MX2008005638A

MX2008005638A - Novel activin receptor and uses thereof

Info

Publication number: MX2008005638A
Application number: MXMX/A/2008/005638A
Authority: MX
Inventors: Han Hq; Soonyung Kwak Keith; Zhou Xiaolan
Original assignee: Amgen Inc; Han Hq; Soonyung Kwak Keith; Zhou Xiaolan
Priority date: 2005-11-01
Filing date: 2008-04-30
Publication date: 2008-10-03

Abstract

The present invention provides novel activin IIB5 receptor polypeptides capable of binding and inhibiting the activities of activin A, myostatin, or GDF-11. The present invention also provides polynucleotides, vectors and host cells capable of producing the receptor polypeptides. Compositions and methods for treating muscle-wasting, metabolic and other disorders are also provided.

Description

NOVEDOUS ACTIVINE RECEPTOR AND USES OF THE SAME Field of the Invention The technical field of this invention relates to transforming TGF-β receptors and members of the β-growth factor family (TGF-β), as well as methods of modulating the activities of family members. TGF-β for the treatment of various disorders. Background of the Invention, The transformation of the family of the growth factor β | (TGF-), of the proteins includes the transformation of the growth factor ß (TGF-ß), activins, bone morphogenic proteins (BMP), nerve growth factors (NGFs), brain-derived neurotropic factor (BDNF), and growth / differentiation factors (GDFs). These family members are involved in the regulation of a wide range of biological processes that include cell proliferation, differentiation, and other functions. Activins were originally discovered as gonadal peptides involved in the regulation of the synthesis of follicular hormone stimulation, and are now believed to be involved in the regulation of a number of biological activities including control of the section and expression of anterior pituitary hormones such as FSH, GH, and ACTH, neuronal survival, secretion of hypothalamic oxytocin, erythropoiesis, gonodal steroidogenesis and placental, early embryonic development, and proliferation of some types of tumors. Activins A, B and AB are the homodimers and heterodimers respectively of two polypeptide chains, BA and BB (Vale et al. Nature 321, 776-779 (1986), Ling et al. Nature 321, 779-782 (1986)). These two β chains can also be dimerized with a related chain that gives an increase to inhibit A and B respectively (Mason et al., Nature 318, 659-663 (1986)). It is well established that inhibins are necessary to maintain normal function in many tissues, particularly those associated with reproductive functions. In these tissues inhibins oppose many, but not all, activities of activin. The growth / differentiation factor 8 (GDF-8), also referred to as myostatin, is a member of the TGF-β family expressed mostly in adult skeletal muscle development and tissue cells. Myostatin seems to play an essential role in the negatively controlled growth of skeletal muscle (McPherron et al. Nature (London) 387, 83-90 (1997)). Antagonization of myostatin has been shown to increase lean muscle mass in animals (McFerron et al., Supra, Zimmers et al., Science 296: 1486 (2002)). Another member of the TGF-β protein family is a related GDF-II growth / differentiation factor. GDF-II has an identity of approximately 90% of the amino acid sequence of myostatin. GDF-II has a role in the axial pattern in the development of animals (Oh et al., Genes Dev 11: 1812-26 (1997)), and also appears to play a role in the development and growth of skeletal muscle. However, the postnatal role of GDF-11 is not currently understood. A family of transmembrane serine / threonine kinases is known to act as receptors for activins and other members of the TGF-β family. These receptors fall into two distinct subfamilies known as type I and type II receptors that act cooperatively to bind a ligand and transduce the signal (Attisano et al., Mol Cell Biol 16 (3), 1066-1073 (1996)). The majority of TGF-β ligands are thought to bind first to a type II receptor and this ligand / type II receptor complex then recruits a type I receptor (Mathews, LS, Endocr Rv 15: 310-325 (1994); Nature Rev: Mol Cell Biol. 1, 169-178 (2000)). The type II receptor kinase then phosphors and activates the type I receptor kinase, which in turn phosphors Smad proteins. Activins bind initially to their type II receptors Act RIIA for activin A, or ActRIIB for activin B. This is followed by recruitment, phosphorylation and subsequent activation of the type I receptor, activin as well as kinase 4 (ALK4). In activation, ALK4 binds and then phosphors a subset of cytoplasmic proteins Smad (Smad2 and Smad3) which produces signal transduction for fortune tellers (Derynck, R et al., Ce.ll 95, 737-740 (1998)). Cross-linking studies have determined that myostatin is capable of binding activin type II ActRIlA and ActRIIB receptors in vitro (Lee et al., PNAS USA 98: 9306-11 (2001)). There is also evidence that GDF-11 links both ActRIlA and ActRIIB (Oh et al., Genes Dev 16: 2749-54 (2002)). TGF-β proteins are known to be associated with a variety of disease states and antagonizing these proteins may be useful as therapeutic treatments for disease states. In particular the antagonization of several TGF-β proteins simultaneously may be particularly effective in treating certain diseases. The present invention provides a composition of matter and novelty methods for using the composition of matter as a treatment for disorders and others related to muscle. Brief Description of the Invention The present invention provides a protein comprising human activin IIB receptor polypeptides (designated ActRIIB5). In one embodiment, the protein comprises polypeptides having an amino acid sequence set forth in SEQ ID NO: 2. In another embodiment, the protein comprises a polypeptide having an amino acid sequence with less about 80% or greater identity with SEQ ID NO: 2, wherein the polypeptide is capable of binding to myostatin, activin A, or GDF-II. In another embodiment, the protein comprises a polypeptide having an amino acid sequence with at least about 80% or greater identity with SEQ ID NO: 2, wherein the C-terminus of the polypeptide consists of the amino acid sequence set forth in SEQ ID NO: 3, and wherein the polypeptide is capable of binding to myostatin, activin A, or GDF-II. In another embodiment, the protein comprises a polypeptide having an amino acid sequence with at least about 80% or greater identity with SEQ ID NO: 2, wherein the C-terminus of the polypeptide has one. amino acid sequence with about at least 80% or greater identity with SEQ ID NO: 3, and wherein the polypeptide is capable of binding to myostatin, activin A, or GDF-11. In one embodiment, the polypeptide lacks an ActRIIB5 signal sequence. In another embodiment, the protein comprises a polypeptide encoded by the polynucleotide having the establissequence SEQ ID NO: 1. In another embodiment, the protein of the present invention comprises ActRIIB5 polypeptides fused to one or more heterologous polypeptides. In one embodiment, the fused ActRIIB5 polypeptides lack a signal sequence. In one embodiment the ActRIIB5 polypeptides are fused to the heterologous polypeptides via one or more linkers of sequences. In another embodiment, the heterologous polypeptides comprise an Fe domain. In another embodiment, the Fe domain is connected to the ActRIIB5 polypeptides by at least one sequence linker. In another embodiment, the ActRIIB5 polypeptides are linked to a non-protein carrier molecule such as a PEG molecule. In another aspect the present invention provides an isolated nucleic acid molecule comprising a polynucleotide encoding an ActRIIB5 polypeptide. In one embodiment, the nucleic acid molecule comprises (a) a polynucleotide having the nucleic acid sequence set forth in SEQ. ID NO: 1 or its complement. In another embodiment, the nucleic