MXPA97004618A - Receiver of the interleucine-11 hum - Google Patents

Abstract

The present invention relates to polynucleotides that encode the human IL-11 receptor and fragments thereof. Also described are IL-11 receptor proteins, methods for their production, inhibitors of the binding of human IL-11 and its receptor, and methods for its identification.

Description

RECEPTOR OF INTERLEUCINE-11 HUMAN Field of the Invention The present invention relates to the human interleukin-11 receptor, fragments thereof and recombinant polynucleotides and cells useful for the expression of such proteins.

Background of the Invention A variety of regulatory molecules, known as cytokines, have been identified including interleukin-11 (IL-11). The various protein forms of IL-11 and the DNA encoding various forms of IL-11 activity are described in Bennett, et al., U.S. Pat. 5,215,895 (June 1, 1993); McCoy, et al., U.S. Patent. 5,270,181 (December 14, 1993); and McCoy, et al., U.S. Pat. 5,292,646 (March 8, 1994), all incorporated herein by reference. Thus, the term "IL-11" includes proteins that have the biological activity described in these patents, whether produced by recombinant genetic engineering techniques; purified from cellular sources that produce the factor naturally or by induction with other factors, or synthesized by chemical techniques; or a combination of these. IL-11 is a pleiotropic cytokine that has been implicated in the production of various biological activities including: the induction of proliferation of hematopoietic and multipotential progenitor cells (Musashi et al (1991) Blood, 78, 1448-1451); the intensification of megakaryocyte and platelet formation (Burstein et al, (1992) J. Cell. Physiol., 153, 305-312); the stimulation of critical phase protein synthesis (Baumann et al (1991) J. Biol. Chem., 266, 20424-20427); the inhibition of the lipoprotein adipocyte lipase activity (Kawashima et al., (1991) FEBS Lett., 283, 199-202); and the effects on the neurotransis phenotype (Fann et al., (1994) Proc. Nati, Acad. Sci. USA, 91, 43-47).

IL-11 can be used in a pharmaceutical preparation or formulation to treat immunological deficiencies, specifically defects in hematopoietic progenitor cells, or disorders related thereto. The treatment of the other disorders or the stimulation of the immune systems of these cells can also employ IL-11. IL-11 can also be used in methods for the treatment of cancer and other diseases. Such pathological conditions may result from disease, radiation exposure or medications and include, for example, leukopenia, bacterial and viral infections, anemia, T cell or B cell deficiencies such as deficiencies in hematopoietic cells or immune cells following a liver transplant. bone marrow. IL-11 can also be used to potentiate the immune response to a variety of vaccines by creating a longer-lasting and more effective immunization. The therapeutic treatment of cancer and other diseases with IL-11 can avoid the undesirable side effects caused by the treatment with currently available drugs. Like most cytokines, IL-11 exhibits certain biological activities by interacting with an IL-11 receptor (IL-11 R) on the surface of target cells. It would be desirable to identify and clone the sequence for the human receptor so that the IL-11R proteins can be produced for various reasons, including the production of therapeutics and the classification for inhibitors of IL-11 binding to the receptor and the signage of the receiver.

OBJECTIVES OF THE INVENTION In accordance with the present invention, polynucleotides encoding the human interleukin-11 receptor are described. In certain embodiments, the invention provides an isolated polynucleotide comprising a nucleotide sequence selected from the group consisting of: (a) the nucleotide sequence of SEQ ID NO: 1, from nucleotide 803 to the 1999 nucleotide; (b) a nucleotide sequence that varies from the sequence of the nucleotide sequence specified in (a) as a result of a degeneracy of the genetic code; and (c) an allelic variation of the nucleotide sequence specified in (to). Preferably, the nucleotide sequence codes for a protein having a biological activity of the human interleukin-11 receptor. The nucleotide sequence may be operably linked to an expression control sequence. In preferred embodiments, the polynucleotide comprises the nucleotide sequence of SEQ ID NO: 1, from nucleotide 803 to nucleotide 1999 or a fragment thereof; the nucleotide sequence of SEQ ID NO: 1, from nucleotide 803 to nucleotide 1828 or a fragment thereof; the sequence of nucleotides of SEQ ID NO: 1, from nucleotide 1904 to nucleotide 1999 or a fragment thereof; the nucleotide sequence of SEQ ID NO: 1, from nucleotide 734 to nucleotide 1999 or a fragment thereof; the nucleotide sequence of SEQ ID NO: 1, from nucleotide 1067 to nucleotide 1828 or a fragment thereof; or the nucleotide sequence of SEQ ID NO: 1, from nucleotide 1067 to nucleotide 1999 or a fragment thereof. In other embodiments, the polynucleotide comprises a nucleotide sequence capable of hybridizing to the nucleotide sequence of SEQ ID NO: 1 under highly stringent conditions. The invention also provides isolated polynucleotides comprising a nucleotide sequence encoding a peptide or a protein comprising an amino acid sequence selected from the group consisting of: (a) the amino acid sequence of SEQ ID NO: 2; (b) the amino acid sequence of SEQ ID NO: 2; of amino acid 24 to (c) the amino acid sequence of SEQ ID NO: 2; of amino acid 24 to 365; (d) the amino acid sequence of SEQ ID NO: 2; of amino acid 391 to 422; (e) the amino acid sequence of SEQ ID NO: 2; of amino acid 112 to 422; (f) the amino acid sequence of SEQ ID NO: 2; of amino acid 112 to 365; (g) fragments of (a) - (f) having biological activity of the human IL-11 receptor. Host cells are also provided, preferably mammalian cells, transformed with the polynucleotides. In other embodiments, the invention provides a process for producing a human IL-11 R protein. The process comprises: (a) cultivating a culture of the host cell of the present invention in an appropriate culture medium; and (b) purifying the human IL-11 R protein from the culture. The proteins produced according to these methods are also provided. The present invention also provides a protein isolated from human IL-11R, which comprises an amino acid sequence selected from the group consisting of: (a) the amino acid sequence of SEQ ID NO: 2; (b) the amino acid sequence of SEQ ID NO: 2; of amino acid 24 to 422; (c) the amino acid sequence of SEQ ID NO: 2; of amino acid 24 to 365; (d) the amino acid sequence of SEQ ID NO: 2; of amino acid 391 to (e) the amino acid sequence of SEQ ID NO: 2; of amino acid 112 to 422; (f) the amino acid sequence of SEQ ID NO: 2; of amino acid 112 to 365; and (g) fragments of (a) - (f) having biological activity of the receptor of the Human IL-11. Preferably the protein comprises the amino acid sequence of SEQ ID NO: 2; the amino acid sequence 24 to 422 of SEQ ID NO: 2; the amino acid sequence 24 to 365 of SEQ ID NO: 2; or the amino acid sequence 391 to 422 of SEQ ID NO: 2. Pharmaceutical compositions comprising a protein of the present invention and a pharmaceutically acceptable carrier are also provided. The present invention also provides compositions comprising an antibody that specifically reacts with a protein of the present invention. Methods for the identification of an inhibitor of IL-11 binding to the human IL-11 receptor are also provided. These methods comprise: (a) combining a human IL-11 R protein or a fragment thereof with IL-11 or a fragment thereof, said combination forming a first binding mixture; (b) measure the degree of binding between the protein and IL-11 or fragment in the first binding mixture; (c) combining a compound with the protein and the IL-11 or fragment to form a second binding mixture; (d) measuring the degree of binding in the second binding mixture; and (e) comparing the degree of binding in the first binding mixture with the degree of binding in the second binding mixture; wherein the compound is capable of inhibiting the binding of IL-11 to the human IL-11 receptor when a decrease in the degree of binding of the second binding mixture occurs. Optionally, the first and / or second binding mixture can further comprise gp130 or a fragment thereof capable of binding to the protein of claim 11 or to IL-11 or fragment used therein. Inhibitors of IL-11R identified by these methods, and pharmaceutical compositions containing them, are also provided. Methods for inhibiting the binding of IL-11 to the human IL-11 receptor in mammalian subjects are also described, which comprise the administration of a therapeutically effective amount of a composition containing a human IL-11 R protein, a IL-11 R inhibitor or an antibody to a human IL-11 R protein. Methods for the treatment or prevention of bone loss in a mammalian subject using these compositions are also provided.

BRIEF DESCRIPTION OF THE DRAWINGS Figure 1 depicts a schematic representation of the structures of the human IL-11 receptor and gp130. Figure 2 presents information demonstrating the biological activity of a soluble form of recombinant human IL-11 R protein.

