-lysine HCl; 32.34 mg / ml of L-methionine; 68.48 mg / ml L-phenylalanine; 40.0 mg / ml L-proline; 26.25 mg / ml L-serine; 101.05 mg / ml L-threonine; 19.22 mg / ml L-tryptophan; 91.79 mg / ml of L-tyrosine-2Na-2H20; and 99.65 mg / ml L-valine; 0.0035 mg / 1 of biotin; 3.24 mg / 1 of D-Ca Pantothenate; 11.78 mg / 1 choline chloride; 4.65 mg / 1 folic acid; 15.60 mg / 1 of i-inositol; 3.02 mg / 1 niacinamide; 3.00 mg / 1 pyridoxal-HCl; 0.031 mg / 1 pyridoxine-HCl; 0.319 mg / 1 riboflavin; 3.17 mg / 1 thiamine HCl; 0.365 mg / 1 thymidine; 0.680 mg / 1 of Vitamin B? 2; HEPES 25 M cushion; 2.39 mg / 1 Na hypoxanthine; 0.105 mg / l of lipoic acid; 0.081 mg / 1 putrescine sodium-2HCl; 55.0 mg / 1 of sodium pyruvate; 0.0067 mg / 1 of sodium selenite; 20 μM ethanolamine; 0.122 mg / 1 ferric citrate; 41.70 mg / 1 methyl-B-cyclodextrin formed in complex with linoleic acid; 33.33 mg / 1 of methyl-B-cyclodextrin complexed with oleic acid; 10 mg / 1 of methyl-B-cyclodextrin formed in complex with retinal acetate. The osmolarity is adjusted to 327 mOsm with glutamine 2 mm and lx penstrep. (100 g dissolved in 1 liter of DMEM for a stock solution of 10% BSA (81-068-3 Bayer)). Media is filtered and 50 μl collected for the endotoxin assay in 15 ml conical polystyrene. The transfection reaction is maintained again, preferably by two people, at the end of the incubation period. Person A aspirates the transfection medium, while Person B adds 1.5 ml of the appropriate medium to each well. It is incubated at 37 ° C for 45 or 72 hours, depending on the means used (1% BSA for 45 hours or CHO-5 for 72 hours). On day 4, using a 300 μl automatic multichannel pipette, 600 μl aliquots are taken in a 1 ml deep well plate and the supernatant remaining in a 2 ml deep well. The supernatants from each well can then be used in the assays described in Examples 13-20. It is specifically understood that when the activity is obtained in any of the assays described below using a supernatant, the activity originates either from the IL-21 polypeptide directly (for example, as a secreted protein) or from IL-21 which induces the expression of other proteins, which are then secreted into the supernatant. Thus, the invention further provides a method for identifying the protein in the supernatant, characterized by an activity in a particular assay.

Ex empl o 12: Cons ti t uci on of the G rea Reporting A signal transduction pathway involved in the differentiation and proliferation of cells is called the Jaks-STATs pathway. The proteins activated in the Jaks-STATs pathway are linked to the elements of the gamma activation site ("GAS") or to the interferon-sensitive response element ("ISRE"), located in the promoter of many genes. The binding of a protein to these elements alters the expression of the associated gene. The GAS and ISRE elements are recognized by a class of transcription factors called Signal Transducers and Transcription Activators or "STATs". There are six members of the STATs family. Statl and Stat3 are present in many cell types, as is Stat2 (since the response to IFN-alpha is widespread). Stat4 is more restricted and is not found in many cell types, although it has been found in helper T cells of class I after treatment with IL-12. Stat5 was originally called mammary development factor, but has been found at higher concentrations in other cells, including myeloid cells. This can be activated in tissue culture cells by many cytokines. STATs are activated to translocate from the cytoplasm to the nucleus after tyrosine phosphorylation by a group of kinases known as the Janus Kinase family ("Jaks"). Jaks represent a different family of tyrosine kinases, soluble, and include Tyk2, Jakl, Jak2 and Jak3. These kinases show significant sequential similarity and are generally catalytically inactive in resting cells. Jaks are activated by a broad range of receptors summarized in the following Table (adapted from review by Schidler and Darnell, Ann. Rev. Bi ochem 64: 621-51 (1995)). A cytokine receptor family, capable of activating Jaks, is divided into two groups: a) Class 1 includes the receptors for IL-2, IL-3, IL-4, IL-6, IL-7, IL- 9, IL-11, IL-12, IL-15, Epo, PRL, GH, G-CSF, GM-CSF, LIF, CNTF, and thrombopoietin; and b) Class 2 includes IFN-alpha, IFN-gamma and IL-10. Class 1 receptors share a conserved cysteine portion (a group of four conserved cysteines and a tryptophan) and a WSXWS portion (a proximal membrane region encoding Trp-Ser-Xxx-Trp-Ser (where "Xxx" represents any amino acid; SEQ ID NO: 14)). Thus, in the binding of a ligand to a receptor, the Jaks are activated, which in turn activate STATs, which are then translocated and linked to the GAS elements. This complete process is encompassed in the path of signal transduction Jaks-STATs.

Therefore, the activation of the Jaks-STATs pathway, reflected by the GAS link or the ISRE element, can be used to indicate the proteins involved in the proliferation and differentiation of cells. For example, growth factors and cytokines are known to activate the Jaks-STATs pathways (see Table below). In this way, through the use of GAS elements linked to reporter molecules, the activators of the Jaks-STATs pathway can be identified.

JAKs STATs GAS (elements) or ISRE Ligando tyk2 Jakl Jak2 Jak3 IFN family IFN-alpha / beta 1,2,3 ISRE IFN-gamma 1 GAS (IRFl> Lys6> IFP) 11-10 1.3 Family gp! 30 IL-6 (pleiotrohic) 1,3 GAS (IRFl> Lys6> IFP) 11-11 (pleiotrohic) 1,3 OnM (pleiotrohic) 1,3 LIF (pleiotrohic) 1,3 CNTF (pleiotrohic) - / + 1, 3 G-CSF (pleiotrohic) 1, 3 n 12 (pleiotrohic) - 1, 3 Family g-C IL-2 (lymphocytes) + 1,3, 5 GAS IL-4 (lymph / myeloid) + 6 GAS (IRF1 == IFP »Ly6) (IgH) IL-7 (lymphocytes) + 5 GAS IL-9 (lymphocytes) + 5 GAS IL-13 (lymphocytes) • > 6 GAS IL-15 + 5 GAS Family gp! 40 IL-3 (myeloid) 5 GAS (IRFl > IFP »Ly6) IL-5 (myeloid) 5 GAS GM-CSF (myeloid) 5 GAS Family Growth Hormone GH + - 5 PRL +/- + - 1.35 EPO + - 5 GAS (B- CAS> IRFl = IFP »Ly6) Tyrosine-kinases EGF Receptor? + 1, 3 GAS (IRF1) PDGF? + 1, 3 CSF-1? + 1, 3 GAS (no IRF1) To construct a synthetic GAS-containing promoter element, which is used in the biological assays described in Examples 13-14, a PCR-based strategy is employed to generate a GAS-SV40 promoter sequence. The 5 'primer contains four tandem copies of the GAS binding site found in the IRF1 promoter, and previously shown to bind to STATs after induction with a range of cytokines (Rothman et al., Immuni ty 1: 457-468 (1994)), although other GAS or ISRE elements can also be used. The 5 'primer contains 18 base pairs of the sequence complementary to the SV40 early promoter sequence and is flanked with the restriction site Xh ol. The primer sequence 5' is '-GCG CCT CGA GAT TTC CCC GAA ATC TAG ATT TCC CCG AAA TGA TTT CCC CGA AAT GAT TTC CCC GAA ATA TCT GCC ATC TCA ATT AG-3' (SEQ ID NO: 15).