acid molecule comprises (b) a polynucleotide that encodes a polypeptide consisting of the amino acid sequence set forth in SEQ ID NO: 2 or its complement. In another embodiment, the nucleic acid molecule comprises (c) a polynucleotide that hybridizes to (a) or (b) under conditions at least moderately rigorously in about 50% formamide, 6X SSC at approximately 42 ° C and washing at about 60 ° C, 0.5X SSC, 0.1% SDS, and wherein the encoded polypeptide comprises a C-terminal having an amino acid sequence set forth in SEQ ID NO: 3, and wherein the polypeptide is capable of binding myostatin , activin A, or GDF-II. In another embodiment, the nucleic acid molecule comprises the polynucleotide of (c) wherein the C-terminal of the encoded polypeptide has an amino sequence of at least about 80% or greater than SEQ ID NO: 3, and wherein the polypeptide is capable of binding to myostatin, activin A, or GDF-11. In another embodiment, the nucleic acid molecule comprises a polynucleotide having at least about 80% or greater identity to SEQ ID NO: 1. In another embodiment, the nucleic acid molecule of the present invention further comprises polynucleotides that encode at least one heterologous protein in the structure with the polynucleotides encoding an ActRIIB5 polypeptide. In one embodiment, the nucleic acid molecule comprises the polynucleotides that encode the peptide linker sequences that bind to the ActRIIB5 polypeptide in at least one heterologous protein. In another embodiment, the heterologous protein is an Fe polypeptide. The present invention further provides a vector comprising the nucleic acid molecules set forth above, as well as a genetically engineered host cell designed to express the nucleic acid molecules described above, and methods of producing the same. ActRIIB5 protein. The present invention further provides a composition containing the protein of the present invention. In one embodiment, the composition is a pharmaceutical composition that contains the protein in the mixture with a carrier pharmaceutically acceptable. In another aspect, the present invention provides a method of inhibiting the activity of the myostatin TGF-β, activin or GDF-II proteins in vitro and in vivo by contacting the proteins with an ActRIIB5 polypeptide. In another aspect the present invention provides a method of increasing lean muscle mass and strength, and a method of increasing the muscle tissue ratio with fat in a subject in need thereof by administering an effective amount of the composition that contains the ActRIIB5 proteins to the subject. In one embodiment of this method, the subject is a fed animal. In another aspect, the present invention provides a method of treating or preventing a muscle wasting disease in a subject suffering such a disorder by administering a therapeutic composition containing an ActRIIB5 protein to the subject. Muscle consumptive disease includes or results from, but is not limited to, the following conditions: muscular dystrophy, amyotrophic lateral sclerosis, congestive obstructive pulmonary disease, chronic cardiac arrest, cancer cachexia, chemical cachexia, HIV / AIDS, renal failure, uremia, rheumatoid arthritis, sarcopenia related to age, organ atrophy, carpal tunnel syndrome, androgen deficiency, and muscle wasting due to inactivity such as prolonged bed rest, spinal cord injury, myocardial infarction, fracture of bone, aging. The wear of Muscle can also result from events such as weightlessness of space flight, insulin resistance, muscle wasting due to burns, androgen deficiency, and other disorders. In another aspect, the present invention provides a method of treating a disease correlated to the expression of activin A. In one embodiment, the disease is cancer. In another aspect, the present invention provides a method of treating a metabolic disorder comprising administering a therapeutic composition to a subject in need of such treatment, wherein the metabolic disorder is selected from diabetes, obesity, impaired glucose tolerance, hyperglycemia, lack of androgen, metabolic syndrome, and bone loss. In another aspect, the present invention provides a method of gene therapy comprising administering a vector encoding the ActRIIB5 proteins of the protein of the present invention to a subject in need thereof, wherein the vector is capable of expressing the ActRBII5 polypeptide. in the subject. The present invention further provides a method of detection and quantification of the TGF-β proteins of myostatin, GDF-11 or activin A by contacting these proteins with an ActRIIB5 polypeptide and detecting the polypeptide. Brief Description of the Figures Figure 1 shows the results of the Biacore® assay determination of EC50 by ActRIIB5 / Fc compared to ActRIIB-ECD / Fc. Figure 2 shows the increase in body weight at some time in C57BI / 6 mice injected with AAV-activin A, AAV-promyostatin / Fc, AAV-ActRI I B5 / Fc, AAV-ActRIIB-ECD / Fc and control empty vector of AAV. Figure 3 shows the percentage of body weight change compared to control in post viral infection in seven weeks in C57BI / 6 mice injected with AAV-activin A, AAV-ActRIIB5 / Fc, AAV-ActR I IB-ECD / Fc , and AAV-promyostatin / Fc vector. Figure 4A shows a decrease in body weight at some time for obese A mice injected with AAV-ActRIIB5 / Fc compared to a control group of obese A mice injected with empty AAV vectors over a period of about three months. Figure 4B shows a decrease in weekly food absorption for the same group of AAV-ActRIIB5 mice compared to the control group in the same time period. Figure 5A shows the change in lean body mass at a certain time by obese A mice injected with AAV-ActRI IB5 / Fc compared to a control group of obese A mice injected with empty AAV vector for approximately three months. Figure 5B shows a large decrease in fat mass for the AAV-ActRIIB5 / Fc mice compared to a control group of the mice of AAV-vacuum in the same period of time. Detailed Description of the Invention The present invention provides a novel human activin receptor designated activin IIB5 receptor (ActRIIB5). This receptor is characterized by its ability to bind to three TGF-β proteins, myostatin (GDF-8), activin A, and GDF-11, and to inhibit the activities of these proteins. As used herein the term "members of the TGF-β family" or "TGF-β proteins" refers to growth factors structurally related to the transformation of the growth factor family including activins, and growth factor and differential (GDF) (Kinglsey et al., Genes Dev. 8: 133-146 (1994), McPherron et al., Growth factors and cytokines in health and disease, Vol IB, D. LeRoith and CBondy. ed., JAI Press Inc., Greenwich, Conn, USA: pp 357-393). GDF-8, also referred to as myostatin, is now known to be a negative regulator of skeletal muscle tissue (McPherron et al., PNAS USA 94: 12457-12461 (1997)). Myostatin is synthesized as an approximately inactive prepotenic complex. of 375 amino acids in length, which have GenBank Access No: AAB86694 for human. The precursor protein is activated by the proteolytic cleavage at a tetrabasic processing site to produce an inactive N-terminal prodomain and a C-terminal protein of the approximately 109 amino acid that dimerizes to form a homodimer of approximately 25 kDa. This homodimer is the mature, biologically active protein (Zimmers et al., Science 296, 1486 (2002)). As used herein, the term "prodomain" or "propeptide" refers to the inactive N-terminal protein that is cleaved to release the active C-terminal protein. As used herein the term "myostatin" or "mature myostatin" refers to the mature, biologically active terminal C-polypeptide in the monomer, dimer or other form, as well as biologically active related fragments or polypeptides including allelic variants, variants of binding, and fusion of peptides and polypeptides. Mature myostatin has been indicated to have 