Detailed Description of the Preferred Modes The inventors of the present application have identified and provided for the first time a polynucleotide encoding the human IL-11 receptor (human IL-11R), SEQ ID NO: 1 provides the nucleotide sequence of a cDNA encoding human IL-11 R. SEQ ID NO: 2 provides the amino acid sequence of the receptor, including a putative signaling sequence of amino acids 1-23. Mature human IL-11 R is believed to have amino acid sequence 24-422 of SEQ ID NO: 2. The mature receptor has at least three distinct domains: an extracellular domain (comprising approximately amino acids 24-365 of SEQ. ID NO: 2). A transmembrane domain (comprising approximately amino acids 366-390 of SEQ ID NO: 2) and an intracellular domain (comprising approximately amino acids 391-422 of SEQ ID NO: 2). The extracellular domain is further divided into a domain similar to immunoglobulin (comprising approximately amino acids 24-111 of SE ID NO: 2) and a cytokine tipol domain (comprising approximately amino acids 112-365 of SEQ ID NO: 2 ). Soluble forms of the human IL-11R protein can also be produced. Said soluble forms include without limitation proteins comprising amino acids 1-365 and 24-365 of SEQ ID NO: 2. Soluble forms of human IL-11 R are further characterized as being soluble in aqueous solution, preferably at room temperature. Human IL-11 R proteins comprising only the intracellular domain or a part thereof can also be produced. Any forms of human IL-11R of less than total length are encompassed within the present invention and are collectively referred to herein as "human IL-11 R" or "human IL-11R proteins." Human IL-11 R proteins of less than total length can be produced by expressing a corresponding fragment of the polynucleotide encoding the full-length human IL-11 R protein (SEQ ID NO: 1). These corresponding polynucleotide fragments are also part of the present invention. Modified polynucleotides as described above can be made by standard molecular biology techniques, including the construction of appropriate mutants of desired deletion, methods of site-directed mutagenesis or by the reaction of the polymerase chain using appropriate oligonucleotide primers. Based on the similarity to the structure of the IL-6 receptor, it is predicted that IL-11 R proteins containing only the cytokine type I domain of the extracellular region of the full length receptor will be able to bind to IL-11 and to induce receptor signaling. As a result, IL-11R proteins comprising amino acids 112 to 365 of SEQ ID NO: 2, IL-11 R proteins comprising amino acids 112 to 390 of SEQ ID NO: 2 are provided by the invention. , and the IL-11 R proteins comprising amino acids 112 to 422 of SEQ ID NO: 2. Polynucleotides encoding such proteins (such as for example a polynucleotide comprising the nucleotide sequence) are also provided by the invention. of SEQ ID NO: 1, from nucleotide 1067 to nucleotide 1828, a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1, from nucleotide 1067 to nucleotide 1906, and a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1, from nucleotide 1067 to nucleotide 1999, respectively). For the purposes of the present invention, a protein has a "biological activity of the human IL-11 receptor" if it possesses one or more of the following characteristics. (1) the ability to bind IL-11 or a fragment of it (preferably a biologically active fragment thereof); (2) the ability to bind to cytosolic molecules or proteins involved in the signaling pathway invoked by the binding of IL-11 to human IL-11 R; (3) the ability to produce a signal characteristic of the binding of IL-11 to human IL-11 R (where the protein in question contains either a portion capable of binding to IL-11 or where the protein in question would produce such a signal if it were linked to another protein that is capable of binding to IL-11); (4) the ability to bind to gp130 or a fragment thereof (either in the presence or absence of IL-11); (5) the ability to induce tyrosine phosphorylation of gp130; (6) the ability to induce tyrosine phosphorylation of JAK kinases; or (7) the ability to induce tyrosine phosphorylation of the STAT family of DNA binding proteins (Zhong et al. (1994) Science 264, 95-98) Preferably, the biological activity possessed by the protein is the ability to binding to IL-11 or a fragment thereof, more preferably with a Ko of about 0.1 to about 100 nM, more preferably with a KD of about 1 to about 10 nM.

Human IL-11R or active fragments thereof (human IL-11R proteins) may be fused to carrier molecules such as immunoglobulins. For example, soluble forms of human IL-11R can be fused through "linker" sequences to the Fe portion of an immunoglobulin. Other fusion proteins can also be used such as those with GST, Lex-A or MBP. The invention also encompasses allelic variations of the nucleotide sequence as set forth in SEQ ID NO: 1, that is, alternative forms, occurring naturally, of the isolated polynucleotide of SEQ ID NO: 1, which also encode proteins of the Human IL-11 R, preferably those proteins that have a biological activity of human IL-11R. Also included in the invention are isolated polynucleotides that hybridize to the nucleotide sequence set forth in SEQ ID NO: 1 under highly stringent conditions (eg, O.lxSSC at 65 ° C). Also encompassed by the present invention are isolated polynucleotides which encode human IL-11 R proteins but which differ from the nucleotide sequence set forth in SEQ ID NO: 1 by virtue of the degeneracy of the genetic code. Also included in the invention are variations in the nucleotide sequence as set forth in SEQ ID NO: 1, which are caused by point mutations or by induced modifications. The isolated polynucleotides of the present invention may be operably linked to an expression control sequence such as the pMT2 or pED expression vectors described in Kaufman et al., Nucleic Acids Res. 19. 4485-4490 (1991), for the purpose of of recombinantly producing the human IL-1 R protein. Many other expression control sequences are known in the art. General methods for the expression of recombinant proteins are also known and are exemplified in R. Kaufman. Methods in Enzymology 185, 537-56 (1990). As defined herein "operably linked" means enzymatically or chemically linked to form a covalent bond between the isolated polynucleotide of the invention and the expression control sequence, such that the human IL-11 R protein is expressed by a cell host that has been transformed (transfected) with the linked polynucleotide / expression control sequence. A number of cell types can act as appropriate host cells for the expression of the human IL-11R protein. Any type of cell capable of expressing functional human IL-11 R protein can be used. Suitable mammalian host cells include, for example, monkey COS cells, Chinese hamster ovary (CHO) cells, human kidney 293 cells, A431 human epidermal cells, human Colo205 cells, 3T3 cells, CV-1 cells, other lines transformed primate cells, normal diploid cells, cell strains derived from in vitro cultures of primary tissue, primary explants, HeLa cells, mouse L cells, BHK cells, HL-60, U937, Hak, Rat2, BaF3, 32D, FDCP -1, PC12, M1x or C2C12. The human IL-11 R protein can also be produced by operably linking the isolated polynucleotide of the invention to appropriate control sequences in one or more insect expression vectors, and employing an insect expression system. Materials and methods for baculovirus / insect cell expression systems are commercially available in the form of games in, for example, Invitrogen, San Diego California, E.U.A. (the MaxBacF Kit), and such methods are well known in the art, as described in Summers and Smith, Texas Aaricultural Experiment Station Bulletin No. 1555 (1987 incorporated herein by reference.) Soluble forms of the human IL-11 R protein they can also be produced in insect cells using appropriate isolated polynucleotides as described above Alternatively, the human IL-11 R protein can also be produced in minor eukaryotes such as yeast or in prokaryotes such as bacteria Suitable strains of yeast include Saccharomyces cerevisiae, Schizosaccharomyces pombe, strains of Kluyveromyces, Candida, or any strain of yeast capable of expressing heterologous proteins. Suitable bacterial strains include Escherichia coli, Bacillus subtilis, Salmonella typhim? Rium, or any bacterial strain capable of expressing heterologous proteins. Expression in bacteria can result in the formation of inclusion bodies that incorporate the recombinate protein. Thus, refolding of the recombinant protein may be required in order to produce active or more active material. Various methods are well known in the art for correctly obtaining folded heterologous proteins from bacterial inclusion bodies. These methods generally involve the solubilization of the protein from the inclusion bodies, subsequently denaturing the protein completely using a chaotropic agent. When cysteine residues are present in the primary amino acid sequence of the protein, it is often necessary to perform the retraction in an environment that allows the correct formation of disulfide bonds (a redox system). General methods of refolding are described in Kohno, Meth. Enzvm., 185: 187-195 (1990), European Patent EP 0433225 and the co-pending application of US Pat. Nos. 08 / 163,877 describe suitable methods. The human IL-11 R protein of the invention can also be expressed as a product of transgenic animals, v. gr., as a component of the milk of transgenic cows, goats, pigs or sheep that are characterized by germ or somatic cells containing a polynucleotide sequence encoding the human IL-11 R protein. The human IL-11 R protein of the invention can be prepared by growing a culture of transformed host cells under culture conditions necessary to express the desired protein. The resulting expressed protein can then be purified from the culture medium or cell extracts. Soluble forms of the human IL-11 R protein of the invention can be purified from conditioned media. Membrane binding forms of the human IL-11R protein of the invention can be purified by preparing a total membrane fraction from the expression cell and extracting the membranes with non-ionic surfactant such as Triton X-100.