The downstream primer (3 ') is complementary to the SV40 promoter and is flanked with a Hindl l l: 5' -GCG GCA AGC TTT TTG CA AGC CTA GGC-3 'site (SEQ ID NO: 16). PCR amplification is performed using the SV40 promoter as a template, present in the B-gal promoter plasmid obtained from Clontech. The resulting PCR fragment is digested with Xhol and HindlII and subcloned into BLSK2 (Stratagene). Sequencing with forward and reverse primers confirms that the insert contains the following sequence: CTCGAGATTTCCCCGAAATCTAGATTCCCCGAAATGATTTCCC CGAAATGATTTCCCCGAAATATCTGCCATCTCAATTAGTCAGCAACCATAGT CCCGCCCCTAACTCCGCCCATCCCGCCCCTAAACTCCGCCCAGTTCCGCCCA TTCTCCGCCCCATGGCTGACTAATTTTTTTTATTTATGCAGAGGCCGAGGCC GCCTCGGCCTCTTAGCTATTCCAGAAGTAGTGAGGAGGCTTTTTTGGAGGCC TAGGCTTTTGCAAAAAGCTT (SEQ ID NO:.. 17). With this GAS promoter element, linked to the SV40 promoter, a construction of the GAS reporter: SEAP2 is immediately engineered. Here, the reporter molecule is a secreted alkaline phosphatase, or "SEAP". Clearly, however, any reporter molecule can be used instead of SEAP, in this or any of the other examples. Well-known reporter molecules that can be used in place of SEAP include chloramphenicol acetyltransferase (CAT), luciferase, alkaline phosphatase, B-galactosidase, green fluorescent protein (GFP), or any protein detectable by an antibody. The previous sequence confirmed that the synthetic GAS-SV40 promoter element is subcloned into the pSEAP-promoter vector, obtained from Clontech, using HindlII and Xhol, effectively replacing the SV40 promoter with the amplified GAS: SV40 promoter element, to create the GAS-vector. SEAP. However, this vector does not contain a neomycin resistance gene, and therefore, is not preferred for mammalian expression systems. In this way, in order to generate stable, mammalian cell lines, which express the GAS-SEAP reporter, the GAS-SEAP cassette is removed from the GAS-SEAP vector using Sali and No tl, and inserted into a spinal vector containing the neomycin resistance gene, such as pGFP-1 (Clontech), using these restriction sites at the multiple cloning site, to create the GAS-SEAP /? eo vector. Once this vector is transfected into mammalian cells, this vector can then be used as a reporter molecule for the GAS binding as described in Examples 13-14. Other constructs can be elaborated using the above description and replacing GAS with a different promoter sequence. For example, the construction of the reporter molecules containing the NF-kappaB and EGR promoter sequences are described in Examples 15 and 16. However, many other promoters can be substituted using the protocols described in these Examples. For example, SRE, IL-2, NFAT, or the Osteocalcin promoters can be substituted, alone or in combination (eg, GAS / NF-kappaB / EGR, GAS / NF-kappaB, I 1-2 / NFAT, or NF-kappaB / GAS). Similarly, other cell lines can be used to test the reporter's construction activity, such as HeLa (epithelial), HUVEC (endothelial), Reh (B cells), Saos-2 (osteoblasts), HUVAC (aortic) or cardiomyocytes Example 13: Testing of Allocation of Al to Rendimi on for the Activity of Cells T The second protocol is used to evaluate the activity of IL-21 T cells by determining whether the IL-21 supernatant proliferates and / or differentiates T cells. T cell activity is evaluated using the GAS / SEAP construct / Neo produced in Example 12. In this way, the factors that increase the activity of SEAP indicate the ability to activate the transfection pathway of the Jaks-STATS signal. The T cell used in this assay is the Jurkat T cell (ATCC Accession No. TIB-152), although the Molt-3 cells (ATCC Accession No. CRL-1552) and the Molt-4 cells (ATCC No. Access CRL-1582) can also be used. Jurkat T cells are CD4 + Thl lymphoblastic helper cells. In order to generate stable cell lines, approximately 2 million Jurkat cells are transfected with the GAS-SEAP / Neo vector using DMRIE-C (Life Technologies: transfection procedure described below). The transfected cells are seeded at a density of approximately 20. 000 cells per well and the transfectants resistant to 1 mg / ml of genticin are selected. The resistant colonies are expanded and then tested for their response at increasing concentrations of gamma-interferon. The response to the dose of a selected clone is demonstrated. Specifically, the following protocol will produce enough cells for 75 wells containing 200 μl of cells. In this way, the scale is raised, or is made in multiples to generate enough cells for multiple 96-well plates. Jurkat cells are maintained on RPMI + 10% serum with 1% Pen-Strep. 2.5 ml of OPTI-MEM (Life Technologies) are combined with 10 μg of the plasmid DNA in a T25 flask. 2.5 ml of OPTI-MEM containing '50 μl of DMRIE-C are added and incubated at room temperature for 15-45 minutes. During the incubation period, the cell concentration is counted, the required number of cells are taken (107 per transfection), and resuspended in OPTI-MEM to a final concentration of 107 cells / ml. Then add 1 ml of 1 x 107 cells in OPTI-MEM to the T25 flask and incubate at 37 ° C for 6 hours. After incubation, 10 ml of RPMI + 15% serum are added.

Jurkat stable reporter lines: GAS-SEAP are maintained in RPMI + 10% serum, 1 mg / ml genticin, and 1% Pen-Strep. These cells are treated with the supernatants containing the IL-21 polypeptides, or the polypeptides induced with IL-21 as produced by the protocol described in Example 11. On the day of treatment with the supernatant, the cells must be washed and resuspended in RPMI + 10% serum at a density of 500,000 cells per ml. The exact number of cells required will depend on the number of supernatants that are selected. For a 96-well plate, approximately 10 million cells are required (for 10 plates, 100 million cells). The cells are transferred to a triangular reservoir, in order to supply the cells in a 96-well plate, using a 12-channel pipette. A 12-channel pipette is used, 200 μl of cells are transferred into each well (therefore, 100,000 cells are added per well). After all the plates have been seeded, 50 μl of supernatants are transferred directly from the 96-well plate containing the supernatants in each well, using a 12-channel pipette. In addition, a dose of exogenous interferon gamma (0.1, 1.0, 10 ng) is added to wells H9, H10 and Hll to serve as additional positive controls for the assay. The 96-well plates containing the Jurkat cells treated with the supernatants are placed in an incubator for 48 hours (note: this time is variable between 48-72 hours). Samples of μl of each well are then transferred to an opaque 96-well plate using a 12-channel pipette. Opaque plates must be covered (using cellophane covers) and stored at -20 ° C until the SEAP assays are performed according to Example 17. Plates containing the remaining treated cells are placed at 4 ° C and serve as a source of material to repeat the assay in a specific well, if desired. As a positive control, 100 units / ml of interferon gamma that is known to activate Jurkat T cells can be used. Induction greater than 30 times is typically observed in positive control wells.

Ex empl o 14: Essay of Sel ection of Al to Rendimi ent I tifi ca Acti vity Mi el oide The following protocol is used to evaluate the myeloid activity of IL-21 by determining whether IL-21 proliferates and / or differentiates myeloid cells. The activity of myeloid cells is evaluated using the GAS / SEAP / Neo construct produced in Example 12. Thus, the factors that increase the activity of SEAP indicate the ability to activate the signal transduction pathway of Jaks-STATS. The myeloid cell used in this assay is U937, a cell line of pre-monocytes, although TF-1, HL60 or KG1 can be used. To transiently transfect U937 cells with the GAS / SEAP / Neo construct produced in Example 12, a DEAE-Dextran method is used (Kharbanda et al., Cel l Growth &Di fferen ti a ti on, 5: 259- 265 (1994)). First, 2 x 10 7 U937 cells are harvested and washed with PBS. U937 cells are usually developed in RPMI 1640 medium containing 10% fetal bovine serum inactivated by heat (FBS) supplemented with 100 units / ml penicillin and 100 mg / ml streptomycin. Then, the cells are suspended in 1 ml of 20 mM Tris-HCl buffer (pH 7.4) containing 0.5 mg / ml of DEAE-Dextran, 8 μg of plasmid DNA GAS-SEAP2, 140 M sodium chloride, potassium chloride 5 mM, Na2HP04-7H20 375 μM, 1 mM magnesium chloride, 675 μM calcium chloride. Incubate at 37 ° C for 45 minutes. The cells are washed with RPMI 1640 medium containing 10% FBS and then resuspended in 10 ml of complete medium and incubated at 37 ° C for 36 hours. Stable GAS-SEAP / U937 cells are obtained by developing the cells in 400 μg / ml of G418. The free medium of G418 is used for routine development but each one to two months, the cells must be developed again in 400 μg / ml of G418 for a couple of passes. These cells are tested when harvesting 1 x 108 cells (this is sufficient for ten assays in 96-well plate) and washed with PBS. The cells are suspended in 200 ml of the growth medium described above, with a final density of 5 x 10 5 cells / ml. 200 μl of cells are plated per well in the 96-well plate (or 1 × 10 5 cells / well). 50 μl of the supernatant prepared by the protocol described in Example 11 is added. It is incubated at 37 ° C for 48 to 72 hours. As a positive control, 100 U / ml of interferon gamma known to activate U937 cells can be used. An induction greater than 30 times is typically observed in positive control wells. The supernatant is evaluated by SEAP according to the protocol described in Example 17.

Example # 15: Allocation Test of Al to Rendimi in T he I ti ti fi ca l to Nevonal Activity When cells undergo differentiation and proliferation, a group of genes are activated through many different signal transduction pathways. One of these genes, EGR1 (gene 1 of early development response), is induced in various tissues and cell types after activation. The EGR1 promoter is responsible for such induction. Using the EGR1 promoter linked to the reporter molecules, the activation of the cells by IL-21 can be evaluated.