100% sequence identity among many species including human, mouse, chicken, porcine, turkey, and rat (Lee et al., PNAS 98, 9306 (2001)). As used herein, GDF-11 refers to the BMP protein having Swiss Pro accession number 095390, as well as variants and homologous species of that protein. GDF-11 has approximately 90% identity with myostatin at the amino acid level. GDF-11 is involved in the regulation of the anterior / posterior pattern of the axial skeleton (McPherron et al., Natr Genet 22 (93): 260-264 (1999); Gamer et al., Dev. Biol. 208 (1), 222- 232 (1999)) but postnatal functions are unknown. Activin A is the homodimer of BA polypeptide chains. As used herein the term "activin A" refers to to the activin protein that has GenBank Accession No: NM_002192, as well as variants and homologous species of the protein. Activin Receptors As used herein, the term "activin type II receptor" (ActRIlB) refers to the human activin receptor precursor having accession number NP_001097 for the protein or any variant or homologs of this receptor. The amino acid and polynucleotide sequences precursor of human ActRIlB are set forth in SEQ ID NO: 4 and 5 respectively. A variation of ActRIlB is set forth in SEQ ID NO: 6, wherein the arginine at position 64 has been replaced by alanine. SEQ ID NO: 5 is referred to as the R form and SEQ ID NO: 6 is referred to as the A form. The extracellular domain of ActRIlB (ActRIIB-ECD) is represented by amino acids 1 to 124 of SEQ ID NO: 5 and 6. The additional murine isoforms for this receptor have been identified as muActRIIBI, muActR! IB2, muActRIIB3 and muActRIIB4. The present invention provides a novel human activin receptor designated as activin receptor IIB5 (ActRIIB5). This receptor is characterized by the terminal sequence C set forth in SEQ ID NO: 3. The cDNA of this receptor was isolated as described in example 1, and found to be missing from the nucleotide bases 152 corresponding to exon 4. This receiver is also characterized as missing from the transmembrane region encoded by exon 4 of ActRIIB. This receptor is also characterized as being a soluble, secreted instead of a receptor that binds to the membrane. The receptor is further characterized as having the ability to bind and inhibit the activity of any activin A, myostatin, or GDF-11. The present invention provides isolated proteins comprising polypeptides of the ActllB5 receptor. As used herein the term "isolated" refers to a nucleic acid molecule purified to a certain degree of endogenous material. In one embodiment, the protein comprises ActRIIB5 polypeptides having the amino acid sequence set forth in SEQ ID NO: 2, and variants and derivatives of this polypeptide, which retain the activity of the polypeptide of SEQ ID NO: 2 In one embodiment, the protein comprises a polypeptide having at least about 80% identity, at least about 85% identity, at least about 90% identity, at least about 95% identity, at least about 98% identity , or at least about 99% identity with the amino acid sequence set forth in SEQ ID NO: 2, wherein the polypeptide retains the activity of the polypeptide of SEQ ID NO: 2. In another embodiment, the protein comprises ActRIIB5 polypeptides described above wherein the polypeptide has a terminal C comprising the amino acid sequence set forth in SEQ ID NO: 3, and wherein the polypeptide retains the activity of the polypeptide of SEQ ID NO: 2. In other embodiments, the protein comprises ActRIIB5 polypeptides described above wherein the terminal C has an amino acid sequence having at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 98%, or at least about 99% of identity with SEQ ID NO: 3, wherein the polypeptide retains the activity of the polypeptide of SEQ ID NO: 2. In one embodiment, the polypeptide ActRIIB5 lacks a signal sequence of SEQ ID NO: 2, for example , amino acids 1 to 17 of SEQ ID NO: 2. As used herein the term "variant" refers to a polypeptide having one or more amino acids grafted, deleted or substituted in the original sequence of the amino acid, but they have a sequence which remains substantially similar to SEQ ID NO: 2, and which retains the activities of the ActRIIBS polypeptides of SEQ ID NO: 2. As used herein fragments of the polypeptides that retain the activity of the polypeptides they are included in the term "variants". For purposes of the present invention, "substantially similar" is at least about 80% identical to the amino acid sequence, at least about 85% identical, at least about 90% identical, at least about 95% identical, at least about 98% identical, at least about 99% identical to the amino acid sequence set forth in S'EC ID NO: 2, and retain the biological activities of the polypeptide of SEQ ID NO: 2. Substitutions of the amino acid that are conservative substitutions that are unlikely to affect biological activity are considered identical for the purposes of this invention and include the following: to be, Val to lie, ASP to Glu, Thr to be, Ala to Gly, Ala to Thr, Ser to Asn, Ala to Val, Ser to Gly, Tyr to Phe, Ala to Pro, Lys to Arg, ASP to Asn , Leu for lie, Leu for Val, Ala for Glu, ASP for Gly, and vice versa, (see, for example, Neurath et al., The Proteins, Academic Press, New York (1979)). Additional information regarding phenotypically silent amino acid exchanges can be found in Bowie et al., 1999, Science 247: 1306-1310. Amino acid substitutions also include substitutions in SEQ ID NO: 2 for amino acids that do not naturally occur, amino acids D, altered amino acids, or peptidomimetics. The amino acid substitutions also include non-conservative amino acid substitutions, such as neutral hydrophobic for polar neutral, acidic for basic, and other class of substitutions, provided that the substituted polypeptides retain the activities of the polypeptides that they have the sequence of the amino acid in SEQ ID NO: 2. The variants further include modifications to the C and N terminus that originate from the process due to expression in various cell types such as mammalian cells, E. coli, yeast and other recombinant host cells. The variants further include polypeptide fragments and polypeptides comprising the inactivated N-glycosylation site (s), inactivated protease processing site (s), or conservative amino acid substitution (s), of the polypeptide sequence set forth in SEC. ID NO: 2. The identity and similarity of the related peptides and polypeptides can be easily calculated by known methods. Such methods include, but are not limited to, those described in Computational Molecular Biology, Lesk, A.M., ed., Oxford University Press, New York (1988); Biocomputing: Informatics and Genome Projects, Smith, D.W., ed., Academic Press, New York (1993); Computer Analysis of Sequence Data, Part 1, Griffin, A.M., and Griffin, H.G., eds. Humana Press, New Jersey (1994); Sequence Analysis in Molecular Biology, von Heinje, G., Academic Press (1987); Sequence Analysis Primer, Gribskov, M. and Devereux, J., eds., M. Stockton Press, New York (1991); and Carillo et al., SIAM J. Applied Math., 48: 1073 (1988). Methods to determine the relevance or identity of the percentage of two polypeptides that are designed to give the greatest equality between the tested sequences.

Preferred computer program methods for determining the identity between two sequences include, but are not limited to, the GCG program package, which includes GAP (Devereux et al., Nucí, Acid Res., 12: 387 (1984)).; Genetics Computer Group, University of Wisconsin, Madison, WI, BLASTP, BLASTN, and FASTA (Altschul et al., J. Mol. Biol., 215: 403-410 (1990).) The BLASTX program that is publicly available from National Center for Biotechnology Information (NCBI) and other sources (BLAST Manual, Altschul et al., NCB / NLM / NIH Bethesda, MD 20894, Altschul et al., Supra (1990).) The well-known Smith Waterman algorithm can also be used to determine the identity.