The human IL-11 R protein can be purified using methods known to those skilled in the art. For example, the human IL-11R protein of the invention can be concentrated using a commercially available protein concentration filter, for example, a Millipore Pellicon or Amicon ultrafiltration unit. Next to the concentration step, the concentrate can be applied to a purification matrix such as a gel filtration medium. Alternatively, an ion exchange resin, for example, a matrix or a substrate having diethylaminoethyl (DEAE) or polyethylene imine (PEI) groups may be employed. The matrices can be acrylamide, agarose, dextran, cellulose or other types commonly used in protein purification. Alternatively, a cation exchange step may be employed. Suitable cation exchangers include various insoluble matrices comprising sulfopropyl or carboxymethyl groups. Sulfopropyl groups are preferred (e.g., S-Sepharose® columns). Purification of the human IL-11 R protein from the culture supernatant can also include one or more column steps on affinity resins such as concavalin A-agarose, heparin-toyopearl®, or Cibacrom blue 3GA SepharoseF, or by chromatography of hydrophobic interaction using such resins as phenyl ether, butyl ether or propyl ether; or by immunoaffinity chromatography. Finally, one or more steps of high resolution reverse phase liquid chromatography (RP-HPLC) employing hydrophobic RP-HPLC media, for example, silica gel having suspended methyl or other aliphatic groups, can be used to further purify the Human IL-11 R protein. Affinity columns including IL-11 or fragments thereof or including antibodies to the IL-11R protein can also be used in the purification according to known methods. Some or all of the preceding purification steps, in various combinations or with other methods, may also be employed to provide an isolated and substantially purified recombinant protein. Preferably, the protein isolated from human IL-11 R is purified so that it is substantially free of other mammalian proteins. The human IL-11 R proteins of the invention can also be used to classify agents that are capable of binding to human IL-11 R or interfering with the binding of IL-11 to human IL-11 R (whether extracellular domains or intracellular) and thus can act as inhibitors of normal binding and action of cytokine (inhibitors of IL-11R). Binding assays using a desired binding protein, immobilized or not, are well known in the art and can be used for this purpose using the human IL-11R protein of the invention. Classification assays based on purified cells or proteins (cell-free) can be used to identify such agents. For example, the human IL-11 R protein can be immobilized in purified form on a carrier and the protein binding of purified human IL-11 R can be measured in the presence and in the absence of potential inhibitory agents. A suitable binding assay can alternatively employ a soluble form of the human IL-11 R of the invention. In said classification assay, a first binding mixture is formed by combining IL-11 or a fragment thereof and human IL-11 R protein, and the degree of binding in the first binding mixture (B0) is measured. A second binding mixture is also formed by combining IL-11 or a fragment thereof, human IL-11 R protein and the compound or agent to be classified, and the degree of binding in the second binding mixture (B) is measured. ). The degrees of union in the first and second binding mixtures are compared, for example, by calculating the B / B0 ratio. A compound or agent is considered to be capable of inhibiting binding if a decrease in binding is observed in the second binding mixture, compared to the first binding mixture. Optionally, gp130 can be added to one or both binding mixtures. The formulation and optimization of the binding mixtures is within the level of knowledge of the art, such binding mixtures may also contain regulators and salts necessary to enhance or optimize the binding, and additional control tests may be included in the classification test of the invention. Compounds that were found to reduce the binding activity of the human IL-11 R protein to IL-11 or its fragments to any degree, preferably at least about 10%, more preferably above about 50% or more they can therefore be identified and subsequently classified secondarily in other binding assays and in vivo assays. By these means, compounds having inhibitory activity for the binding of IL-11 R, which may be suitable as therapeutic agents, can be identified. Human IL-11 R proteins, and polynucleotides that encode them, can also be used as diagnostic agents to detect the expression or presence of IL-11 R, IL-11 or cells expressing IL-11R or IL- eleven. The proteins or polynucleotides can be employed for that purpose in standard procedures for diagnostic assays using these types of materials. Appropriate methods are well known to those skilled in the art. Human IL-11 R acts as a mediator of the known biological activities of IL-11. As a result, isolated human IL-11R protein and IL-11 R inhibitors may be useful in the treatment or modulation of various medical conditions. in which IL-11 is involved or that are effected by the activity (or lack of it) of IL-11 (collectively "conditions related to IL-11") Conditions related to IL-11 include, without limitation, ipmunological deficiencies, specifically deficiencies in hematopoietic progenitor cells, or related disorders, cancer and other diseases.

Said disease states can result from disease, radiation exposure or medications, and include, for example, leukopenia, bacterial and viral infections, anemia, B cell deficiencies or T cells such as those deficiencies in immune cells or in hematopoietic cells that follow to bone marrow transplants. It is also believed that IL-11 and IL-11 R may play a role in the regulation of bone maturation and repair (Girasole et al., (1994) J. Clin. Invest. 93, 1516-1524; Passeri et al. al. (1992) J. Bone Miner, Res., 7 (S1), S110 Abst, Passeri et al., (1993) J. Bone Miner, Res., 8 (S1), S162 Abst.). As a result, the human IL-11 R protein and the IL-11 R inhibitors may be useful in the treatment of bone loss (including that associated with osteoporosis, post-menopausal osteoporosis, senile osteoporosis, idiopathic osteoporosis, Pagets, multiple myeloma and hypogonadic conditions). The human IL-11 R protein and the IL-11 R inhibitors, purified from cells or recombinantly produced, can be used as a pharmaceutical composition when combined with a pharmaceutically acceptable carrier. Said composition may contain, in addition to human IL-11R or ligand and carrier, diluents, fillers, salts, regulators, stabilizers, solubilizers, and other materials that are well known in the art. The term "pharmaceutically acceptable" means a non-toxic material that does not interfere with the effectiveness of the biological activity of the active ingredient (s). The characteristics of the carrier will depend on the administration route. The pharmaceutical composition of the invention may also contain cytokines, lymphokines, or other hematopoietic factors such as M-CSF, GM-CSF, IL-1, IL-2, IL-3, IL-4, IL-5, IL-6. , IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-15, G-CSF, progenitor cell factor and erythropoietin. The pharmaceutical composition may contain thrombolytic or anti-thrombotic factors such as plasminogen activator and Factor VIII. The pharmaceutical composition may also contain other anti-inflammatory agents. Such additional factors and / or agents can be included in the pharmaceutical composition to produce a synergistic effect with the isolated human IL-11 R protein or the IL-11 R inhibitor, or to minimize side effects caused by IL-11. Isolated human R or IL-11 inhibitor R. Conversely, isolated human IL-11 R or IL-11R inhibitor can be included in the particular cytocipase formulation, lymphokine, other hematopoietic factor, thrombolytic factor or anti-thrombotic, or anti-inflammatory agent to minimize side effects of the cytokine, lymphokine, another hematopoietic factor, thrombolytic or anti-thrombotic factor, or anti-inflammatory agent. The pharmaceutical composition of the invention may be in the form of a liposome wherein the human IL-11 R protein, asylated, or the inhibitor of IL-11 R is combined, in addition to other pharmaceutically acceptable carriers, with amphipathic agents such as lipids that exist in aggregate form such as miscelas, insoluble monolayers, liquid crystals, or laminar layers that are in aqueous solution. Suitable lipids for liposomal formulation include, without limitation, monoglycerides, diglycerides, sulfatides, lysolecithin, phospholipids, saponins, bile acids, and the like. The preparation of such liposomal formulations is within the skill of the art, as described, for example, in U.S. Pat. No. 4,235,871; Patent of the U.S.A. No. 4,501,728; Patent of the U.S.A. No. 4,837,028 and U.S. Pat. No. 4,737, 323; all of which is incorporated herein by reference. As used herein, the term "therapeutically effective amount" means the total amount of each active component of the pharmaceutical composition or method that is sufficient to show a significant benefit to the patient, for example, improvement of the symptoms, cure or increase in the speed of cure of said conditions. When an active ingredient is applied to an individual, administered alone, the term refers to that ingredient alone. When applied to a combination, the term refers to combined amounts of the active ingredients that result in the therapeutic effect, whether administered in combination, serially or simultaneously. In the practice of the method of treatment or use of the present invention, a therapeutically effective amount of the protein isolated from the human IL-11R or the inhibitor of the IL-11 R is administered to a mammal. The human IL-11R protein, isolated, or the inhibitor of IL-11 R can be administered according to the method of the invention, either alone or in combination with other therapies such as treatments employing cytokines, lymphokines or other factors hematopoietic When co-administered with one or more cytokines, lymphokines or other hematopoietic factors, the human IL-11 R protein or the IL-11R inhibitor can be administered either simultaneously with cytokine (s), lymphokine (s), other (s) hematopoietic factor (s), thrombolytic or anti-thrombotic factors, or sequentially. If administered sequentially, the treating physician will decide on the appropriate sequence of administration of the human IL-11R protein or the IL-11 R inhibitor in combination with cytokine (s), lymphokine (s), other factor (s) (is) hematopoietic (s), thrombolytic or anti-thrombotic factors. Administration of the human IL-11 R protein or the IL-11 R inhibitor used in the pharmaceutical composition or to practice the method of the present invention can be carried out in a variety of conventional ways, such as oral ingestion, inhalation, skin injection, subcutaneous or intravenous: Intravenous administration to the patient is the most preferred. When a therapeutically effective amount of the human IL-11 R protein or the IL-11 R inhibitor is administered orally, the human IL-11R or IL-11 inhibitor protein Will subside in the form of a tablet, capsule, powder, solution or elixir. When administered in tablet form, the pharmaceutical composition of the invention may additionally contain a solid carrier such as gelatin or an adjuvant. The tablet, capsule and powder contains from about 5 to 95% of the human IL-11 R protein or inhibitor of IL-11 R and preferably from about 25 to 90% of human IL-11 R protein or IL-11 inhibitor. 11 R, When administered in liquid form, a liquid carrier such as water, petroleum, animal or vegetable oils such as peanut oil, mineral oil, soybean oil, sesame oil, or synthetic oils can be added. The liquid form of the pharmaceutical composition may further contain physiological saline, dextrose or other saccharide solution, or glycols such as ethylene glycol, propylene glycol or polyethylene glycol. When administered in liquid form, the pharmaceutical composition contains from about 0.5 to 90% by weight of the human IL-11R protein or the IL-11R inhibitor, and preferably from about 1 to 50% of IL protein. -11R human or inhibitor of IL-11. When a therapeutically effective amount of the human IL-11R protein or IL-11 R inhibitor is administered by intravenous, cutaneous or subcutaneous injection, the human IL-11 R protein or IL-11 R inhibitor will be in the form of a parenterally acceptable and pyrogen-free aqueous solution. It is within the skill in the art to prepare such parenterally acceptable protein solutions, with due regard to pH, isotonicity, stability and the like. A pharmaceutically preferred composition for intravenous, cutaneous or subcutaneous injection must contain, in addition to the human IL-11 R protein or IL-11R inhibitor, an isotonic vehicle such as Sodium Chloride Injection, Ringer's Injection, Dextrose Injection, Injection of Dextrose and Sodium Chloride, Ringer's Injection with Lactate, or other vehicle as is known in the art. The pharmaceutical composition of the present invention may also contain stabilizers, preservatives, regulators, antioxidants, or other additives known to those skilled in the art. The amount of the human IL-11 R protein or inhibitor of IL-11 R in the pharmaceutical composition of the present invention will depend on the nature and severity of the condition being treated, and on the nature of previous treatments that the patient I had. Finally, the attending physician will decide the amount of the human IL-11 R protein or IL-11 R inhibitor that will treat each individual patient. Initially, the attending physician will administer low doses of the human IL-11 R protein or IL-11R inhibitor and observe the patient's response. Higher doses of the human IL-11R protein or IL-11 R inhibitor can be administered until the optimal therapeutic effect is obtained for the patient, and at that point the dose will generally not be increased further. It is contemplated that the various pharmaceutical compositions used in the practice of the method of the present invention should contain about 0.1 μg to about 100 mg of the human IL-11 R protein or IL-11 R inhibitor per kilogram of body weight. The duration of intravenous therapy using the pharmaceutical composition of the present invention will vary depending on the severity of the disease being treated and the potential idiosyncratic condition and response of each individual patient. It is contemplated that the duration of each application of the human IL-11R protein or IL-11R inhibitor will be in the range of 12 to 24 hours of continuous intravenous administration. Finally the attending physician will decide about the appropriate duration of the intravenous therapy using the pharmaceutical composition of the present invention. Human IL-11 R proteins of the invention can also be used to immunize animals to obtain polyclonal and monoclonal antibodies that specifically react with the human IL-11 R protein and which can inhibit the binding of IL-11 or fragments of This one to the receiver. Such antibodies can be obtained using whole human IL-11 R as an immunogen, or using fragments of human IL-11R, such as soluble mature IL-11R. Minor fragments of human IL-11 R can also be used to immunize animals. The peptide immunogens may additionally contain a cysteine residue at the carboxyl terminus, and be conjugated to a hapten such as keyhole limpet hemocyanin (KLH). Additional peptide immunogens can be generated by replacing tyrosine residues with sulphated tyrosine residues. Methods for synthesizing such peptides are known in the art, for example, as in R.P. Merrifield, J. Amer. Chem. Soc. 85, 2149-2154 (1963); J.L. Krstenansky, et al, FEBS Lett. 2jM 10 (1987). Neutralization or non-neutralizing antibodies (preferably monoclonal antibodies) that bind to human IL-11R protein may also be useful therapeutics for certain tumors and also in the treatment of the conditions described above. These monoclonal neutralizing antibodies may be able to block the binding of IL-11 to human IL-11 R.