In particular, the following protocol is used to evaluate neuronal activity in PC12 cell lines. PC12 cells (rat phenochromocytoma cells) are known to proliferate and / or differentiate by activation with a number of mitogens, such as TPA (tetradecanoyl-phorbol acetate), NGF (nerve growth factor) and EGF (epidermal growth factor). The expression of the EGR1 gene is activated during this treatment. Thus, by stably transfecting PC12 cells with a construct containing an EGR promoter linked to the SEAP reporter, activation of PC12 cells by IL-21 can be assessed. The EGR construction / SEAP reporter can be assembled using the following protocol. The promoter sequence EGR-1 (nucleotides -633 to +1; Sakamoto, K. Et al., Oncogene 6: 867-871 (1991)) can be amplified by PCR from human genomic DNA using the following primers: (A) 5 'primer: 5' -GCG CTC GAG GGA TGA CAG CGA TAG AAC CCC GG-3 '(SEQ ID NO: 18) and (B) 3' primer: '-GCG AAG CTT CGC GAC TCC CCG GAT CCG CCT C-3' (SEQ ID NO: 19).

Using the GAS: SEAP / Neo vector produced in Example 12, the product amplified by EGR1 can then be inserted into this vector. The GAS: SEAP / Neo vector is linearized using the restriction enzymes Xhol and íTindlII, removing the larger GAS / SV40 fragment. The amplified product is digested by EGR1 with the same enzymes. The vector and the EGR1 promoter are ligated. To prepare the 96-well plates for cell culture, 2 ml of a coating solution (1:30 dilution of type I collagen (Upstate Biotech Inc. Cat. # 08-115) and 30% ethanol (sterilized by filtration)) through a 10 cm or 50 ml plate per well of the 96-well plate, and allowed to air dry for 2 hours. PC12 cells are routinely grown in RPMI-1640 medium (Bio Whittaker) containing 10% horse serum (JRH BIOSCIENCES, Cat. # 12449-78P), 5% heat inactivated bovine fetal serum (FBS), supplemented with 100 units / ml of penicillin and 100 μg / ml of streptomycin on a 10 cm tissue culture box, pre-coated. A 1: 4 division is made every three to four days. The cells are removed from the plates by scraping and resuspended by repeatedly sucking and ejecting with a pipette for more than 15 times. The EGR / SEAP / Neo construct is transfected into PC12 using the Lipofectamine protocol described in Example 11. The stable cells of EGR-SEAP / PC12 are obtained by developing the cells in 300 μg / ml of G418. The free medium of G418 is used for routine development but each one to two months, the cells must be developed again in 300 μg / ml of G418 for several passes. To evaluate neuronal activity, a 10 cm plate with cells around 70 to 80% confluence is selected by removing the old medium. The cells are washed once with PBS. The cells are then starved in low serum medium (RPMI-1640 containing 1% horse serum and 0.5% FBS with antibiotics) overnight. The next morning the medium is removed and the cells are washed with PBS. The cells are scraped off the plate, the cells are suspended perfectly in 2 ml of medium with low serum content. The number of cells is counted and more medium is added with low serum content to reach the final cell density of 5 x 10 5 cells / ml. 200 μl of the cell suspension is added to each well of the 96 well plate (equivalent to 1 x 105 cells / well). Add 50 μl of supernatant produced by Example 11, 37 ° C for 48 to 72 hours. As a positive control, one can use the growth factor known to activate PC12 cells through EGR, such as 50 ng / μl of Neuronal Growth Factor (NGF). An induction of more than fifty times of SEAP is typically observed in positive control wells. The supernatant is evaluated by SEAP according to Example 17.

Ex empl o 1 6: Allocation Test of Al to Rendimi on for the Acti vity of Cellos T NF-kappaB (Nuclear Factor kappaB) is a transcription factor activated by a wide variety of agents including the inflammatory cytokines IL-1 and TNF, CD30 and CD40, lymphotoxin a and lymphotoxin b, by exposure to LPS or thrombin, and by the expression of certain viral gene products. As a transcription factor, NF-kappaB regulates the expression of genes involved in the activation of immune cells, the control of apoptosis (NF-kappaB seems to protect cells from apoptosis), the development of B and T cells , antiviral and antimicrobial responses, and multiple responses to stress. Under non-stimulated conditions, NF-kappaB is retained in the cytoplasm with I-kappaB (kappaB inhibitor). However, after stimulation, I-kappaB is phosphorylated and degraded, causing NF-kappaB to be released into the nucleus, thereby activating the transcription of the target genes. The target genes activated by NF-kappaB include IL-2, IL-6, GM-CSF, ICAM-1 and MHC of class 1. Due to their central role and the ability to respond to a wide range of stimuli, constructions reporters using the NF-kappaB promoter element are used to select the supernatants produced in Example 11. Activators or inhibitors of NF-kappaB could be useful in the treatment of diseases. For example, inhibitors of NF-kappaB could be used to treat those diseases related to the acute or chronic activation of NF-kappaB, such as rheumatoid arthritis. To construct a vector containing the NF-kappaB promoter element, a PCR-based strategy is employed. The upstream primer (5 ') contains four tandem copies of the NF-kappaB binding site (5' -GGG GAC TTT CCC-3 '; I KNOW THAT. ID. NO: 20), 18 base pairs of sequence complementary to the 5 'end of the SV40 early promoter sequence, and flanked by an Xhol site: 5' -GCG GCC TCG AGG GGA CTT TCC CGG GGA CTT TCC GGG GAC TTT CCG GGA CTT TCC ATC CTG CCA TCT CAÁ TTA G-3 '(SEQ ID NO: 21). The downstream primer (3 ') is complementary to the 3' end of the SV40 promoter and is flanked by a HindlII site: 5 '-GCG GCA AGC TTT TTG CAA AGC CTA GGC-3' (SEQ ID NO: 22). PCR amplification is performed using the SV40 promoter template present in the plasmid pB-gal: promoter obtained from Clontech. The resulting PCR fragment is deferred with Xhol and ífindIII and subcloned into BLSK2- (Stratagene). Sequencing with T7 and T3 primers confirms that the insert contains the following sequence: 5'-CTCGAGGGGACTTTCCCGGGGACTTTCCGGGGACTTTCCGGGACTTTCCATC TGCCATCTCAATTAGTCAGCAACCATAGTCCCGCCCCTAACTCCGCCCATCC CGCCCCTAACTCCGCCCAGTTCCGCCCATTCTCCGCCCCATGGCTGACTAAT TTTTTTTATTTATGCAGAGGCCGAGGCCGCCTCGGCCTCTGAGCTATTCCAG AAGTAGTGAGGAGGCTTTTTTGGAGGCCTAGGCTTTTGCAAAAAGCTT-3 '(SEQ ID NO:.. 23). Then, the minimal promoter element SV40 present in the plasmid pSEAP2-promoter (Clontech) is replaced with this fragment NF-kappaB / SV40 using Xhol and JindiII. However, this vector does not contain a neomycin resistance gene, and therefore, it is not preferred for mammalian expression systems. In order to generate stable mammalian cell lines, the NF-kappaB / SV40 / SEAP cassette is removed from the aforementioned NF-kappaB / SEAP vector, using the SalI and No tl restriction enzymes, and inserted into a vector containing resistance to neomycin. Particularly, the cassette? F-kappaB / SV40 / SEAP was inserted into pGFP-1 (Clontech), replacing the GFP gene, after restricting pGFP-1 with Sali and No l.

Once the NF-kappaB / SV40 / SEAP / Neo vector is created, the stable Jurkat T cells are created and maintained according to the protocol described in Example 13. Similarly, the method for evaluating the supernatants with the T cells of Stable Jurkat is also described in Example 13. As a positive control, exogenous TNF-α (0.1, 1, 10 ng) is added to wells H9, H10 and Hll, with 5- to 10-fold higher activation typically observed.

Ex empl o 1 7: Essay for the Ac ti vi ity of SEAP As a reporter molecule for the assays described in Examples 13-16, SEAP activity is evaluated using the Tropix Phospho-light Kit (Cat. BP-400) according to the following general procedure. The Tropix Phospho-light Kit provides the Dilution, Test, and Reaction Shocks used later. A dispenser is prepared with the 2.5x Dilution Damper and 15 μl of buffer is supplied at a dilution of 2.5x inside the Optiplates (optical plates) containing 35 μl of a supernatant. The plates are sealed with a plastic sealer and incubated at 65 ° C for 30 minutes. The Optiplates are separated to avoid non-uniform heating. The samples are cooled to room temperature for 15 minutes. The spout is emptied and prepared with the Test Shock Absorber. 50 μl of Assay Buffer is added and incubated at room temperature for 5 minutes. The spout is emptied and prepared with the Reaction Damper (see the table below). 50 μl of Reaction Buffer is added and incubated at room temperature for 20 minutes. Since the intensity of the chemiluminescent signal is time dependent, and it takes approximately 10 minutes to read 5 plates on the luminometer, 5 plates should be treated at a time and the second group started 10 minutes later. The unit of relative light in the luminometer is read. H12 is set as blank and the results are printed. An increase in chemiluminescence indicates reporter activity.