As used herein the term "derivative" of the ActRIIB5 polypeptides refers to the attachment of at least one additional chemical moiety, or at least one additional polypeptide, to form covalent or aggregated conjugates such as terminal fusion proteins. -C or terminal-N, glycosyl groups, lipids, acetyl groups, conjugation to PEG molecules, and other modifications that are described more fully below. As used herein, the term "an ActRIIB5 polypeptide activity" or "a biological activity of the ActRIIB5 polypeptide" refers to one or more of the in vitro or in vivo activities of the ActRIIB5 polypeptides, including but not limited to those shown in the examples below. The activities of ActRHB5 polypeptides include, but are not limited to, the ability to bind to myostatin or activin A or GDF-II, the ability to reduce or neutralize myostatin or activin A or GDF-11 activity. For example, assay based on the pMARE C2C12 cell described in Example 3 below measures the neutralizing activity of activin A, myostatin neutralizing activity, and GDF-11 neutralizing activity. In vivo activities include but are not limited to the increase in body weight, increase in lean muscle mass, and a decrease in fat mass as demonstrated in the animal models below. Biological activities also include the reduction or prevention of cachexia caused by certain types of tumors, and preventing the metastasis of certain tumor cells. In addition, the discussion of ActRIIB5 polypeptide activities is provided below. The proteins of the present invention further comprise the heterologous proteins linked to the ActRIIB5 polypeptide either directly or through a sequence linker to form a fusion protein. As used herein the term "fusion protein" refers to a protein having a heterologous polypeptide bound via recombinant DNA techniques. Heterologous proteins include but are not limited to Fe polypeptides, their labels, and leucine zipper domains to promote oligomerization and stabilization of the ActRIIB5 polypeptides as described, for example, in the WO 00/29581, which is incorporated herein by reference. As used herein the term "Fe" or "Fe polypeptide" refers to polypeptides that contain the Fe domain of an antibody. The "Fe domain" refers to the portion of the antibody that is responsible for binding to the antibody receptors in the cells. A Fe domain can contain one, two or all of the following: constant heavy domain 1 (CH1), constant heavy domain 2 (CH2), constant heavy domain 3 (CH3), and the region of the joint. The Fe domain of human IgGI, for example, contains the CH2 domain, and the CH3 domain and the joint region, but not the CH1 domain. Truncated forms of such polypeptides that contain the region of the joint that promotes dimerization are also included. See, for example, C. A. Hasemann and J. Donald Capra, Immunoglobins: Structure and Function, in William E. Paul, ed. A Fe is a fully human Fe that can originate from any of the immunoglobulins, such as IgGI and IgG2. However, Fe molecules that are partially human, or that originate from non-human species are also included in the present. The Fe molecules can be composed of monomeric polypeptides which can be linked in dimeric or multimeric forms by covalent (i.e., disulfide linkage) and non-covalent association. The number of intermolecular disulfide bonds between the monomeric subunits of the native Fe molecules have a range of 1 to 4 depending of the class (for example, IgG, IgA, IgE) or subclass (for example, IgGI, IgG2, IgG3, I g Al, IgG2). The term "Fe" as used herein is used to refer to monomeric, dimeric, and multimeric forms. As used herein, the term "Fe variant" refers to a modified form of a native Fe sequence. Fe variants can be constructed, for example, by replacing or removing residues, grafting residues or truncating the portions containing the site. The grafted or substituted residues can also be altered amino acids, such as peptidomimetics or D-amino acids. The proteins of the present invention may also optionally comprise a "linker" group. The linkers serve primarily as a spacer between a polypeptide and a second heterologous protein or other type of fusion or between two or more ActRIIB5 polypeptides. In one embodiment, the linker is composed of amino acids linked together by peptide bonds, preferably from 1 to 20 amino acids linked by peptide bonds, wherein the amino acids are selected from the 20 naturally occurring amino acids. One or more of these amino acids can be glycosylated, as understood by them in the art. In one embodiment, from 1 to 20 amino acids are selected from glycine, alanine, proline, asparagine, glutamine, and Usin. Preferably, a linker is composed of a majority of amino acids without spherical hindrance, such as glycine and to the girl. The exemplary linkers are polyglycines (particularly (Gly) 5, (Gly) 8, poly (Gly-Ala), and polyalanines The linkers of the present invention are also non-peptide linkers, for example, alkyl linkers such as -NH- (CH2) sC ( 0) -, where s = 2-20 can be used.These alkyl linkers can be further substituted by any group with non-steric hindrance such as (for example, C-C6) lower acyl, halogen (e.g., Cl, Br ), CN, NH2, phenyl, etc. The proteins of the present invention can also bind to a molecule without proteins in order to confer desired properties such as reduction of degradation and / or period increase, reduction of toxicity, reduction of immunogeneticity, and / or increase in biological activity of the ActRIIB polypeptides Exemplary molecules include but are not limited to linear polymers such as polyethylene glycol (PEG), polysylysine, dextran, a lipid, a cholesterol group (such as a steroid); a carbohydrate, or a oligosaccharide molecule. In another aspect, the present invention provides isolated nucleic acid molecules comprising polynucleotides that encode the ActRIIB5 polypeptides of the present invention. As used herein the term "isolated" refers to nucleic acid molecules purified to a certain degree from the endogenous material. In one embodiment, the nucleotide acid molecule of the present invention comprises a polynucleotide encoding SEQ ID NO: 2. Due to the known degeneracy of the genetic code, wherein more than one codon can encode the same amino acid, a DNA sequence may vary from that shown in SEQ ID NO: 1, and still encode a polypeptide having the amino acid sequence of SEQ ID NO: 2. Such variable DNA sequences may result from silent mutations that occur during production, or may be the product of the deliberate mutagenesis of SEQ ID NO: 2. In another embodiment, the nucleic acid molecule comprises a polynucleotide encoding a polypeptide having at least about 80% identity to SEQ ID NO: 2, at least about 90% identity to SEQ ID NO: 2 , at least about 95% identity with SEQ ID NO: 2, at least about 99% identity with SEQ ID NO: 2. Percentage identity can be determined by visual inspection and mathematical calculation penthouse. Alternatively, the percent identity of two nucleic acid sequences can be determined by comparing the sequence information using the GAP computer program, version 6.0 described by (Devereux et al., Nucí Acids Res., 12: 387 (1984) ) and available from University of Wisconsin Genetics Computer Group (UWGCG). The preferred pre-set parameters for the GAP program include: (1) a comparison matrix (containing a value of 1 for identities and 0 for non-identifiers) identities) for the nucleotides, and the heavy comparison matrix of (Gribskov and Burgess, Nucí Acids Res., 14: 6745 (1986)), as described by (Schwartz and Dayhoff, eds., Atlas of Protein Sequence and Structure , National Biomedical Research Foundation, pp. 353-358 (1979)); (2) a penalty of 3.0 for each space and an additional penalty of 0.10 for each symbol in each space; and (3) no sanction for extreme spaces. Other programs used by the expert in the sequence comparison technique can also be used. In another embodiment, the nucleic acid molecule of the present invention comprises a polynucleotide having the polynucleotide sequence set forth in SEQ ID NO: 1, or complementary strand of SEQ ID NO: 1. In another embodiment, the present invention provides molecules of nucleic acid which hybridize under stringent or moderate conditions with the regions encoding the polypeptide of SEQ ID NO: 1, wherein the encoded polypeptide