Example 1 Isolation of human IL-11 R cDNA Generation of DNA Probes: DNA probes derived from the murine Etl-2 sequence (SEQ ID No: 3) were obtained by PCR from murine placental cDNA . The probe with amino terminus corresponds to base pairs 418-570 and the probe with carboxy termination at base pairs 847-1038 of the murine Etl-2 sequence. The DNA probes were gel purified and radiolabelled using a32P-dATP and a32P-dCTP.

Classification of the cDNA Library cDNA was generated from activated human PBMC using the Superscript Choice System and cloned into the Ecorl site of ZAP II (Stratagene). The resulting phage was used to infect the BB4 strain of E. coli. One million phages were plated on 150 mm NZCYM plates at a density of 15,000 pfu / plate. The plates were transferred to duplicates of Duralose nitrocellulose filters (Stratagene). Next to alkaline denaturation and heat fixation the filters were prehybridized in 5X SSC, 5X Denhardts, 01% SDS, and 50 μg / ml yeast tRNA for 2 hours at 65 ° C. One set of filters was hybridized with the amino-terminated probe and the other set with the carboxy-terminated probe (5 x 105 cpm / ml) for 48 hours at 55 ° C in the pre-hybridization buffer. The filters were washed with 4X SSC, 0.1% SDS once at 25 ° C and twice at 55 ° C. The plates that hybridized to both probes were identified by autoradiography. Of the million plates classified, two plates hybridized to both probes.

These plates were harvested and the phage eluted within SM media containing chloroform. The resulting phage was used to reinfect E. coli strain BB4 and plated on NZCYM plates at a density of 100-300 pfu / plate for a second classification. After secondary sorting, plasmid DNA was isolated from ZAPII plates by cut using helper phage (Stratagenß). The DNA sequence of the inserts was determined in an Applied Biosystems DNA sequencer. The clone phlL11 R14-2 containing the polynucleotide having the sequence of SEQ ID No .: 1 was deposited with the ATCC, accession number 69731, on December 22, 1994.

Example 2 Soluble Protein Expression of Human IL-11R v Activity Assay A soluble form of the human IL-11 R protein was expressed in mammalian cells. The expressed recombinant protein was able to effect the transduction of a signal in BAF130-9 cells. A portion of the full-length human IL-11 R sequence (nucleotides 734-1828 of SEQ ID No .: 1 encoding amino acids 1-365 of SEQ ID No .: 2) corresponding to a soluble form was cloned into the mammalian pED expression vector and used to transfect COSM6 cells. 40 hours after the transfection medium conditioned culture was removed, concentrated 5 times and used in proliferation assays with the murine BAF130-9 cell line (Hibi, M, et al. (1990) Cell 63, 1149-57 ), a derivative of the BAFB03 cell line that expresses the signal transducer of human gp130. BAF130-9 cells do not proliferate in response to IL-11 or IL-6 alone, but proliferated in response to the combination of IL-6 and soluble IL-6R (Hibi et al., Supra), BAF130- cells 9 (1 x 10 4 cells in 0.1 ml) were cultured in RPMI 1640/10% FCS culture medium with increasing concentrations of recombinant human IL-11 in the absence or presence of 10 μl of conditioned medium from imitated transfected cells or cells transfected with the sequence of soluble human IL-11 R. After forty hours the cells were labeled by pulse with 3H-thymidine (0.5 μCi / well) for eight hours and the incorporated nucleotide was determined. As shown in Figure 2, BAF-130-9 cells do not proliferate in response to IL-11 or soluble IL-11 R alone, but do proliferate in the presence of both IL-11 and IL- 11R. Other proteins of human IL-11 R can be tested in this model to determine whether they exhibit a "biological activity" of human IL-11R as defined herein.

Example 3 Other Systems for the Determination of Biological Activity of Human IL-11 R Protein Other systems can be used to determine whether a specific human IL-11R protein has a "biological activity" of human IL-11R as defined herein. The following are examples of such systems.

Assays for IL-11 Binding The ability of a human IL-11R protein to bind IL-11 or a fragment thereof can be determined by any appropriate assay that can detect such binding. Some suitable examples are: The binding of IL-11 to the extracellular region of the human IL-11R protein will specifically cause a rapid induction of phosphotyrosine on the receptor protein. The assays for ligand binding activity, measured by induction of phosphorylation, are described below. Alternatively, a human IL-11 R protein (such as, for example, a soluble form of the extracellular domain) is produced and used to detect the binding of IL-11. For example, a DNA construct is prepared wherein the extracellular domain (truncated before, preferably immediately before, of the predicted transmembrane domain) is ligated in frame to a cDNA encoding the hinge domains CH2 and CH3 of a human immunoglobulin (ig) ? This construct is generated in an expression vector appropriate for COS cells, such as pED? C or pMT2. The plasmid is transiently transfected into COS cells. The protein of the secreted IL-11 R-lg fusion is harvested in the conditioned culture medium and purified by protein A chromatography. The purified IL-11 R-lg fusion protein is used to demonstrate the binding of IL-11. 11 in a number of applications. IL-11 can be coated on the surface of an enzyme-linked immunosorbent assay plate (ELISA) and subsequently additional binding sites blocked with casein or bovine serum albumin using standard ELISA regulators. The IL-11 R-lg fusion protein is then bound to the solid phase IL-11 and the binding is detected with a secondary, anti-human, goat Ig conjugated to horseradish peroxidase. The specifically bound enzyme activity can be measured with a colorimetric substrate, such as tetramethyl benzidine and absorbance readings.