Table III: Reaction Shock Formulation 60 3 11 65 3.25 12 70 3.5 13 75 3.75 14 80 4 15 85 4.25 16 90 4.5 17 95 4.75 18 100 5 19 105 5.25 20 110 5.5 21 115 5.75 22 120 6 23 125 6.25 24 130 6.5 25 135 6.75 26 140 7 27 145 7.25 28 150 7.5 29 155 7.75 30 160 8 31 165 8.25 32 170 8.5 33 175 8.75 34 180 9 35 185 9.25 36 190 9.5 37 195 9.75 38 200 10 39 205 10.25 40 210 10.5 41 215 10.75 42 220 11 43 225 11.25 44 230 11.5 45 235 11.75 46 240 12 47 245 12.25 48 250 12.5 49 255 12.75 50 260 13 Ex empl o 18: Al to Rendimi S ection Test to Identify the Changes in Concentration of Small Molecules and the Permeability of the Membrane It is known that the binding of a ligand to a receptor alters the intracellular levels of small molecules, such as calcium, potassium, sodium and pH, as well as alters the potential of the membrane. These alterations can be measured in an assay to identify supernatants that bind to the receptors of a particular cell. Although the following protocol describes an assay for calcium, this protocol can easily be modified to detect changes in potassium, in sodium, pH, in the membrane potential or in any other small molecule that is detectable by a fluorescent probe. The following test uses the Reader of Fluorometric Image Plate ("FLIPR") to measure changes in fluorescent molecules (Molecular Probes) that bind to small molecules. Clearly, any fluorescent molecule that detects a small molecule can be used instead of a fluorescent molecule to calcium, fluo-3, used here. For adherent cells, cells are seeded at 10,000-20,000 cells / well in a black 96-well Co-star plate with a clear background. The plate is incubated in a CO 2 incubator for 20 hours. The adherent cells are washed twice in the Biotek scrubber with 200 μl of HBSS (Hank's Balanced Salt Solution) leaving 100 μl of buffer after the final wash. A stock solution of 1 mg / ml fluo-3 is made in DMSO with 10% pluronic acid. To load the cells with fluo-3, 50 μl of 12 μl / ml of fluo-3 are added to each well. The plate is incubated at 37 ° C in a C02 incubator for 60 minutes. The plate is washed four times with the Biotek scrubber with HBSS leaving 100 μl of buffer. For non-adherent cells, the cells are centrifuged from the culture medium. The cells are re-suspended at 2-5 x 10 6 cells / ml with HBSS in a 50 ml conical tube. Four μl of 1 mg / ml of fluo-3 solution in DMSO with 10% pluronic acid are added to each ml of cell suspension. The tube is then placed in a 37 ° C water bath for 30 to 60 minutes. The cells are washed twice with HBSS, and resuspend at 1 x 106 cells / ml, and are dispensed into a microplate, 100 μl / well. The plate is centrifuged at 1000 rpm for 5 minutes. The plate is then washed once in Denley CellWash (Denley cell wash) with 200 μl, followed by an aspiration step to a final volume of 100 μl. For a non-cell-based assay, each well contains a fluorescent molecule, such as fluo-3. The supernatant is added to the well, and a change in fluorescence is detected. To measure the flow rate of intracellular calcium, the FLIPR is adjusted for the following parameters: (1) the gain of the system is 300-800 mW; (2) the exposure time is 0.4 seconds; (3) Chamber F / stop is F / 2; (4) Excitation is 488 nm; (5) the emission is 530 nm; and (6) the addition of the sample is 50 μl. The increased emission at 530 nm indicates an extracellular signaling event caused by the molecule, either IL-21 or a molecule induced by IL-21, which has resulted in an increase in intracellular Ca2 + concentration.

Ex empl o 19: Al to Rendimi Sel s Test to Identify the Tyrosine Activity - Kinase Protein tyrosine kinases (PTKs) represent a diverse group of transmembrane and cytoplasmic kinases. Within the receptor protein tyrosine kinase group are the receptors for a range of mitogenic and metabolic growth factors including PDGF, FGF, EGF, NGF, HGFa and subfamilies of the insulin receptor. In addition there is a large family of RPTKs for which the corresponding ligand is unknown. The ligands for the RPTKs mainly include small secreted proteins, but also intracellular matrix proteins bound to the membrane. Activation of RPTK by the ligands involves receptor dimerization, mediated by the ligand, resulting in transphosphorylation of receptor subunits and activation of cytoplasmic tyrosine kinases. Tyrosine kinases include tyrosine kinases associated with the receptor of the src family (eg, src, yes, l ck, lyn, fyn) and cytosolic protein tyrosine kinases and not linked to the receptor, such as the Jak family. , the members of which are mediators of signal transduction triggered by the cytokine receptor superfamily (e.g., Interleukins, Interferons, GM-CSF, and Leptin). Due to the wide range of known factors capable of stimulating tyrosine kinase activity, the identification of whether IL-21 or a molecule induced by IL-21 is able or not to activate the pathways of tyrosine signal transduction kinase, is of interest. Therefore, the following protocol is designed to identify such molecules capable of activating the pathways of tyrosine kinase signal transduction. Target cells (e.g., primary keratinocytes) are seeded at a density of approximately 25,000 cells per well in 96-well Loprodine Silent Selection Plates purchased from Nalge Nunc (Naperville, IL). The plates are sterilized with two 30-minute rinses with 100% ethanol, rinsed with water and dried overnight. Some plates are coated for 2 hours with 100 ml of type I collagen grade cell culture (50 mg / ml), gelatin (2%) or polylysine (50 mg / ml), all of which can be purchased from Sigma Chemicals (Saint L uis MO) or 10% of JMatrigel purchased from Becton Dickinson (Bedford, MA), or calf serum, rinsed _with -PBS and stored at 4 ° C. The depment of the cells on these plates is evaluated by sowing 5, 000 cells / well in the growth medium and indirect quantification of ^? of cells through the use of Alamar jajz.nl as described by the manufacturer Alamar Biosciences, Inc. (Sacramento, CA) after 48 hours. Falcon Plate Covers # 3071 from Becton Dickixson iBedford, MA) are used to fit the Loprodyne Silent Selection Plates. The jzelnlax. Falcon Microtest III culture plates can also be used in some proliferation experiments ... To prepare the extracts, A431 _se -sJ-etpbran cells on nylon footpipes of Loprodyne plates (20,000 / 200 mg / well) and are grown overnight in complete medium. the cells are at rest by incubation in serum-free basal medium for 24 hours. After 5-20 minutes, treatment with EGF (60 ng / ml) or 50 μl of the supernatant produced in Example 11, the medium was removed and 100 ml of extraction buffer was added (20 mM HEPE.S, pH 7 , 5, 0.15 M sodium chloride, 1% Triton X-100, 0.1% SDS, 2 mM Na3V04, 2 mM Na4P207 and a cocktail of inhibitors of pxotease (# 1836170) obtained from Boeheringer Mannheim (Indianapolis, IN) a each well, the plate is shaken on a rotary shaker for 5 minutes at 4 ° C. The extracts are collected in a 9-well trap / test well at the bottom of the vacuum tube immediately placed on ice. clarified by centrifugation, the content of each p.ozo, after solubilization with detergent for 5 minutes, is removed and centrifuged for 15 minutes at 4 ° C at 16,000 xg. Filtered extracts are tested for the activity levels of tyrosine-cina = a ^ Although many methods of detecting shooting activity ina-kinase are known, a method is described here. In general, the tyrosine kinase activity of a supernatant is evaluated by determining its ability to phosphorylate a tyrosine residue on a specific sustxate (a biotinylated peptide). Biotinylated peptides that can be used for this purpose include PSK1 (corresponding to amino acids 6-20 of the cell division kinase cdc2-p34) and PSK2 (corresponding to gastrin JL-17 amino acids). Both peptides are substrates for a wide range of tixosine kinases and are available from Boehringer Mannheim. The tyrosine kinase reaction is established by the addition of the following components, in order. Firstly, 10 μl of the Biotinylated Peptide 5 JJ.M, then 10 μl of ATP / Mg2 + (5 mM ATP / 50 mM MgCl2), then 10 μl of 5x Assay Buffer (40 mM imidazole hydrochloride, pH 7.3, b) are added. -glycerofsof to 0 m EGTA 1 mM, 100 mM magnesium chloride, 5 mM manganese chloride, 0.5 mg / ml BSA), then 5 μl of Sodium Vanadatp (1 mM), and then 5 μl of water. The components are mixed gently and the reaction mixture is preincubated at 30 ° C for 2 minutes. The reaction is initiated by the addition of 10 μl of the control enzyme or the filtered supernatant. The test-shot reaction, sina-kinase is then terminated by the addition of 10 μl of EDTA 120 mm and the reactions are placed on ice. The tyrosine kinase activity is determined by transferring an aliquot of 50 μl of the reaction mixture to a 3 I) module. microtiter plate (MTP) and incubating at 37 ° C for 20 minutes. This allows the 96-well plate coated with streptavidin to be associated with the biotinylated peptide. The MPT module is washed with 300 μl / well of PBS four times. Next, 75 μl of the anti-phosphotyrosine antibody conjugated to the horseradish peroxidase (anti-P-Tyr-POD (0.5 μl / ml)) is added to each ozo and incubated at 37 ° C for one hour. The well is layed as described above. Then 100 μl of the peroxidase substrate solution (Boehringer Mannheim) is added and incubated at room temperature for at least 5 minutes (up to 30 minutes). The absorbance of the sample is measured at 405 n using a yectox ELISA. The level of activity of bound peroxidase is quantified using an ELISA reader and reflects the level of tyrosine kinase activity.