comprises a C-terminal amino acid sequence as set forth in SEQ ID NO: 3, and wherein the encoded polypeptide maintains an activity of the ActRIIB5 polypeptides. In another embodiment, the present invention provides the nucleic acid molecules which hybridize under stringent or moderate conditions to the regions encoding the polypeptide of SEQ ID NO: 1, wherein the encoded polypeptide comprises a C-terminal amino acid sequence. which has at least about 80% identity, at least about 85% identity, at least about 90% identity, at least about 95% identity, at least about 98% identity, at least approximately 99% identity with the amino acid sequence set forth in SEQ ID NO: 3, and wherein the encoded polypeptide has at least one activity of the ActRIIB5 polypeptides. As used herein, moderate stringent conditions can be readily determined by those of skill in the art based on, for example, the length of the DNA. The basic conditions are established by (Sambrook et al., Molecular Cloning: A Laboratory Manual, 2ed.Vol. 1, pp. 1,101-104, Cold Spring Harbor Laboratory Press, (1989)), and include the use of a pre-washed solution by the nitrocellulose filters 5X SSC, 0.5% SDS, 1.0 mM EDTA (pH 8.0), hybridization conditions of approximately 50% formamide, 6X SSC at approximately 42 ° C (or other similar hybridization solution, such as the Stark solution, in approximately 50% formamide at approximately 42 ° C), and wash conditions of approximately 60 ° C, 0.5X SSC, 0.1% SDS. High stringency conditions can also be easily determined by an expert based on, for example, the length of the DNA. Generally, such defined conditions as "highly stringent conditions" for hybridization and washing are 0.015 M sodium chloride, 0.0015 M sodium citrate at 65-68 ° C or 0.015 M sodium chloride, 0.0015 M sodium citrate, and 50% formamide at 42 ° C. Other conditions include hybridization and washing at approximately 68 ° C, 0.2X SSC, 0.1% SDS. The skilled person will recognize that the temperature and washing of the concentration of the salt solution can be adjusted as necessary according to factors such as sequence length. See Sambrook, Fritsch & Maniatis, Molecular Cloning: A Laboratory Manual (2nd ed., Cold Spring Harbor Laboratory, 1989); Anderson et al., Nucleic Acid Hybridization: A Practical Approach, Ch.4 (IRL Press Limited). The nucleic acid molecules of the invention include DNA in both forms, single chain and double chain, as well as the complement of RNA. The DNA includes, for example, cDNA, genomic DNA, synthetic DNA, DNA amplified by PCR, and combinations thereof. Genomic DNA can be isolated by conventional techniques, such as using the cDNA of SEQ ID NO: 1, or a convenient fragment thereof, as a test. The genomic DNA encoding the ActRIIB5 polypeptides is obtained from the genomic libraries that are available for a number of species. Synthetic DNA is available from the chemical synthesis that overlaps the oligonucleotide fragments followed by the assembly of the fragments to reconstitute part or all of the coding regions and flanking sequences. RNA can be obtained from the prokaryotic expression vectors which direct the high-level synthesis of mRNA, such as vectors using the T7 promoters and RNA polymerase. The ADCc is obtained from libraries prepared from mRNA isolated from the various tissues expressing ActRIIB5. The DNA molecules of the invention include full length genes as well as polynucleotides and fragments thereof. The full-length gene can also include sequences encoding the N-terminal signal sequence. The invention also provides methods of producing and identifying the ActRIIB5 polynucleotides. The well-known polymerase chain reaction (PCR) method can be used to isolate and amplify a DNA sequence encoding a desired fragment of the protein. The oligonucleotides that define the desired term of the DNA fragment are used as 5 'and 3' primers. The oligonucleotides may additionally contain recognition sites for the restriction endonucleases, to facilitate insertion of the amplified DNA fragment into an expression vector. PCR techniques are described in Saiki et al., Science, 239: 487 (1988); Wu et al., Recombinant DNA Methodology, eds., Academic Press, Inc., San Diego, pp. 189-196 (1989); and Innis et al., PCR Protocols: A Guide to Methods and Applications, eds., Academic Press, Inc. (1990). In another aspect of the present invention, the vectors of expression containing the nucleic acid sequences are also provided, and host cells transformed with such vectors and methods of producing the ActRIIB5 polypeptides are also provided. The term "expression vector" refers to a plasmid, phage, virus or vector for expressing a polypeptide of a polynucleotide sequence. The vectors for the expression of ActRII5 polypeptides contain minimal sequences required for the propagation of the vector and for the expression of the cloned graft. An expression vector comprises a transcriptional unit comprising an assembly (1) of a genetic element or elements that have a regulatory role in gene expression, eg, promoters or enhancers, (2) a sequence encoding the ActRIIB5 polypeptides which are transcribed into mRNA and translated into protein, and (3) appropriate sequences of transcription initiation and termination. These sequences may also include a selection marker. Suitable vectors for expression in host cells are readily available and nucleic acid molecules are grafted into the vectors using recombinant DNA techniques. Such vectors may include promoters that function in specific tissues, and viral vectors for the expression of ActRIIB5 in human or animal labeled cells. A certain expression of exemplary vectors suitable for the expression of ActRIIB5 includes, but is not limited to, pDSRa, (described in the document WO 90/14363, incorporated herein by reference) and its derivatives, containing the polynucleotides ActRIIB5, and vectors pDC323 or pDC324 (described in Bianchi et al., Biotech and Bioengineering, Vol 84 (4): 439-444 (2003)) containing ActRII5 polynucleotides, as well as additional convenient vectors known in the art or described below, are provided by the present invention. The application further provides methods for creating ActRIIB5 polypeptides and proteins. A variety of other expression / host systems can be used. These systems include but are not limited to microorganisms such as bacteria transformed with recombinant bacteriophage, plasmid or cosmid DNA expression vectors; yeast transformed with yeast expression vectors; insect cell systems infected with virus expression vectors (eg, baculovirus); plant cell systems transfected with virus expression vectors (eg, cauliflower mosaic virus, CaMV, tobacco mosaic virus, TMV) or transformed with bacterial expression vectors (eg, Ti or plasmid pBR322); or animal cell systems. Mammalian cells useful in recombinant protein productions include but are not limited to VERO cells, HeLa cells, Chinese hamster ovarian cell lines (CHO), COS cells (such as COS-7), W138, BHK, HepG2, 3T3 cells, RIN, MDCK, A549, PC12, K562 and 293. Host cells mammals may be preferred when they are posttranslational modifications such as glycosylation processing and polypeptide that are important for activity. Mammalian expression allows the production of secreted or soluble polypeptides which can be recovered from the growth medium.

Using an appropriate vector-host system, ActRIIB5 proteins and polypeptides are recombinantly produced by growing a host cell transformed with an expression vector containing the nucleic acid molecules of the present invention under conditions that allow production. The transformed cells can be used for long-term, high-yield protein production. Once such cells are transformed with the vectors containing selectable markers as well as the desired expression band, the cells can be allowed to grow for 1-2 days in an enriched medium before they are switched to selective media. The selectable marker is designed to allow the growth and recovery of cells that successfully express the introduced sequences. Groups of stable transformed cells can be proliferated using tissue culture techniques appropriate to the cell line used. An overview of the expression of recombinant proteins is found in Methods of Enzymology, v. 185, Goeddell, D.V., ed., Academic Press (1990).