IL-11 can also be expressed on the cell surface, for example by providing a glycosylphosphatidyl inositol (GPI) bond or transmembrane domain. Cells expressing membrane-bound IL-11 can be identified using the IL-11 R-Ig fusion protein. The fusion of soluble IL-11 R-lg is ligated to the surface of these cells and detected with goat antihuman Ig conjugated to a fluorochrome, such as flow cytometry and fluorescent isothiocyanate.

Interaction trap A yeast genetic selection method, the "interaction trap" [Gyuris et al., Cell 75: 791-803, 1993], can be used to determine whether a human IL-11R protein has a biological activity of human IL-11R as defined here. In this system, the expression of the reporper genes of both LexAop-Leu2 and LexAop-LacZ depends on the interaction between the bait protein, for example in this case a species that interacts with human IL-11R and the prey, for example in this case, the human IL-11R protein. Thus, one can measure the strength of the interaction by the level of expression of LacZ or Leu2. The simplest method is to measure the activity of the protein encoded by LacZ, β-galactosidase. This activity can be judged by the degree of blue color on the filter or medium containing X-Gal. For quantitative measurement of β-galactosidase activity, standard assays can be found in "Methods in Yeast Genetics" Cold Spring Harbor, New York, 1990 (by Rose, MD, Winston, F. and Hieter, P.), In such methods, if one wishes to determine whether the human IL-11R protein interacts with a partar species (such as, for example, a cystolic protein that binds to the intracellular domain of human IL-11 R in vivo), that species can be used as the "bait" in the interaction trap with the human IL-11R protein to be tested serving as the "prey", or vice versa.

CAT Induction System Transcription of critical phase plasma protein genes, such as the rat β-fibrinogen gene, is activated by IL-11 in a variety of cell lines. In an exemplary system, COSM6 cells are transfected with plasmids encoding the human IL-11 R protein (such as full-length human IL-11 R or a soluble form thereof), the signal transducer of human gp130 and a reporter gene containing the promoter region of 350 base pairs of the rat b-fibrinogen gene fused to a reporter gene, CAT (Baumann et al (1991) J. Biol. Chem 266, 20424-27), The cells are stimulated with increasing concentrations of recombinant human IL-11 and transcription of the reporter gene is monitored by assay to determine the presence of CAT activity.

Phosphorylation of ap130 Activity can also be determined by examining the ability of the IL-11 to induce tyrosine phosphorylation of gp130 in cells transfected with a sequence encoding the human IL-11 R protein (such as full-length human IL-11R or a soluble form thereof) (Luttcken et al. (1994) Science 263, 89-92).