Ex empl o 20 Assay of Allocation of Allocation Rendimi gue Iden tifi ca Acti vity of Fosforil aci ón As a potential alternative / or as a complement to the tyrosine kinase protein activity assay described in Example 19, an assay that detects the activation (phosphorylation) of the major intracellular signal transduction intermediates can be used. For example, as described below, a particular assay can detect the phosphorylation of tixosin of the Erk-1 and Erk-2 kinases. However, the phosphorylation of other molecules ^ such as Raf JNK ^, p38 MAP ,, the Map kinase (MEK), the MEK kinase, Src, the specific kinase of the Muscle ÍMuSK) j IRAK, Tec and Janus, as well as any other Phosphoserine molecule, phosphothyrinine, or phosphothreonine, can be detected by replacing these molecules with Erk-1 or Erk-2 in the following assay. Specifically, the assay plates are prepared by coating the wells of a 96-well ELISA plate with 0.1 ml of G protein (1 μg / ml.) For 2 hours at room temperature (RT). The plates are then rinsed with PBS and blocked with 3% BSA / PBS for 1 hour at room temperature. The G protein plates are then treated with two commercial monoclonal antibodies (100 ng / well) against Erk-1 and Erk-2. (1 hour at room temperature, available from Santa Cruz Biotechnolo.gy), To detect other molecules, this step can be easily modified by substituting a monoclonal antibody detecting any of the molecules 3D-8 previously described. After 3 to 5 rinses with JPBS, the plates are stored at 4 ° C until use. A431 cells are seeded at 20,000 / well in a 96 well Loprodyne filter plate and grown overnight in growth medium. The cells are then subjected to starvation for 48 hours in basal medium (DMEM) and then treated with EGF (6 ng / well or 50 μl of the supernatants obtained in Example 11 for 5 to 20 minutes) The cells are then solubilized and the extracts are filtered directly into the test plate.After incubation with the extract for 1 hour at room temperature, the wells are again rinsed, As a positive control, a commercial preparation of MAP kinase (10 ng / well) is used. Instead of the A431 extract, the plates are then treated with a commercial polyclonal antibody (rabbit) (1 .μg / ml) that specifically recognizes the phosphorylated epitope of the Erk-1 and Erk-2 kinases (1 hour at room temperature). This antibody is biotinylated by standard procedures.The polyclonal antibody binding is then quantified by successive incubations with Europium-streptavidin and the 303-augmenting reagent.

Europium fluorescence in the Wallac DELFIA instrument (fluorescence resolved by time). An increased fluorescent signal on the foreground indicates a phosphorylation by 11-21 or a molecule induced by IL-21.

Example 21: Method of Determination of Alterations in Gene I -21 RNA isolated from whole families or from individual patients who present a phenotype of interest (such as a disease) is isolated. The cDNA is then generated from these RNA samples using protocols known in the art (see, Sambrook et al., Supra). The cDNA is then used as a template for PCR, employing primers that surround the regions of interest in the SEQ. ID. NO: 1. Suggested PCR conditions consist of 35 cycles at 95 ° C for 30 seconds, - 60-120 seconds at 52-58 ° C; and 60-120 seconds at 70 ° C, using the buffer solutions described by Sidransky et al. (Sci en 252: 706 (1991)). The PCR products are then sequenced using the primers labeled at the 5 'end with the T4 polynucleotide kinase, using the SequiTJaexm Polymerase (Epicenfxe Tecnnologies). The intron-exon boundaries of the selected exons of IL-21 are also detexinized and the genomic PCR pipelines are analyzed to confirm the results. The PCR products harboring the suspected mutations in IL-21 are then cloned and sequenced to give the results of direct sequencing. The IL-21 PCR products are cloned into the T-designed vectors as described by Holton and Gxaham [Nucí, Aci ds, Res. 19: 1156 (1991)) and sequenced with T7 polymerase (United States Biochemical). Affected individuals are identified by IL-21 mutations not present in unaffected individuals. Genomic rearrangements are also observed as a method to detexminax alterations in the IL-21 gene. The genomic clones isolated according to Example 2 are pseudotransducted with digoxigenindeoxyuridine 5'-triphosphate (Boehringer Manheim, J, and FISH performed as described by Jonson et al. (Me th ods Cel l Bi., 35.- 13 -99 (1991 )) Hybridization with the labeled probe is carried out using a large excess of cot-1 DNA for specific hybridization of the IL-21 genomic locus.The chromosomes are counterstained with 4-6-diamino-2-phenylindole and iodide propidio, producing a combination of bands C and R. The images aligned for the precise mapping of the map are obtained using a three-band filter group (Chroma Technology, Grattleboro, VT) in combination with a charge-coupled device camera, cooled (Photometrics, Tucson, AZ) and variable excitation wavelength filters IJon = on, C. et al., Genet, Anal. Tech. Appl. 8: 75 (1991)). The image collection, the analysis and the chromosomal fractional length measurements are carried out using the ISee Graphical Program System. (Inovision Corporation, Durham, NC). Chromosomal alterations of the genomic region of IL-21 (hybridized by the probe) are identified as insertions, deletions, and translocations. These alterations of IL-21 are used as a diagnostic marker for an associated disease.

Ex empl o 22: Detection method of the Ni-level is abnormal of IL-21 in a Biological Sample IL-21 polypeptides can be detected in a biological sample, and if an increased or decreased level of IL-21 is detected, this polypeptide is a marker for a particular phenotype. The detection methods are numerous, and thus, it is understood that a person skilled in the art can modify the following test to suit his particular needs. For example, antibody sandwich ELISAs are used to detect 11-21 in a sample, preferably a biological sample. The wells of a microtitre plate are coated with antibodies specific for IL-21, at a final concentration of 0.2 to 10 μg / ml. The antibodies are either monoclonal or polyclonal and are produced by the method described in Example 10. The wells are blocked so that the non-specific binding of IL-21 to the well is reduced. The coated wells are then incubated by > 2 hours at room temperature with a sample containing IL-21. Preferably, serial dilutions of the sample should be used to validate the results. The plates are then washed three times with deionized or distilled water to remove unbound IL-21. Then, 50 μl of the specific antibody-alkaline phosphatase conjugate, at a concentration of 25-400 ng, is added and incubated for 2 hours at room temperature. The plates are again washed three times with deionized or distilled water to remove the unbound conjugate. 75 x1 of the 4-methylumbelliferyl phosphatase (MUP) or the p-nitrophenyl phosphate substrate solution (NPP) are added to each well and incubated 1 hour at room temperature. The reaction is measured by a microtiter plate reader. A standard curve is prepared, using serial dilutions of a control sample, and the concentration of the IL-21 polypeptide on the X-axis (logarithmic scale) and the fluorescence or absorbance of the Y-axis (linear scale) are plotted. The concentration of IL-21 in the sample is interpolated using the standard curve.