In some cases, such as the expression using prokaryotic systems, the expressed polypeptides of this invention may need to be "refolded" and oxidized in an appropriate tertiary structure and the generated disulfide bonds are biologically active. Refolding can be accomplished using a number of procedures well known in the art. Such methods include, for example, exposing the solubilized polypeptide at a pH generally above 7 in the presence of a chaotropic agent. The selection of the chaotrope is similar to the options used for the solubility of the inclusion body, however, a chaotrope is normally used in a lower concentration. Exemplary chaotropic agents are guanidine and urea. In most cases, the refolding / oxidation solution will also contain a reducing agent in addition to its oxidized form in a specific ratio to generate a particular redox potential that allows the disulfide mixture to occur for the formation of cysteine bridges. Some commonly used redox combinations include cysteine / cystamine, glutathione / dithiobisGSH, cupric chloride, dithiothreitol DTT / dithiane DTT, and 2-mercaptoethanol (bME) / dithio-bME. In many cases, a co-solvent can be used to increase the effectiveness of refolding. Commonly used cosolvents include glycerol, polyethylene glycol of various molecular weights, and arginine. The proteins and polypeptides of the present may synthesized in the solution or in a solid support according to conventional techniques. The various automatic synthesizers are commercially available and can be used in accordance with known protocols. See, for example, Stewart and Young (supra); Tarn et al., J Am Chem Soc, 105: 6442, (1983); Errifield, Science 232: 341-347 (1986); Barany and Merrifield, The Peptides, Gross and Meienhofer, eds, Academic Press, New York, 1-284; Barany et al., Int J Pep Protein Res, 30: 705 It is necessary to purify the proteins and polypeptides of the present invention. The purification techniques of the protein are well known to those skilled in the art. These techniques involve, at one level, the crude fractionation of the protein and non-protein fractions. Once the peptide polypeptides are separated from other proteins, the peptide or polypeptide of interest can further be purified using chromatographic and electrophoretic techniques to achieve complete or partial purification (or purification for homogeneity). Suitable analytical methods particularly for the preparation of polypeptides or of the present invention are ion exchange chromatography, exclusion chromatography; polyacrylamide gel electrophoresis; isoelectric focus. A particularly efficient method of peptide purification is rapid liquid chromatography of the protein or even HPLC. The term "isolated polypeptide" or "purified polypeptide" as used herein, it is desired to refer to a composition, isolable from other components, wherein the polypeptide is purified to any degree in relation to its naturally obtainable state. A purified polypeptide therefore also refers to a polypeptide that is free from the environment in which it may occur naturally. Generally, "purified" will refer to a polypeptide composition that has been subjected to fractionation to remove several other components, and that the composition substantially retains its expressed biological activity. Where the term "substantially purified" is used, this designation will refer to a peptide or a polypeptide composition wherein the polypeptide or peptide forms the major component of the composition, such as constituting approximately 50%, approximately 60%, approximately 70% , about 80%, about 90%, about 95% or more of the proteins in the composition. The various methods for quantifying the degree of purification of the polypeptide will be known to those skilled in the art in view of the present disclosure. These include, for example, determining the specific binding activity of an active fraction, or evaluating the amount of peptide or polypeptide within a fraction by SDS / PAGE analysis. A preferred method for evaluating the purity of a polypeptide fraction is to calculate the binding activity of the fraction, to compare it to the binding activity of the initial extract, and thus calculate the degree of purification, evaluated herein by "the number of times of purification". The actual units used represent the amount of binding activity, of course, they will be dependent on the particular assay technique chosen to continue the purification and whether or not the polypeptide or peptide exhibits a detectable binding activity. The various techniques suitable for use in purification will be well known to those skilled in the art. These include, for example, precipitation with ammonium sulfate, PEG, antibodies (immunoprecipitation) and the like or by denaturation of heat, followed by centrifugation; chromatography steps such as affinity chromatography (e.g., Protein-A-Sepharose), ion exchange, gel filtration, reversed phase, hydroxylapatite and affinity chromatography; isoelectric focus; gel electrophoresis; and combinations of these techniques. As is generally known in the art, it is believed that the order of conduction of the various steps of purification can be changed, or that certain steps can be omitted, and still result in a convenient method for the preparation of a substantially purified polypeptide. Antibodies The present invention further includes antibodies that specifically bind to the ActRIIB5 receptor polypeptides of the present invention. As used in this the "specifically linked" term refers to antibodies that have a binding affinity (Ka) for ActRIIB5 polypeptides of 1 O6 M "1 or greater. As used herein, the term" antibody "refers to intact antibodies that include polyclonal antibodies (see, for example, Antibodies: A Laboratory Manual, Harlow and Lane (eds), Cold Spring Harbor Press, (1988)), and monoclonal antibodies (see, for example, US Patent Nos. RE 32,011, 4,902,614, 4,543,439, and 4,411,993, and Monoclonal Antibodies: A New Dimension in Biological Analysis, Plenum Press, Kennett, McKearn and Bechtol (eds.) (1980).) As used herein, the term "antibody" also refers to a fragment of an antibody such as F (ab), F (ab '), F (ab') 2, Fv, Fe, and only chain antibodies that are produced by recombinant DNA techniques or by the enzymatic or chemical division of intact antibodies. The term "antibody" also refers to bispecific or bifunctional antibodies, which are an artificial hybrid antibody that has two different heavy / light chain pairs and two different binding sites. Bispecific antibodies can be produced by a variety of methods including fusion of hybridomas or ligation of Fab fragments. (See Songsivilai et al., Clin. Exp. Immunol., 79: 315-321 (1990), Kostelny et al., J. Immunol., 148: 1547-1553 (1992)). As used herein the term "antibody" also refers to chimeric antibodies, i.e., antibodies having a constant antibody immunoglobulin domain coupled to one or more non-human antibody immunoglobulin variable domains, or fragments thereof (see, for example, U.S. Patent No. 5,595,898 and U.S. Patent No. 5,693,493). The antibodies also refer to "humanized" antibodies (see, for example, US Patent No. 4,816,567 and WO 94/10332), minibodies (WO 94/09817), antibodies, and antibodies produced by transgenic animals, wherein a transgenic animal contains a proportion of the human antibody that produces genes but is deficient in the production of endogenous antibodies are capable of producing human antibodies (see, for example, Méndez et al., Nature Genetics 15: 146-156 (1997), and North American Patent No 6,300,129). The term "antibodies" also includes multimeric antibodies, or a higher order complex of proteins such as heterodimeric antibodies, and anti-idiotypic antibodies. The "antibodies" also include anti-idiotypic antibodies. The antibodies against ActRIIB5 can be used, for example, to identify and quantify ActRIIB5 in vitro and in vivo. Pharmaceutical Compositions Pharmaceutical compositions containing the ActRIIB5 polypeptides and proteins of the present invention are also provided. Such compositions comprise the therapeutically or prophylactically effective amount of polypeptide in the mixture with pharmaceutically acceptable materials, and physiologically acceptable materials of the formulation. The pharmaceutical composition may contain the materials of the formulation to modify, maintain or preserve, for example, pH, osmolarity, viscosity, clarity, color, isotonicity, odor, sterility, stability, rate of dissolution or release, adsorption or penetration of the composition. Suitable formulation materials include, but are not limited to, amino acids (such as glycine, glutamine, asparagine, arginine or Usin); antimicrobials; antioxidants (such as ascorbic acid, sodium sulfite or sodium hydrogen sulfite); buffers (such as borate, bicarbonate, Tris-HCl, citrates, phosphates, other organic acids); volumetric agents (such as mannitol or glycine), chelating agents (such as ethylenediamine tetraacetic acid (EDTA)); complexing agents (such as caffeine, polyvinylpyrrolidone, beta-cyclodextrin or hydroxypropyl-beta-cyclodextrin); fillers; monosaccharides; disaccharides and other carbohydrates (such as glucose, mannose, or dextrins); proteins (such as serum albumin, gelatin or immunoglobulins); Colorant; flavoring, and diluting agents; emulsifying agents; hydrophilic polymers (such as polyvinyl pyrrolidone); low molecular weight polypeptides; counterions that form salt (such as sodium); preservatives (such as benzalkonium chloride, benzoic acid, acid salicylic, thimerosal, phenethyl alcohol, methylparaben, propylparaben, chlorhexidine, sorbic acid or hydrogen peroxide); solvents (such as glycerin, propylene glycol or polyethylene glycol); sugar alcohols (such as mannitol or sorbitol); suspension agents; or wetting agents or surfactants (such as pluronics, PEGs, sorbitan esters, polysorbates such as polysorbate 20, polysorbate 80, triton, tromethamine, lecithin, cholesterol, tyloxapal); stability promoting agents (sucrose or sorbitol); tonicity promoting agents (such as alkali metal halides (preferably sodium or potassium chloride, sorbitol and mannitol), delivery vehicles, diluents, excipients and / or pharmaceutical adjuvants (Remington's Pharmaceutical Sciences, 18th Edition, AR Gennaro, ed. ., Mack Publishing Company, 1990.) The optimal pharmaceutical composition will be determined by one skilled in the art depending on, for example, the intended route of administration, delivery format, and desired dosage, see for example, Remington's Pharmaceutical Sciences, supra. Such compositions may influence the physical state, stability, in vivo release index and polypeptide separation index in vivo For example, suitable compositions may be water for injection, physiological saline for parenteral administration. in a pharmaceutical composition it can be either aqueous or non-aqueous in nature.