Phosphorylation of STATs Activity can also be determined by examining the ability of IL-11 to induce tyrosine phosphorylation of STATs (ransducers and activators of transcriptional signal, a family of DNA binding proteins) in cells transfected with a sequence encoding the human IL-11R protein (such as full length human IL-11 R or a soluble form thereof) (Zhong et al (1994) Science 264, 95-98) Phosphorylation of JAK kinases Activity can also be determined by examining the ability of IL-11 to induce tyrosine phosphorylation of JAK kinase in cells transfected with a sequence encoding the human IL-11R protein (such as IL-11). Full-length human R or a soluble form thereof) (Yin et al (1993) J Immunol 151 2555-61) All of the patents and literature references cited here are incorporated by reference as if they had been fully exposed, LIST OF SEQUENCES (1) GENERAL INFORMATION (i) APPLICANT: Tobin, James (ii) TITLE OF THE INVENTION: RECEPTOR OF INTERLEUCINE-11 HUMAN (iii) SEQUENCE NUMBER: 4 (iv) ADDRESS FOR CORRESPONDENCE: (A) RECIPIENT: Genetics Institute, Inc. (B) STREET: 87 Cambridge Park Drive (C) CITY: Cambridge (D) STATE: MA (E) COUNTRY: USA (F) POSTAL CODE: 02140 (V) COMPUTER LEADABLE FORM: (A) MEDIA TYPE: Floppy disk (B) COMPUTER: IBM compatible PC (C) OPERATING SYSTEM: PC-DOS / MS-DOS (D) SOFTWARE: Patentln Reeléase # 1.0, Version # 1.25 (vi) DATA OF THE CURRENT APPLICATION: (A) APPLICATION NUMBER: (B) DATE OF PRESENTATION: (C) CLASSIFICATION: (viii) EMPOWERED / AGENT INFORMATION: (A) NAME: Bro n, Scott A. (B) REGISTRATION NUMBER: 32,724 (C) REFERENCE NUMBER / FILE: GI5252 (ix) TELECOMMUNICATIONS INFORMATION: (A) PHONE: (617) 498-8224 (B) TELEFAX: ( 617) 876-5851 (2) INFORMATION FOR SEQ ID NO: 1: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 2456 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: cDNA (iii) HYPOTHETICAL: NO (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 734..1999 (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: l: TCGCCCACCC CCAGCCTCTG GCAGCAGCCA GGGCATCTGG ATCTGCTTAA CTACACAGCC 60 CCAGCCTGCA CCCTAGCCCC ATCCAGCTTC ACAAACTGGA GACCAACGAA GTGTCAAGAG 120 CCAGGCCCAG CTGAGTGGCC CAAGTAGCCA GACCAAGGAG CCAGGTTCAG GCGAGAAGCC 180 TGGCAGCCAG GGCAGGGGTG GGCCTCAGGG TGGGAGTGCA GGATGGGCTC AGATCCATGA 240 TGACACCCTT CCCCCAGGGT GATAA3GTCT GCCTAGGTTA ATCAGAGGCA GTGATAAGCC 300 CTGGACCAGG TGGGGGTAAA TACCAGAATT CCCAACAGCT GGACTGGAGG GGTTAATGGG 360 AGTGGCTGAG CTGGTGCCAG TGCTTGGTGC CAGGGGTGGG CGCCAAGGGC AGTGGAGGGG 420 GAGTTGCTGG CACAGTCTGT TGCCTCCGGC TTTTGTTCTG GGCCCTAAGC CCAGGACTGA 480 GATGGAGGGT GTGAGGGGGT GTGTGTGTCC GTGTGTGTGT GTGTGTGTGT GTGCGCGCGC 540 ACGCACATGC AAAGCACTGG GTATACAGTG GGAAAGGGGA CCTCAGGTCA GTTCCCGCAG 600 TGATTTCTAA CAGCCTTACC CCACTTGGTG CATCAATTTT TCTCCTAGGA AGCCTCAGTT 660 TTGGAGAGGA AGAGCCAGGC TTTAGCCTCC CATCTCAGGG GTCGGGGATT TTTGACTCTA 720 CCTCTCCCCA CAG ATG AGC AGC AGC TGC TCA GGG CTG AGC GTC CTG 769 Met Ser Ser Cys Ser Gly Leu Ser Arg Val Leu 1 5 10 GTG GCC GTG GCT ACÁ GCC CTG GTG TCT GCC TCC TCC CCC TGC CCC CAG 817 Val Ala Val Ala Thr Ala Leu Val Ser Wing Ser Pro Cys Pro Gln 15 20 25 GCC TGG GGC CCC CCA GGG GTC CAG TAT GGG CAG CCA GGC AGG TCC GTG 865 Wing Trp Gly Pro Pro Gly Val Gln Tyr Gly Gln Pro Gly Arg Ser Val 30 35 40 AAG CTG TGT TGT CCT GGA GTG ACT GCC GGG GAC CCA GTG TCC TGG TTT 913 Lys Leu Cys Cys Pro Gly Val Thr Wing Gly Asp Pro Val Ser Trp Phe 45 50 55 60 CGG GAT GGG GAG CCA AAG CTG CTC CAG GGA CCT GAC TCT GGG CTA GGG 961 Arg Asp Gly Glu Pro Lys Leu Leu Gln Gly Pro Asp Ser Gly Leu Gly 65 70 75 CAT GAA CTG GTC CTG GCC CAG GAC GAC AGC ACT GAT GAG GGC ACC TAC 1009 His Glu Leu Val Leu Ala Gln Ala Asp Ser Thr Asp Glu Gly Thr Tyr 80 85 90 ATC TGC CAG ACC CTG GAT GGT GCA CTT GGG GGC ACA GTG ACC CTG CAG 1057 He Cys Gln Thr Leu Asp Gly Ala Leu Gly Gly Thr Val Thr Leu Gln 95 100 105 CTG GGC T AC CCT CCA 'GCC CGC CCT GTT GTC TCC TGC CAG GCA GCC GAC 1105 Leu Gly Tyr Pro Pro Wing Arg Pro Val Val Ser Cys Gln Wing Wing Asp 110 115 120 TAT GAG AAC TTC TCT TGC ACT TGG AGT CCC AGC CAG ATC AGC GGT TTA 1153 Tyr Glu Asn Phe Ser Cys Thr Trp Ser Pro Ser Gln He Ser Gly Leu 125 130 135 140 CCC ACC CGC TAC CTC ACC TCC TAC AGG AAG AA GTC CTA GGA GCT 1201 Pro Thr Arg Tyr Leu Thr Ser Tyr Arg Lys Lys Thr Val Leu Gly Wing 145 150 155 GAT AGC CAG AGG AGT AGC CCA TCC ACA GGG CCC TGG CCA TGC CCA CAG 1249 Asp Ser Gln Arg Arg Ser Pro Ser Thr Gly Pro Trp Pro Cys Pro Gln 160 165 170 GAT CCC CTA GGG GCT GCC CGC TGT GTT GTC CAC GGG GCT TAG TGG 1297 Asp Pro Leu Gly Wing Wing Arg Cys Val Val His Gly Wing Glu Phe Trp 175 180 185 AGC CAG TAC CGG ATT AAT GTG ACT GAG GTG AAC CCA CTG GGT GCC AGC 1345 Ser Gln Tyr Arg He Asn Val Thr Glu Val Asn Pro Leu Gly Wing Ser 190 195 200 ACÁ CGC CTG CTG GAT GTG AGC TTG CAG AGC ATC TTG CGC CCT GAC CCA 1393 Thr Arg Leu Leu Asp Val Ser Leu Gln Ser He Leu Arg P ro Asp Pro 205 210 215 220 CCC CAG GGC CTG CGG GTA GAG TCA GTA CCA GGT TAC CCC CGA CGC CTG 1441 Pro Gln Gly Leu Arg Val Glu Ser Val Pro Gly Tyr Pro Arg Arg Leu 225 230 235 CGA GCC AGC TGG ACÁ TAC CCT GCC TCC TGG CCG TGC CAG CCC CTC TTC 1489 Arg Wing Ser Trp Thr Tyr Pro Wing Ser Trp Pro Cys Gln Pro His Phe 240 245 250 CTG CTC AAG TTC CGT TTG CAG TAC CGT CCG GCG CAG CAT CCA GCC TGG 1537 Leu Leu Lys Phe Arg Leu Gln Tyr Arg Pro Wing Gln His Pro Wing Trp 255 260 265 TCC ACG GTG GAG CCA GCT GGA CTG GAG GAG GTG ATC ACA GAT GCT GTG 1585 Ser Thr Val Glu Pro Wing Gly Leu Glu Glu Val He Thr Asp Wing Val 270 275 280 GCT GGG CTG CCC CAT GCT GTA CGA GTC AGT GCC CGG GAC TTT CTA GAT 1633 Wing Gly Leu Pro His Wing Val Arg Val Ser Wing Arg Asp Phe Leu Asp 285 290 295 300 GCT GGC ACC TGG AGC ACC TGG AGC CCG GAG GCC TGG GGA ACT CCG AGC 1681 Wing Gly Thr Trp Ser Thr Trp Ser Pro Glu Wing Trp Gly Thr Pro Ser 305 310 315 ACT GGG ACC ATA CCA AAG GAG ATA CCA GCA TGG GGC CAG CTA CAC ACG 1729 Thr Gly Thr He Pr or Lys Glu He Pro Wing Trp Gly Gln Gln Leu His Thr 320 325 330 CAG CCA GAG GTG GAG CCT CAG GTG GAC AGC CCT GCT CCT CCA AGG CCC 1777 Gln Pro Glu Val Glu Pro Gln Val Asp Ser Pro Pro Wing Pro Pro Arg Pro 335 340 345 TCC CTC CAA CCA CAC CCT CGG CTA CTT GAT CAC AGG GAC TCT GTG GAG 1825 Ser Leu Gln Pro His Pro Arg Leu Leu Asp His Arg Asp Ser Val Glu 350 355 360 CAG GTA GCT GTG CTG'GCG TCT TTG GGA ATC CTT TCT TTC CTG GGA CTG 1873 Gln Val Wing Val Leu Wing Ser Leu Gly He Leu Ser Phe Leu Gly Leu 365 370 375 380 GTG GCT GGG GCC CTG GCA CTG GGG CTC TGG CTG AGG CTG AGA CGG GGT 1921 Val Ala Gly Ala Leu Ala Leu 31y Leu Trp Leu Arg Leu Arg Arg Gly 385 390 395 GGG AAG GAT GGA TCC CCA AAG CCT GGG TTC TTG GCC TCA GTG ATT CCA 1969 Gly Lys Asp Gly Ser Pro Lys Pro Gly Phe Leu Wing Ser Val He Pro 400 405 410 GTG GAC AGG CGT CCA GGA GCT CCA AAC CTG TAGAGGACCC AGGAGGGCTT 2019 Val Asp Arg Arg Pro Gly Wing Pro Asn Leu 415 420 CGGCAGATTC CACCTATAAT TCTGTCTTGC TGGTGTGGAT GGATGGACAG ATAGAAACCA 2079 GGCAGGACAG TAGATCCCTA TGGTTGGATC TCAGCTGGAA GTTCTGTTTG GAGCCCATTT 2139 CTGTGAGACC CTGTATTTCA AATTTGCAGC TGAAAGGTGC TTGTACCTCT GATTTCACCC 2199 CAGAGTTGGA GTTCTGCTCA AGGAACGTGT GTAATGTGTA CATCTGTGTC CATGTGTGAC 2259 CATGTGTCTG TGAGGCAGGG AACATGTATT CTCTGCATGC ATGTATGTAG GTGCCTGGGG 2319 AGTGTGTGTG GGTCCTTGGC TCTTGGCCTT TCCCCTTGCA GGGGTTGTGC AGGTGTGAAT 2379 AAAGAGAATA AGGAAGTTCT TGGAGATTAT ACTCAGAAAA AAAAAAAAAA AGTCGACGCG 2439 GCCGCGAATT CCTGCAG 2456 (2) INFORMATION FOR SEQ ID NO: 2: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 422 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 2: Met Ser Ser Cys Ser Gly Leu Ser Arg Val Leu Val Ala Val Ala 1 5 10 15 Thr Ala Leu Val Ser Ala Ser Ser Pro Cys Pro Gln Ala Trp Gly Pro 20 25 30 Pro Gly Val Gln Tyr Gly Gln Pro Gly Arg Ser Val Lys Leu Cys Cys 35 40 45 Pro Gly Val Thr Ala Gly Asp Pro Val Ser Trp Phe Arg Asp Gly Glu 50 55 60 Pro Lys Leu Leu Gln Gly Pro Asp Ser Gly Leu Gly His Glu Leu Val 65 70 75 80 Leu Wing Gln Wing Asp Ser Thr Asp Glu Gly Thr Tyr He Cys Gln Thr 85 90 95 Leu Asp Gly Wing Leu Gly Gly Thr Val Thr Leu Gln Leu Gly Tyr Pro 100 105 110 Pro Wing Arg Pro Val Val Ser Cys Gln Wing Wing Asp Tyr Glu Asn Phe 115 120 125 Ser Cys Thr Trp Ser Pro Ser Gln He Ser Gly Leu Pro Thr Arg Tyr 130 135 140 Leu Thr Ser Tyr Arg Lys Thr Val Leu Gly Wing Asp Ser Gln Arg 145 150 155 160 Arg Ser Pro Be Thr Gly Pro Trp Pro Cys Pro Gln Asp Pro Leu Gly 165 170 175 Wing Wing Arg Cys Val Val His Gly Wing Glu Phe Trp Ser Gln Tyr Arg 180 185 