Ex empl o 23: Formulation of a Polirjéti do The composition of 11-21 will be formulated and dosed in a manner consistent with good medical practice, taking into account the clinical condition of the individual patient (especially the side effects of treatment with the IL-2.1 polypeptide on the administration site, the method of administration, administration scheme, and other factors known to medical practitioners The "effective amount" for purposes of this is thus determined by such considerations As a general proportion, the total pharmaceutically effective amount of IL-21 parenterally administered per dose will be in the range of about 1 μg / kg / day to 10 mg / kg / day of the patient's body weight, although, as noted above, this will be subject to therapeutic discretion. less than 0.01 mg / kg / day, and more preferably for humans between about 0.01 and 1 mg / kg / day for the hormone. is administered continuously, IL-21 is typically administered at a dose rate of about 1 μg / kg / hour to about 50 μg / kg / hour, either for 1 to 4 injections per day or for continuous subcutaneous infusions, for example, , using a mini-pump. An intravenous bag solution can also be used. The length of the treatment, necessary to observe changes, and the following treatment interval so that O-c x the response, seems to vary depending on the desired effect. The pharmaceutical compositions contain 11-21 are administered, orally, rectally, parenterally, intracisternally, intravaginally, intraperitoneally, topically (such as powders, ointments, gels, drops or transdermal patch), Jbncalmente, or as an oral or nasal spray. The "pharmaceutically acceptable carrier" refers to a solid filler., semi-solid, or non-toxic liquid, a diluent, encapsulation material or formulation aid of any kind. The term "parenteral" as used herein refers to modes of administration that include intravenous, intramuscular, intraperitoneal, intrasternal, subcutaneous and intraarticular injection and infusion. IL-21 is also properly administered by sustained release systems. Suitable examples of sustained release compositions include semi-permeable polymer matrices in the form of shaped articles, eg, films or jaicrocapsules. Sustained-release matrices include polylactides (U.S. Pat. No. 3, 773, .91.9, European Patent EP-58,881), L-glutamic acid copolymers and gamma-ethyl-L-glutamate iSidman, U. et al., Bi opolymers 22. 547-556 (1983)), poly (2-hydroxyethyl methacrylate) (Langer, R, Et al., J. Bi. Or Ed. Ma ter. Res. 15: 167- 277 (1981); Langer, R. Ch em. Tech. 12: 98-105-11982).), Ethylene vinyl acetate iLanger, H. Et al.) Or poly-D- (-) -3-hydroxybutyric acid (European Patent EP-133,988). Sustained-release compositions include liposomally entrapped IL-21 polypeptides. Liposomes containing 1I > -21 are prepared by methods known per se (German Patent JDE-3, 218, 121; Epstein et al., PX C ^ Na ti. Acad. Sci. USA 82: 3688-3692 (1985); Hwang et al-, Pxoc, Na tl ^? Cad ^ Sci. USA 11 4D3D-4D34 _ (L98Q) European Patents EP-52,322; EP-36,676; EP-88,046; EP-143, 949; EP-142, 641, - Patent Application. Japan 83-118008; U.S. Patent Nos. 4,485,045 and 4,544,545, and European Patent EP-102, 324). Ordinarily, liposomes are of the small unilamellar type (approximately 200-800 Angstroms) in which the lipid content is greater than about 30% mol of cholesterol, the selected proportion being adjusted for optimal secreted polypeptide therapy. For parenteral administration, in one embodiment, IL-21 is formulated in general by mixing it to the desired degree of purity, in a unit dose injectable form (solution, suspension or emulsion), with a pharmaceutically acceptable carrier, by example, one that is non-toxic to patients at the doses and concentrations employed, and that is compatible with other ingredients of the formulation. For example, the formulation preferably does not include oxidizing agents and other compounds that are known to be harmful to polypeptides. In general, formulations are prepared by contacting IL-21 uniformly and intimately with liquid carriers or solid carriers. finely divided, or both. Then, if necessary, the product is shaped into the desired formulation. Preferably, the carrier is a parenteral carrier, more preferably a solution that is isotonic with the patient's blood.

Examples of such carriers include water, saline, Ringer's solution, and dextrose solution. Non-aqueous vehicles such as fixed oils and ethyl oleate are also useful here, as well as liposomes. The carrier suitably contains minor amounts of additives such as substances that increase isotonicity and chemical stability. Such materials are non-toxic to patients at the doses and concentrations employed, and include buffers such as phosphate, citrate, succinate, acetic acid, and other organic acids or their salts; antioxidants such as ascorbic acid; low molecular weight polypeptides (less than about ten residues), eg, polyarginine or tripeptides, - proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids, such as glycine, glutamic acid, aspartic acid, or arginine; monosaccharides, disaccharides and other carbohydrates including cellulose or its derivatives, glucose, mannose or dextrins; chelating agents such as -EDTA sugar alcohols such as mannitol or sorbitol; counterions such as sodium; and / or anionic surfactants such as polysorbates, poloxamers, or PEG. IL-21 is typically formulated in such vehicles at a concentration of about 0.1 mg / ml to 100 mg / ml, preferably 1-10 mg / ml, at a pH of about 3 to 8, It will be understood that the use of some of the aforementioned excipients, carriers or stabilizers,? axá as a result the formation of polypeptide salts. IL-21 used for therapeutic administration may be sterile. Sterility is easily elaborated by filtration through sterile filtration membranes (eg, 0.2 micron membranes). Therapeutic polypeptide compositions are generally placed in a container having a sterile access gate, for example, a solution bag. intravenous or a bottle that has a plug pierced by a hypodermic injection needle. IL-21 polypeptides will ordinarily be stored in single-dose or multi-dose containers, for example, sealed vials or flasks, as an aqueous solution or as a lyophilized formulation for reconstitution. As an example of a lyophilized formulation, fill a 10 ml bottle with 5 ml of polynucleotide solution = 1% aqueous IL / 21 v / v. sterilized by filtration, and the resulting mixture is lyophilized. The infusion solution is prepared by reconstituting the lyophilized IL-21 polypeptide by using Water For Bacteriostatic Injection (WFI). The invention also provides a pharmaceutical package or kit comprising one or more containers filled with one or more of the ingredients of the pharmaceutical compositions of the invention. Associated with such containers may be a notice in the form of a government agency that regulates the manufacture, use or sale of fax products or biological products, the notice of which reflects the agency's approval of the manufacture, use or sale of the product. human administration. In addition, IL-21 can be used in conjunction with other therapeutic compounds, Ex empl o 24: N amalgama ting Method is Dimin ated of IL-21 The present invention relates to a method for treating an individual in need of a decreased level of IL-21 activity in the body, comprising administering to such an individual, a composition comprising a therapeutically effective amount of the IL- antagonist. twenty-one. Preferred antagonists for use in the present invention are antibodies specific for IL-21. In addition, it will be appreciated that conditions caused by a decrease in the standard or normal expression level of IL-21 in an individual can be treated by administering ± and il-1, preferably in the secreted form. Thus, the invention also provides a method of treating an individual in need of an increased level of IL-21 polypeptide, which comprises administering to such an individual a pharmaceutical composition comprising an amount of IL-21 to increase the level of IL-21 activity in such an individual. For example, a patient with decreased levels of the IL-21 polypeptide receives a daily dose of 0.1-100 μg / kg of the polypeptide for six consecutive days. Preferably, the polypeptide is in the secreted form. The exact details of the dosage scheme, based on the administration of the formulation are given in Example 23.

Ex empl o 25: N ov e tra ting Method is Increased from I -21 The present invention also relates to a method for treating an individual in need of an increased level of IL-21 activity in the body, comprising administering to such an individual a composition comprising a therapeutically effective amount of IL-21 or a agonist thereof. Antisense technology is used to inhibit IL-21 production. This technology is an example of a method to decrease the levels of IL-21 polypeptide, preferably a secreted fox, due to a variety of etiologies, such as cancer. . For example, a patient diagnosed with abnormally unrecorded levels of 11-21 years administered intravenously with antisense polynucleotides at 0.5, 1.0, 1.5, 2.0 and 3.0 mg / kg per day for 21 days. This treatment is repeated after a rest period of 7 days if the treatment was well tolerated. The formulation of the antisense polynucleotide is provided in Example 23.

Ex empl o 26: Method of Tra tami in Medi an te the Use of Terapi a Gen i ca One method of gene therapy is the transplants of fibroblasts, which are capable of expressing IL-21 polypeptides, in a patient. In general, fibroblasts are obtained from a subject by skin biopsy. The resulting tissue is placed in a tissue culture medium and separated into small pieces. The small traces of the tissue are placed on a wet surface of a flask for tissue culture, approximately 10 pieces are placed in each flask. The flask is turned down, hermetically sealed and left at room temperature overnight. After 24 hours at room temperature, the flask is inverted and the pieces of tissue remain fixed to the bottom of the flask and fresh medium is added (for example Ham's F12 medium, with 10% FBS, with penicillin and streptomycin). The flasks are then incubated at 37 ° C for about a week.

At this time, fresh media is added and subsequently changed every few days.