For example, a suitable vehicle or carrier can be water for injection, physiological saline or artificial cerebrospinal fluid, possibly supplemented with other common materials in compositions for parenteral administration. The neutral or saline buffered saline mixed with serum albumin are additional exemplary vehicles. Other exemplary pharmaceutical compositions comprise Tris buffer of about pH 7.0-8.5, or acetate buffer of about pH 4.0-5.5, which may also include sorbitol or, therefore, a convenient substitute. In one embodiment of the present invention, the compositions can be prepared for storage by mixing the selected composition having the desired degree of purity with the optionally formulating agents (Remington's Pharmaceutical Sciences, supra) in the form of a lyophilized cake or an aqueous solution . In addition, the therapeutic composition can be formulated as lyophilized using appropriate excipients such as sucrose. The formulations can be supplied in a variety of methods, for example, by inhalation, oral, or injection therapy. When parenteral administration is contemplated, the therapeutic compositions for use in this invention may be in the form of a parenterally pyrogen-free aqueous solution comprising the desired polypeptide in a pharmaceutically acceptable carrier. A particularly convenient vehicle for parenteral injection is distilled water sterile in which a polypeptide is formulated as a sterile, isotonic solution, correctly preserved. Still another preparation may involve the formulation of the desired molecule with an agent, such as injectable microspheres, bioerodible particles, polymeric compounds (polylactic acid, polyglycolic acid), granules, or liposomes, which are provided for controlled or sustained release of the product that can then supplied via a deposit injection. Hyaluronic acid can also be used, and this can have the effect of promoting sustained duration in the circulation. Other convenient means for the introduction of the desired molecule include the implantable drug delivery devices. In another aspect, pharmaceutical formulations suitable for injectable administration can be formulated in aqueous solutions, preferably in physiologically compatible buffers such as the Hanks solution, ringer solution, or physiologically buffered saline. Aqueous injection suspensions may contain substances that increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Additionally, suspensions of the active compounds can be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils, such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate, triglycerides, or liposomes. Non-lipid polycationic amino polymers can also be used for delivery. Optionally, the suspension may also contain stabilizers or convenient agents to increase the solubility of the compounds and allow the preparation of highly concentrated solutions. In another embodiment, a pharmaceutical composition can be formulated for inhalation. The inhalation solutions can also be formulated with a propellant for aerosol delivery. In another embodiment, the solutions can be nebulized. Pulmonary administration is further described in PCT Application No. PCT / US94 / 001875, which describes the pulmonary delivery of chemically modified proteins. It is also contemplated that certain formulations may be administered orally. In one embodiment of the present invention, the molecules that are administered in this manner can be formulated with or without the carriers normally used in the composition of solid dosage forms such as tablets and capsules. For example, a capsule can be designed to release the active portion of the formulation at the point in the gastrointestinal tract when bioavailability is maximized and pre-systemic degradation is minimized. Additional agents can be included to facilitate absorption of the therapeutic molecule. Diluents, flavors, low melting waxes, vegetable oils, lubricants, suspending agents, tablet disintegrating agents, and binders may also used. Pharmaceutical compositions for oral administration can also be formulated using pharmaceutically acceptable carriers well known in the art at convenient doses for oral administration. Such carriers allow the pharmaceutical compositions to be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, mixtures, suspensions, and the like, for ingestion by the patient. Pharmaceutical preparations for oral use can be obtained by combining active compounds with the solid excipient and processing the resulting mixture of granules (optionally, after grinding) to obtain tablets or dragee cores. Suitable auxiliaries can be added, if desired. Suitable excipients include carbohydrate or protein fillers, such as sugars, including lactose, sucrose, mannitol, and sorbitol; corn starch, wheat, rice, potato, or other plants; cellulose, such as methyl cellulose, hydroxypropylmethyl cellulose, or sodium carboxymethylcellulose sodium; gums, including gum arabic and tragacanth; and proteins, such as gelatin and collagen. If desired, the disintegration or solubilization agents may be added, such as cross-linked polyvinyl pyrrolidone, agar, and alginic acid or a salt thereof, such as sodium alginate. The dragee cores can be used in conjunction with convenient coatings, such as solutions of concentrated sugar, which may also contain gum arabic, talc, polyvinylpyrrolidone, carbopol gel, polyethylene glycol, and / or titanium dioxide, lacquer solutions, and suitable organic solvents or solvent mixtures. The dyes or pigments can be added to the tablets or dragee coatings for the identification of the product or to characterize the amount of the active compound, i.e., doses. Pharmaceutical preparations that can be used orally also include hard capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a coating, such as glycerol or sorbitol. The hard capsules may contain active ingredients mixed with fillers or binders, such as lactose or starches, lubricants, such as talc or magnesium stearate, and, optionally, stabilizers. In soft capsules, the active compounds can be dissolved or suspended in convenient liquids, such as fatty oils, liquid, or liquid polyethylene glycol with or without stabilizers. Additional pharmaceutical compositions will be apparent to those skilled in the art, including formulations involving polypeptides in sustained or controlled delivery formulations. Techniques for formulating a variety of other controlled or sustained delivery media, such as liposome carriers, biodrodable microparticles or porous granules and depot injections, are also known to the experts in the art. See, for example, PCT / US93 / 00829 which describes the controlled release of porous polymeric microparticles for the delivery of pharmaceutical compositions. Additional examples of sustained release preparations include semipermeable polymer matrices in the form of formed articles, e.g., films, or microcapsules. Sustained-release matrices may include polyesters, hydrogels, polylactides (US 3,773,919, EP 58,881), L-glutamic acid copolymers and ethyl-L-glutamate range (Sidman et al., Biopolymers, 22: 547-556 (1983), poly (2-hydroxyethyl-methacrylate) (Langer et al., J. Biomed, Mater. Res., 15: 167-277, (1981); Langer et al., Chem. Tech., 12: 98-105 (1982) ), ethylene vinyl acetate (Langer et al., supra) or poly-D (-) - 3-hydroxybutyric acid (EP 133,988). Sustained release compositions also include liposomes, which can be prepared by any of several methods known in the art. See, for example, Eppstein et al., PNAS (USA), 82: 3688 (1985), EP 36,676, EP 88,046, EP 143 949. The pharmaceutical composition that is used for in vivo administration should normally be sterile. by the filtration through the sterile filtration membranes.Where the