190 He Asn Val Thr Glu Val Asn Pro Leu Gly Ala Ser Thr Arg Leu Leu 195 200 205 Asp Val Ser Leu Gln Ser He Leu Arg Pro Asp Pro Pro Gln Gly Leu 210 215 220 Arg Val Glu Ser Val Pro Gly Tyr Pro Arg Arg Leu Arg Ala Ser Trp 225 230 235 240 Thr Tyr Pro Wing Ser Trp Pro Cys Gln Pro His Phe Leu Leu Lys Phe 245 250 255 Arg Leu Gln Tyr Arg Pro Wing Gln His Pro Wing Trp Ser Thr Val Glu 260 265 270 Pro Wing Gly Leu Glu Glu Val He Thr Asp Wing Val Wing Wing Gly Leu Pro 275 280 285 His Wing Val Arg Val Wing Wing Arg Asp Phe Leu Asp Wing Gly Thr .Trp 290 295 300 Ser Thr Trp Ser Pro Glu Wing Trp Gly Thr Pro Ser Thr Gly Thr He 305 310 315 320 Pro Lys Glu He Pro Wing Trp Gly Gln Leu His Thr Gln Pro Glu Val 325 330 335 Glu Pro Gln Val Asp Pro Pro Wing Pro Pro Arg Pro Ser Leu Gln Pro 340 345 350 His Pro Arg Leu Leu Asp His Arg Asp Ser Val Glu Val Val Gl Val 355 360 365 Leu Ala Ser Leu Gly He Leu Ser Phe Leu Gly Leu Val Ala Gly Ala 370 375 380 Leu Ala Leu Gly Leu Trp Leu Arg Leu Arg Arg Gly Gly Lys Asp Gly 385 390 395 400 Ser Pro Lys Pro Gly Phe Leu Wing Ser Val He Pro Val Asp Arg Arg 405 410 415 Pro Gly Ala Pro Asn Leu 420 (2) INFORMATION FOR SEQ ID NO: 3: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 1714 base pairs (B) TYPE: nucleic acid (C) TYPE OF CHAIN: double (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: cDNA (iii) HYPOTHETICAL: No (ix) CHARACTERISTICS: (A) NAME / KEY: CDS (B) LOCATION: 34..1359 (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 3: TTCTTAGCCT GATAGGAGGA AGTCTTGGAG GCC ATG GCA CTC AGT CAC TGT GAT 54 Met Ala Leu Ser His Cys Asp 1 5 TAT CAA GAT GAG CAG CAG CTG CTC AGG GCT GAC CAG GGT CCT GGT GGC 102 Tyr Gln Asp Glu Gln Gln Leu Leu Arg Ala Asp Gln Gly Pro Gly Gly 10 15 20 CGT GCT ACÁ GCC CTG GTG TCT TCC TCC TCC CCC TGC CCC CAÁ GCT TGG 150 Arg Ala Thr Ala Leu Val Ser Ser Ser Pro Pro Cys Pro Gln Ala Trp 25 30 35 GGT CCT CCA GGG GTC CAG TAT GGA CAA CCT GGC AGG CCC GTG ATG CTG 198 Gly Pro Pro Gly Val Gln Tyr Gly Gln Pro Gly Arg Pro Val Met Leu 40 45 50 55 TGC TGC CCC GGA GTG AGT GCT GGG ACT CCA GTG TCC TGG TTT CGG GAT 246 Cys Cys Pro Gly Val Be Wing Gly Thr Pro Val Ser Trp Phe Arg Asp 60 65 70 GGA GAT TCA AGG CTG CTC CAG GGA CCT GAC TCT GGG TTA GGA CAC AGA 294 Gly Asp Ser Arg Leu Leu Gln Gly Pro Asp Ser Gly Leu Gly His Arg 75 80 85 CTG GTC TTG GCC CAG GTG GAC AGC CCT GAT GAA GGC ACT TAT GTC TGC 342 Leu Val Leu Ala Gln Val Asp Ser Pro Asp Glu Gly Thr Tyr Val Cys 90 95 100 CAG ACC CTG GAT GGT GTA TCA GGG ATG GTG ACC CTG AAG CTG GGC 390 Gln Thr Leu Asp Gly Val Ser Gly Gly Met Val Thr Leu Lys Leu Gly 105 110 115 TTT CCC CCA GCA CGT CCT GAA GTC TCC TGC CAG GCG GTA GAC TAT GAA 438 Phe Pro Pro Wing Arg Pro Glu Val Ser Cys Gln Wing Val Asp Tyr Glu 120 125 130 135 AAC TTC TTC TGT ACT TGG AGT CCA GGC CAG GTC AGC GGT TTG CCC ACC 486 Asn Phe Ser Cys Thr Trp Ser Pro Gly Gln Val Ser Gly Leu Pro Thr 140 145 150 CGC TAC CTT ACT TCC TAC AGG AAG AAG ACG CTG CCA GGA GCT GAG AGT 534 Arg Tyr Leu Thr Ser Tyr Arg Lys Lys Thr Leu Pro Gly Wing Glu Ser 155 160 165 CAG AGG GAA AGT CCA TCC ACC GGG CCT TGG CCG TGT CCA CAG GAC CCT 582 Gln Arg Glu Ser Pro Be Thr Gly Pro Trp Pro Cys Pro Gln Asp Pro 170 175 180 CTG GAG GCC TCC CGA TGT GTG GTC CAT GGG GCA GAG TTC TGG AGT GAG 630 Leu Glu Wing Ser Arg Cys Val Val His Gly Wing Glu Phe Trp Ser Glu 185 190 195 TAC CGG ATC AAT GTG ACC GAG GTG AAC CCA CTG GGT GCC AGC ACG TGC 678 Tyr Arg He Asn Val Thr Glu Val Asn Pro Leu Gly Ala Ser Thr Cys 200 205 210 215 CTA CTG GAT GTG AGA TTA CAG ^ GC ATC TTG CGT CCT GAT CCA CCC CAA 726 Leu Leu Asp Val Arg Leu Gln Ser He Leu Arg Pro Asp Pro Pro Gln 220 225 230 GGA CTG CGG GTG GAA TCC GTA CCT GGT TAC CCG AGA CGC CTG CAT GCC 774 Gly Leu Arg Val Glu Ser Val Pro Gly Tyr Pro Arg Arg Leu His Wing 235 240 245 AGC TGG ACÁ TAC CCT GCC TCC TGG CGT CGC CAAC CCC CAC TTT CTG CTC 822 Ser Trp Thr Tyr Pro Wing Ser Trp Arg Arg Gln Pro His Phe Leu Leu 250 255 260 AAG TTC CGG TTG CAA TAC CGA CCA GCA CAG CAT CCA GCC TGG TCC ACG 870 Lys Phe Arg Leu Gln Tyr Arg Pro Wing Gln His Pro Wing Trp Ser Thr 265 270 275 GTG GAG CCC ATT GGC TTG GAG GAA GTG ATA ACA GAT GCT GTG GCT GGG 918 Val Glu Pro He Gly Leu Glu Glu Val He Thr Asp Ala Val Wing Gly 280 285 290 295 CTG CCA CAC GCG GTA CGA GTC AGT GCC AGG GAC TTT CTG GAT GCT GGC 966 Leu Pro His Wing Val Arg Val Wing Wing Arg Asp Phe Leu Asp Wing Gly 300 305 310. ACC TGG AGC GCC TGG AGC CCA GAG GCC TGG GGT ACT CCT AGC ACT GGT 1014 Thr Trp Ser Wing Trp Ser Pro Glu Wing Trp Gly Thr Pro Ser Thr Gly 315 320 325 CCC CTG CAG GAT GAG ATA CCT GAT TGG AGC CAG GGA CAT GGA CAG CAG 1062 Pro Leu Gln Asp Glu He Pro Asp Trp Ser Gln Gly His Gly Gln Gln 330 335 340 CTA GAG GCA GTA GTA GCT GAC GAC AGC CCG GCT CCT GCA AGG CCT 1110 Leu Glu Ala Val Val Ala Gln Glu Asp Ser Pro Wing Pro Wing Arg Pro 345 350 355 TCC TTG CAG CCG GAC CCA AGG CCA CTT GAT CAC AGG GAC CCC TTG GAG 1158 Ser Leu Gln Pro Asp Pro Arg Pro Leu Asp His Arg Asp Pro Leu Glu 360 365 370 375 CAA GTA GCT GTG TTA GCG TCT CTG GGA ATC TTC TCT TGC CTT GGC CTG 1206 Gln Val Wing Val Leu Wing Ser Leu Gly He Phe Ser Cys Leu Gly Leu 380 385 390 GCT GTT GGA GCT CTG GCA CTG GGG CTC TGG CTG AGG CTG AGA CGG AGT 1254 Wing Val Gly Ala Leu Ala Leu Gly Leu Trp Leu Arg Leu Arg Arg Ser 395 400 405 GGG AAG GAT GGA CCG CAA AAA CCT GGG CTC TTG GCA CCC ATG ATC CCG 1302 Gly Lys Asp Gly Pro Gln Lys Pro Gly Leu Leu Al a Pro Met He Pro 410 415 420 GTG GAA AAG CTT CCA "GGA ATT CCA AAC CTG CAG AGG ACC CCA GAG AAC 1350 Val Glu Lys Leu Pro Gly He Pro Asn Leu Gln Arg Thr Pro Glu Asn 425 430 435 TTC AGC TGATTTCATC TGTAACCCGG TCAGACTTGG GGTGGTTAAA AGGACAGGCA 1406 Phe Ser 440 GAAAGAGGCG GGGCAGTGGA TCCCTGTGGA TGGAGGTCTC AGCTGAAAGT CTGAGCTCTT 1466 TTCTTTGACA CCTATACTCC AAACTTGCTG CCGGCTGAAG GCTGTCTGGA CTTCCGATGT 1526 CCTGAGGTGG AAGTCCACCT GAGGAATGTG TACAGAAGTC TGTGTTCCTG TGATCGTGTG 1586 TGTATGTGAG ACAGGGAGCA AAAGTTCTCT GCATGTGTGT ACAGATGATT GGAGAGTGTG 1646 TGCGGTCTTG GGCTTGGCCC TTCTGGGAAG TGTGAAGAGT TGAAATAAAA GAGACGGAAG 1706 TTTTTGGA 1714 (2) INFORMATION FOR SEQ ID NO: 4: (i) CHARACTERISTICS OF THE SEQUENCE: (A) LENGTH: 441 amino acids (B) TYPE: amino acid (D) TOPOLOGY: linear (ii) TYPE OF MOLECULE: protein (xi) DESCRIPTION OF THE SEQUENCE: SEQ ID NO: 4: Met Ala Leu Ser His Cys Asp Tyr Gln Asp Glu Gln Gln Leu Leu Arg 1 5 10 15 Wing Asp Gln Gly Pro Gly Gly Arg Wing Thr Wing Leu Val Being Ser - Ser 20 25 30 Ser Pro Cys Pro Gln Wing Trp Gly Pro Pro Gly Val Gln Tyr Gly Gln 35 40 45 Pro Gly Arg Pro Val Met Leu Cys Cys Pro Gly Val Ser Wing Gly Thr 50 55 60 Pro Val Ser Trp Phe Arg Asp Gly Asp Ser Arg Leu Leu Gln Gly Pro 65 70 75 80 Asp Ser Gly Leu Gly His Arg Leu Val Leu Wing Gln Val Asp Ser Pro 85 90 95 Asp Glu Gly Thr Tyr Val Cys Gln Thr Leu Asp Gly Val Ser Gly Gly 100 105 110 Met Val Thr Leu Lys Leu Gly Phe Pro Pro Wing Arg Pro Glu Val Ser 115 120 125 Cys Gln Wing Val Asp Tyr Glu Asn Phe Ser Cys Thr Trp Ser Pro Gly 130 135 140 Gln Val Ser Gly Leu Pro Thr Arg Tyr Leu Thr Ser Tyr Arg Lys Lys 145 150 155 160 Thr Leu Pro Gly Wing Glu Ser Gln Arg Glu Ser Pro Thr Gly Pro 165 170 175 Trp Pro Cys Pro Gln'Asp Pro Leu Glu Wing Ser Arg Cys Val Val His 180 185 190 Gly Wing Glu Phe Trp Ser Glu Tyr Arg He Asn Val Thr Glu Val Asn 195 200 205 Pro Leu Gly Wing Be Thr Cys Leu Leu Asp Val Arg Leu Gln Ser He 210 215 220 Leu Arg Pro Asp Pro Pro Gln Gly Leu Arg Val Glu Ser Val Pro Gly 225 230 235 240 Tyr Pro Arg Arg Leu His Wing Ser Trp Thr Tyr Pro Wing Ser Trp Arg 245 250 255 Arg Gln Pro His Phe Leu Leu Lys Phe Arg Leu Gln Tyr Arg Pro Wing 260 265 270 Gln His Pro Wing Trp Ser Thr Val Glu Pro He Gly Leu Glu Glu Val 275 280 285 He Thr Asp Wing Val Wing Gly Leu Pro His Wing Val Arg Val Ser Wing 290 295 300 Arg Asp Phe Leu Asp Wing Gly Thr Trp Ser Wing Trp Ser Pro Glu Wing 305 310 315 320 Trp Gly Thr Pro Ser Thr Gly Pro Leu Gln Asp Glu He Pro Asp Trp 325 330 335 Being Gln Gly His Gly Gln Gln Leu Glu Wing Val Val Wing Gln Glu Asp 340 345 350 Ser Pro Wing Pro Wing Arg Pro Ser Leu Gln Pro Asp Pro Arg Pro Leu 355 360 365 Asp His Arg Asp Pro Leu Glu Gln Val Wing Val Leu Wing Ser Leu Gly 370 375 380 He Phe Ser Cys Leu Gly Leu Wing Val Gly Wing Leu Wing Leu Gly Leu 385 390 395 400 Trp Leu Arg Leu Arg Arg Ser Gly Lys Asp Gly Pro Gln Lys Pro Gly 405 410 415 Leu Leu Wing Pro Met He Pro Val Glu Lys Leu Pro Gly He Pro Asn 420 425 430 Leu Gln Arg Thr Pro Glu Asn Phe Ser