After two additional weeks in culture, a monolayer of fibroblasts emerges. The monolayer is trypsinized and changed to larger flasks. pMV-7 (Kirschmeier, PT et al., DNA 1: 219-25 (1988)), flanked by the large end repeats of Moloney murine sarcoma virus, is digested with £ coRI and HindIII and subsequently treated with intestinal phosphatase. veal. The linear vector is fractionated on agarose gel and purified, using glass spheres. The cDNA encoding IL-21 can be amplified using PCR primers which correspond to the sequence of the 5 'and 3' ends respectively as described in Example 1. Preferably, the 5 'primer contains an .EcoRI site and the 3' primer includes a HindI I site. Equal amounts of the linear backbone of Moloney murine sarcoma virus and the amplified £ coRI and Hino'III fragment are aggregated together in the presence of the T4 DNA ligase. The resulting mixture is maintained under conditions appropriate for the ligation of the two fragments. The ligation mixture is then used to transform HB101 bacteria, which are then plated on agar containing kanamycin, for purposes of confirming that the vector contains properly inserted IL-21. Amphotropic packaging cells pA317 or GP + aml2 are grown in tissue culture to the confluent density in the Dulbecco Modified Eagle Medium (DMEM) with 10% calf serum (CS), penicillin and streptomycin. The MSV vector containing the IL-21 gene is then added to the medium and the packaging cells transduced with the vector. The packaging cells now produce infectious viral particles that contain the IL-21 gene (packaging cells are now referred to as producer cells). Fresh medium is added to the translated producer cells, and subsequently, the medium is harvested from a 10 cm plate of the confluent producer cells. The medium based, which contains the infectious viral particles, is filtered through a millipore filter to eliminate the detached production cells, and this medium is then used to infect fibroblast cells. Media is removed from a subconfluent plate of the fibroblasts and rapidly replaced with the medium from the producer cells. This medium is removed and replaced with fresh medium. If the virus titer is high, it is high, then virtually all fibroblasts will be infected and selection is not required. If the titer is very low, then it is necessary to use a retroviral vector that has a selectable marker, such as neo or his. Once the fibroblasts have been efficiently infected, the fibroblasts are analyzed to determine whether the IL-21 protein is produced or not. The genetically engineered fibroblasts are then transplanted onto the host, either alone or after being developed to confluence on cytodex 3 microcarrier spheres. It will be clear that the invention can be practiced otherwise than is particularly described in the description and previous examples. Numerous modifications and variations of the present invention are possible in light of the above teachings and, therefore, are within the scope of the appended claims.

The complete description of each cited document (including patents, patent applications, journal articles, extracts, laboratory manuals, books or other descriptions) in the background of the invention, the detailed description and the examples, are incorporated by reference in the I presented. In addition, the sequence listing sent with the present and each of the sequence listings sent with the provisional application of the United States Serial No. 60 / 087,340, filed May 29, 1998, the co-pending North American Provisional Application No. of Series 60 / 099,805, filed on September 10, 1998, and the Co-pending Provisional North American application No. of Series 60 / 131,965, filed on April 30, 1999 (for each of which the present application claims the benefit of the filing dates of 35 USC section 119 (e)), in forms written on paper and computer readable are incorporated herein by reference in its entirety. It is noted that in relation to this date, the best method known to the applicant to carry out the aforementioned invention, is that which is clear from the present description of the invention.

Claims

CLAIMS Having described the invention as above, the content of the following claims is claimed as property:

1. An isolated nucleic acid molecule, characterized in that it comprises a polynucleotide having a nucleotide sequence at least 95% identical to a sequence selected from the group consisting of: a) a polynucleotide fragment of SEQ. ID. NO: 1 or a polynucleotide fragment of the cDNA sequence included in the ATCC Deposit No. 209666; b) a polynucleotide encoding a polypeptide fragment of SEQ. ID. NO: 2 or the cDNA sequence included in the ATCC Deposit No. 209666; c) a polynucleotide encoding the conserved polypeptide I domain of SEQ. ID. NO: 2 or the cDNA sequence included in the ATCC Deposit No. 209666; d) a polynucleotide encoding the conserved polypeptide II domain of SEQ. ID. NO: 2 or the cDNA sequence included in the ATCC Deposit No. 209666; e) a polynucleotide encoding the polypeptide III domain deleted, SEQ. ID. NO: 2 or the cDNA sequence included in the ATCC Deposit No. 209666; f) a polynucleotide encoding the conserved polypeptide IV domain of SEQ. ID. NO: 2 or the cDNA sequence included in the ATCC Deposit No. 209666; g) a polynucleotide encoding a polypeptide epitope of SEQ. ID. NO: 2 or the cDNA sequence included in ATCC Deposit No. 209666; h) a polynucleotide encoding a polypeptide of SEQ. ID. NO: 2 or the cDNA sequence included in the ATCC Deposit No. 209666, which has biological activity; i) a polynucleotide that is a variant of SEQ. ID. NO: 1; j) a polynucleotide that is an allelic variant of SEQ. ID. NO: 1; k) a polynucleotide encoding a homologous species of the polypeptide whose amino acid sequence is shown in SEQ. ID. NO: 2; 1) a polynucleotide capable of hybridizing under stringent conditions to any of the polynucleotides specified in (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), or (k), wherein the polynucleotide does not hybridize under stringent conditions to a nucleic acid molecule having a nucleotide sequence only waste A or only waste T; and m) a polynucleotide that is the complement of any of the polynucleotides specified in (a), (b), (c), (d), (e), (f), (g), (h), (i) , (j), (k) or (m).

2. An isolated nucleic acid molecule, characterized in that it comprises a polynucleotide having a nucleotide sequence at least 95% identical to a sequence selected from the group consisting of: a) a polynucleotide fragment of SEQ. ID. NO: 28; b) a polynucleotide encoding a polypeptide fragment of SEQ. ID. NO: 28, - c) a polynucleotide encoding the conserved polypeptide I domain of SEQ. ID. NO: 28; d) a polynucleotide encoding the polypeptide II domain conserved from SEQ. ID. NO: 28; e) a polynucleotide encoding the polypeptide III domain conserved from SEQ. ID. NO: 28; f) a polynucleotide encoding the polypeptide IV domain conserved from SEQ. ID. NO: 28; g) a polynucleotide encoding the conserved polypeptide V domain of SEQ. ID. NO: 28; h) a polynucleotide encoding the polypeptide VI domain conserved from SEQ. ID. NO: 28; i) a polynucleotide encoding the polypeptide VII domain conserved from SEQ. ID. NO: 28; j) a polynucleotide encoding a polypeptide epitope of SEQ. ID. NO: 28; k) a polynucleotide encoding a polypeptide of SEQ. ID. NO: 28 that has biological activity; 1) a polynucleotide that is a variant of SEQ. ID. NO: 28; m) a polynucleotide that is an allelic variant of SEQ. ID. NO: 28; n) a polynucleotide that codes for a homologous species of the polypeptide whose amino acid sequence is shown in SEQ. ID. NO: 28; o) a polynucleotide capable of hybridizing under stringent conditions to any of the polynucleotides specified in (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (1), (m), or (n), wherein the polynucleotide does not hybridize under conditions strictures to a nucleic acid molecule having a nucleotide sequence residing only A or residues T only; p) a polynucleotide that is the complement of any of the polynucleotides specified in (a), (b), (c), (d), (e), (f), (g), (h), (i), (j), (k), (1), (), (n), u (o).

3. An isolated nucleic acid molecule, characterized in that it comprises a polynucleotide having a nucleotide sequence at least 95% identical to a sequence selected from the group consisting of: a) a polynucleotide fragment of SEQ. ID. NO: 3 or a polynucleotide fragment of the cDNA sequence included in the ATCC Deposit No. 209665; b) a polynucleotide encoding a polypeptide fragment of SEQ. ID. NO: 4 or the cDNA sequence included in the ATCC Deposit No. 299665; c) a polynucleotide encoding the conserved polypeptide I domain of SEQ. ID. NO: 4 or the cDNA sequence included in the ATCC Deposit No. 209665; d) a polynucleotide encoding the conserved polypeptide II domain of SEQ. ID. NO: 4 or the cDNA sequence included in the ATCC Deposit No. 209665; e) a polynucleotide encoding the conserved polypeptide III domain of SEQ. ID. NO: 4 or the cDNA sequence included in the ATCC Deposit No. 209665; f) a polynucleotide encoding the conserved polypeptide IV domain of SEQ. ID. NO: 4 or the cDNA sequence included in the ATCC Deposit No. 209665; g) a polynucleotide encoding a polypeptide epitope of SEQ. ID. NO: 4 or the cDNA sequence included in the ATCC Depósio No. 209665; h) a polynucleotide encoding a polypeptide of SEQ. ID. NO: 4 or the cDNA sequence included in the ATCC Deposit No. 209665 having biological activity; i) a polynucleotide that is a variant of SEQ. ID. NO: 3; j) a polynucleotide that is an allelic variant of SEQ. ID. NO: 3; k) a polynucleotide encoding a homologous species of the polypeptide whose amino acid sequence is shown in SEQ. ID. NO: 4; 1) a polynucleotide capable of hybridizing under stringent conditions to any of the polynucleotides specified in (a), (b), (c), (d), (e), (f), (g), (h), (i), (j) or (k), wherein the polynucleotide does not hybridize under stringent conditions to the nucleic acid molecule having a nucleotide sequence only of waste A or only waste T; and m) a polynucleotide that is the complement of any of the polynucleotides specified in (a), (b), (c), (d), (e), (f), (g), (h), (i) , (j), (k) or (1).

4. The isolated nucleic acid molecule according to claim 1, characterized in that the polynucleotide fragment comprises a nucleotide sequence that codes for a mature form or a secreted protein.