Claims (30)

1. Novelty of the Invention 1. An isolated polynucleotide comprising a nucleotide sequence selected from the group consisting of: (a) the nucleotide sequence of SEQ ID No .: 1, from nucleotide 803 to nucleotide 1999; (b) a nucleotide sequence that varies from the sequence of the nucleotide sequence specified in (a) as a result of a degeneracy of the genetic code; and (c) an alollic variant of the nucleotide sequence specified in (a).
2. The polynucleotide of claim 1, wherein said nucleotide sequence codes for a protein having a biological activity of the human IL-11 receptor.
3. 3. The polynucleotide of claim 1, wherein said nucleotide sequence is operably linked to an expression control sequence.
4. 4. The polynucleotide of claim 2, comprising the nucleotide sequence of SEQ ID No .: 1, from nucleotide 803 to nucleotide 1999.
5. 5. The polynucleotide of claim 2, comprising the nucleotide sequence of SEQ ID No .: 1, from nucleotide 803 to nucleotide 1828 or a fragment thereof.
6. 6. The polynucleotide of claim 2, comprising the nucleotide sequence of SEQ ID No .: 1, from nucleotide 1907 to nucleotide 1999 or a fragment thereof.
7. 7. The polynucleotide of claim 1, comprising the nucleotide sequence of SEQ ID No .: 1, from nucleotide 734 to nucleotide 1999.
8. 8. The polynucleotide of claim 1, comprising the nucleotide sequence of SEQ ID No .: 1, from nucleotide 1607 to nucleotide 1828.
9. 9. The polynucleotide of claim 1, comprising the nucleotide sequence of SEQ ID No .: 1, from nucleotide 1607 to nucleotide 1999.
10. 10, A host cell transformed with the polynucleotide of claim 3,
11. 11. The host cell of claim 8, wherein said cell is a mammalian cell.
12. 12. A process for producing a human IL-11R protein, said process comprising: (a) growing a culture of the host cell of claim 10 in a suitable culture medium; and (b) purifying the human IL-11 R protein from said culture.
13. 13. An isolated, human IL-11 R protein, which comprises an amino acid sequence selected from the group consisting of: (a) the amino acid sequence of SEQ ID NO: 2; (b) the amino acid sequence of SEQ ID NO: 2; of amino acid 24 to 422; (c) the amino acid sequence of SEQ ID NO: 2; from amino acid 24 to (d) the amino acid sequence of SEQ ID NO: 2; of amino acid 391 to 422; (e) the amino acid sequence of SEQ ID NO: 2; of amino acid 102 to 422; (f) the amino acid sequence of SEQ ID NO: 2; of amino acid 102 to 365; and (g) fragments of (a) - (f) having a biological activity of the human IL-11 receptor.
14. 14. The protein of claim 13, comprising the amino acid sequence of SEQ ID No .: 2.
15. 15. The protein of claim 13, comprising the amino acid sequence 24 to 365 of SEQ ID No: 2.
16. 16. A pharmaceutical composition comprising a protein of claim 13 and a pharmaceutically acceptable carrier.
17. 17. A protein produced in accordance with the process of claim 12.
18. 18. A composition comprising an antibody that specifically reacts with a protein of claim 13.
19. 19. A method for identifying an inhibitor of the binding of IL-11 to the human IL-11 receptor, which comprises: (a) combining a protein of claim 13 with IL-11 or a fragment thereof, said combination forming a first binding mixture; (b) measuring the degree of binding between the protein and IL-11 or fragment in the first binding mixture; (c) combining a compound with the protein and the IL-11 or fragment to form a second binding mixture; (d) measuring the degree of binding in the second binding mixture; and (e) comparing the degree of binding in the first binding mixture with the degree of binding in the second binding mixture; wherein the compound is capable of inhibiting the binding of IL-11 to the human IL-11 receptor when a decrease in the degree of binding of the second binding mixture occurs,
20. 20, The method of claim 19, wherein the first and second binding mixtures comprise gp130 or a fragment, capable of binding to the protein of claim 13 or to IL-11 or a fragment used therein.
21. 21. An inhibitor identified by the method of claim 19.
22. 22. A pharmaceutical composition comprising the inhibitor of claim 21 and a pharmaceutically acceptable carrier.
23. 23. A method to inhibit the binding of IL-11 to the receptor d? human IL-11 in a mammalian subject, said method comprising administering a therapeutically effective amount of a composition of claim 22.
24. 24. A method for inhibiting the binding of IL-11 to the human IL-11 receptor in a mammalian subject, said method comprising administering a therapeutically effective amount of a composition of claim 16.
25. 25. A method for inhibiting the binding of IL-11 to the human IL-11 receptor in a mammalian subject, said method comprising administering a therapeutically effective amount of a composition of claim 18.
26. 26. A method for treating or preventing the loss of bone mass in a mammalian subject, said method comprising administering a therapeutically effective amount of a composition of claim 22.
27. 27. A method for treating or preventing the loss of bone mass in a mammalian subject, said method comprising administering a therapeutically effective amount of a composition of claim 16.
28. 28. A method for treating or preventing the loss of bone mass in a mammalian subject, said method comprising administering a therapeutically effective amount of a composition of claim 18.
29. 29. An isolated polynucleotide comprising a nucleotide sequence capable of hybridizing under stringent conditions to the polynucleotide of claim 4.
30. 30. An isolated polynucleotide comprising a sequence of nucleotides encoding a peptide or protein comprising an amino acid sequence selected from the group consisting of: (a) the amino acid sequence of SEQ ID NO: 2; (b) the amino acid sequence of SEQ ID NO: 2; of amino acid 24 to 422; (c) the amino acid sequence of SEQ ID NO: 2; of amino acid 24 to 365; (d) the amino acid sequence of SEQ ID NO: 2; of amino acid 391 to 422; (e) the amino acid sequence of SEQ ID NO: 2; of amino acid 112 to 422; (f) the amino acid sequence of SEQ ID NO: 2; of amino acid 112 to 365; and (g) fragments of (a) - (f) having a biological activity of the human IL-11 receptor.