5. The isolated nucleic acid molecule according to claim 2, characterized in that the polynucleotide fragment comprises a nucleotide sequence that codes for a mature form or a secreted protein.

6. The isolated nucleic acid molecule according to claim 3, characterized in that the polynucleotide fragment comprises a nucleotide sequence that codes for a mature form or a secreted protein.

7. The isolated nucleic acid molecule according to claim 1, characterized in that the polynucleotide fragment comprises a nucleotide sequence that codes for the sequence identified as SEQ. ID. NO: 2 or the coding sequence included in the ATCC Deposit No. 209666.

8. The isolated nucleic acid molecule according to claim 2, characterized in that the polynucleotide fragment comprises a nucleotide sequence that codes for the sequence identified as SEQ. ID. NO: 28

9. The nucleic acid molecule according to claim 3, characterized in that the polynucleotide fragment comprises a nucleotide sequence coding for the sequence identified as SEQ. ID. NQ: 4 or the coding sequence included in the ATCC Deposit No. 209665.

10. The isolated nucleic acid molecule according to claim 1, characterized in that the polynucleotide fragment comprises the complete nucleotide sequence of SEQ. ID. NO: 1 or the cDNA sequence included in the ATCC Deposit No. 209666.

11. The isolated nucleic acid molecule according to claim 2, characterized in that the polynucleotide fragment comprises the complete nucleotide sequence of SEQ. ID. NO: 28

12. The isolated nucleic acid molecule according to claim 3, characterized in that the polynucleotide fragment comprises the complete nucleotide sequence of SEQ. ID. NO: 3 or the cDNA sequence included in the ATCC Deposit No. 209665.

13. The isolated nucleic acid molecule according to claim 5, characterized in that the nucleotide sequence comprises the sequential nucleotide deletions either of the C-terminus or the N-terminus.

14. The isolated nucleic acid molecule according to claim 8, characterized in that the nucleotide sequence comprises the sequential nucleotide deletions of either the C-terminus or the N-terminus.

15. A recombinant vector, characterized in that it comprises the isolated nucleic acid molecule according to claim 2.

16. A method for the preparation of a recombinant host cell, characterized in that it comprises the isolated nucleic acid molecule according to claim 1.

17. A method for the preparation of a recombinant host cell, characterized in that it comprises the isolated nucleic acid molecule according to claim 2.

18. A method for the preparation of a recombinant host cell, characterized in that it comprises the isolated nucleic acid molecule according to claim 3.

19. A recombinant host cell, characterized in that it is produced by the method according to claim 16.

20. A recombinant host cell, characterized in that it is produced by the method according to claim 17.

21. A recombinant host cell, characterized in that it is produced by the method according to claim 18.

22. The recombinant host cell according to claim 19, characterized in that it comprises vector sequences.

23. The recombinant host cell according to claim 20, characterized in that it comprises vector sequences.

24. The recombinant host cell according to claim 21, characterized in that it comprises vector sequences.

25. An isolated polypeptide, characterized in that it comprises at least 95% amino acid sequence identical to a sequence selected from the group consisting of: a) a polypeptide fragment of SEQ. ID. NO: 2 or the encoded sequence included in the ATCC Deposit No. 209666; b) a polypeptide fragment of SEQ. ID. NO: 2 or the encoded sequence included in the ATCC Deposit No. 209666 having biological activity; c) a polypeptide domain of SEQ. ID. NO: 2 or the encoded sequence included in the ATCC Deposit No. 209666; d) a polypeptide epitope of SEQ. ID. NO: 2 or the sequence codes included in the ATCC Deposit No. 209666; e) a mature form of a secreted protein; f) a full-length secreted protein; g) a variant of SEQ. ID. NO: 2; and h) an allelic variant of SEQ. ID. NO: 2; and i) a homologue of species of the SEQ. ID. NO: 2

26. An isolated polypeptide, characterized by an at least 95% amino acid sequence identical to a sequence selected from the group consisting of: a) a polypeptide fragment of SEQ. ID. NO: 29; b) a polypeptide fragment of SEQ. ID. NO: 29 that has biological activity; c) a polypeptide domain of SEQ. ID. NO: 29; d) a polypeptide epitope of SEQ. ID, NO: 29; e) a mature form of a protein secreted from SEQ. ID. NO: 29; f) a full-length secreted protein of SEQ. ID. NO: 29; g) a variant of SEQ. ID. NO: 29; h) an allelic variant of SEQ. ID. DO NOT: 29; and a homologue of species of the SEQ. ID. NO: 29

27. An isolated polypeptide, characterized in that it comprises at least 95% amino acid sequence identical to a sequence selected from the group consisting of: a) a polypeptide fragment of SEQ. ID. NO: 4 or the encoded sequence included in the ATCC Deposit No. 209665; b) a polypeptide fragment of SEQ. ID. NO: 4 or the encoded sequence included in the ATCC Deposit No. 209665 that has biological activity; c) a polypeptide domain of SEQ. ID. NO: 4 or the encoded sequence included in the ATCC Deposit No. 209665; d) a polypeptide epitope of SEQ. ID. NO: 4 or the encoded sequence included in the ATCC Deposit No. 209665; e) a mature form of a secreted protein; f) a full-length secreted protein; g) a variant of SEQ. ID. NO: 4; h) an allelic variant of SEQ. ID. NO: 4; and i) a homologue of species of the SEQ. ID. NO: 4

28. The isolated polypeptide according to claim 26, characterized in that the mature form or the full-length secreted protein comprises the deletions of sequential amino acids either from the C-terminus or the N-terminus.

29. An isolated antibody, characterized in that it binds specifically to the isolated polypeptide according to claim 26.

30. A recombinant host cell, characterized in that it expresses the isolated polypeptide according to claim 25.

31. A recombinant host cell, characterized in that it expresses the isolated polypeptide according to claim 26.

32. A recombinant host cell, characterized in that it expresses the isolated polypeptide according to claim 27.

33. A method for making an isolated polypeptide, characterized in that the method comprises: a) cultivating the recombinant host cell according to claim 30 under conditions such that the polypeptide is expressed; and b) recovery of the polypeptide.

34. A method for making an isolated polypeptide, characterized in that the method comprises: a) the culture of the recombinant host cell according to claim 31 under conditions such that the polypeptide is expressed; and b) recovery of the polypeptide.

35. A method for making an isolated polypeptide, characterized in that the method comprises: a) the culture of the recombinant host cell according to claim 32, under conditions such that the polypeptide is expressed; and b) recovery of the polypeptide.

36. The polypeptide, characterized in that it is produced in accordance with claim 34.

37. A method for preventing, treating or improving a medical condition, characterized in that the method comprises administering to a mammalian subject a therapeutically effective amount of the polypeptide according to claim 26.

38. A method for diagnosing a pathological condition or a susceptibility to a pathological condition in a subject, related to the expression or activity of a secreted protein, characterized the method because it comprises: a) determining the presence or absence of a mutation in the polynucleotide in accordance with claim 2; b) diagnose a pathological condition or a susceptibility to a pathological condition, based on the presence or absence of said mutation.

39. A method for diagnosing a pathological condition or a susceptibility to a pathological condition in a subject, related to the expression or activity of a secreted protein, characterized in the method because it comprises: a) determining the presence or amount of expression of the polypeptide in accordance with the claim 26, in a biological sample; b) diagnosing a pathological condition or a susceptibility to a pathological condition, based on the presence or amount of expression of the polypeptide.

40. A method for identifying the polypeptide binding partner according to claim 26, characterized in that it comprises: a) contacting the polypeptide according to claim 26 with a binding partner; and b) determining whether the link partner performs a polypeptide activity.

41. The gene, characterized in that it corresponds to the cDNA sequence according to SEQ. ID. NO: 1.

42. The gene, characterized in that it corresponds to the cDNA sequence according to SEQ. ID. NO: 28

43. The gene, characterized in that it corresponds to the cDNA sequence according to SEQ. ID. NO: 3

44. A method for identifying an activity in a biological assay, characterized in the method because it comprises: a) expressing the SEQ. ID. NO: 1 in a cell; b) isolate the supernatant; c) detect an activity in a biological assay; and d) identify the protein that has the activity, in the supernatant.

45. A method for identifying an activity in a biological assay, characterized in the method because it comprises: a) expressing the SEQ. ID. NO: 28 in a cell; b) isolate the supernatant; c) detect an activity in a biological assay; and d) identify the protein that has the activity, in the supernatant.

46. A method for identifying an activity in a biological assay, characterized in the method because it comprises: a) expressing the SEQ. ID. NO: 3 in a cell; b) isolate the supernatant; c) detect an activity in a biological assay; and d) identify the protein that has the activity, in the supernatant.

47. The product, characterized in that it is produced by the method according to claim 44.

48. The product, characterized in that it is produced by the method according to claim 45.

49. The product, characterized in that it is produced by the method according to claim 46.