composition is li Therefore, the sterilization using this method can be conducted either before or after lyophilization and reconstitution. The composition for Parenteral administration can be stored in lyophilized form or in solution. In addition, parenteral compositions are generally placed in a container having a sterile access port, for example, an intravenous solution bag or bottle having a stopper punctured by a hypodermic injection needle. Once the pharmaceutical composition has been formulated, it can be stored in sterile bottles as a solution, suspension, gel, emulsion, solid, or a dehydrated or lyophilized powder. Such formulations can be stored in a ready-to-use form or in a form (eg, lyophilized) which requires reconstitution before administration. In a specific embodiment, the present invention is directed to kits for producing a single-dose administration unit. The kits can each contain both, a first container having a dry protein and a second container having an aqueous formulation. Also included within the scope of this invention are kits containing single and multi-chamber pre-filled syringes (eg syringes and liquid syringes). An effective amount of a pharmaceutical composition that is used therapeutically will depend, for example, on the context and therapeutic objectives. The person skilled in the art will appreciate that the appropriate dose levels for the treatment they will thus vary, depending in part, on the molecule delivered, the indication for what the polypeptide is being used, the route of administration, and the size (body weight, body surface or organ size) and condition (age and general health) ) of the patient. Therefore, the clinician can titrate the dosage and modify the route of administration to obtain the optimal therapeutic effect. A typical dose may range from about 0.1 mg / kg to about 100 mg / kg or more, depending on the factors mentioned above. The polypeptide compositions may preferably be injected or administered intravenously. The long-acting pharmaceutical compositions can be administered every 3 to 4 days, every week, or biweekly depending on the separation index and particular formulation period. The frequency of the dose will depend on the pharmacokinetic parameters of the polypeptide in the formulation used. Normally, a composition is administered until a dose achieves the desired effect. The composition can, therefore, be administered as a single dose, or as a multiple dose (in the same or different concentrations / doses) over time, or as a continuous infusion. Further refinement of the appropriate dose is done routinely. Appropriate doses can be checked through the use of appropriate dose response data. The route of administration of the pharmaceutical composition is in accordance with known methods, for example orally, through injection by intravenous, intraperitoneal, intracerebral (intra-parenchymal), intracerebroventricular, intramuscular, intraocular, intraarterial, intraportal, intralesional, intramedullary, intrathecal, intraventricular, transdermal routes, subcutaneous, or intraperitoneal; as well as intranasal, enteric, topical, sublingual, urethral, vaginal, or rectal media, by sustained release systems or implantation devices. Where desired, the compositions may be administered by bolus injection or continuously by infusion, or by the implantation device. Alternatively or additionally, the composition can be administered locally via implantation of a membrane, sponge, or other suitable material wherein the desired molecule has been absorbed or encapsulated. Where an implantation device is used, the device can be implanted in any convenient tissue or organ, and the delivery of the desired molecule can be the diffusion route, programmed release bolus, or continuous administration. In some cases, the ActRIIB5 polypeptides of the present invention can be delivered by implanting certain cells that have been genetically engineered, using methods such as those described herein, to express and secrete polypeptide. Such cells can be animal or human cells, and can be autologous, heterologous, or xenogeneic. Optionally, the cells can be immortalized. To decrease the chance of a immunological response, the cells can be encapsulated to prevent infiltration of surrounding tissues. The encapsulation materials are semi-permeable, biocompatible polymeric membranes or containers that allow the release of the protein product (s) but prevent the destruction of the cells by the patient's immune system or by other damaging factors of the surrounding tissues. ActRIIB5 gene therapy in vivo is also visualized wherein a nucleic acid molecule encoding ActRIIB5, or a variant or derivative of ActRIIB5 is introduced directly into the subject. For example, a nucleic acid sequence encoding an ActRIIB5 is introduced into labeled cells via the local injection of a nucleic acid construct with or without an appropriate delivery vector, such as an associated adeno-virus vector. Alternative viral vectors include, but are not limited to, retroviruses, adenoviruses, herpes simplex, papilloma virus vectors and viruses. The physical transfer of the virus vector can be achieved in vivo by the local injection of the desired nucleic acid construct or other appropriate delivery vector containing the desired nucleic acid sequence, transfer through the liposome, direct injection (naked DNA), or microparticle bombing (gene gun).

Uses of the Compositions of ActRIIB5 The present invention provides methods and compositions for reducing or neutralizing the amount or activity of myostatin, activin A, or GDF-11 in vivo and in vitro by contacting the proteins with an ActRIIB5 protein. The examples below show that ActRIIB5 proteins have a high affinity for myostatin, activin A, and GDF-11, and are capable of reducing and inhibiting the biological activities of myostatin, activin A and GDF-11. The examples show that ActRIIB5 have a higher activity compared to ActRIIB-ECD as shown by the IC50 values in example 3, and the biological response in animals is higher for the animals ActRIIB5 compared to the animals ActRIIB-ECD as shown in Examples 5 and 6. In one aspect, the present invention provides methods and reagents for treating myostatin-related and / or activin-A related disorders in a subject in need thereof by administering an effective dose of an ActRIIB5 composition to the subject . As used herein, the term "subject" refers to any animal, such as mammals that include humans. In another aspect, the present invention provides the use of the ActRIIB proteins in the preparation of a pharmaceutical composition for the treatment of disorders related to activin A, metabolicps, and muscle wasting listed down. In another aspect, the present invention provides the use of nucleic acids and ActRIIB vectors in the preparation of a pharmaceutical composition for the treatment of the activin A, metabolic, and muscle loss disorders listed below. The compositions of the present invention have been shown with the increase of lean muscle mass as a percentage of body weight and decrease in fat mass as a percentage of body weight in animal models as shown in the examples below. Disorders that can be treated by an ActRIIB5 composition include but are not limited to various forms of muscle loss, as well as metabolic disorders such as diabetes and related disorders, and bone degenerative diseases such as osteoporosis. Disorders from muscle loss include dystrophies such as Duchenne muscular dystrophy, progressive muscular dystrophy, Becker-type muscular dystrophy, Dejerine-Landouzy muscular dystrophy, Erb muscular dystrophy, and infant neuroaxonal muscular dystrophy. Additional muscle loss disorders arise from chronic diseases or disorders such as amyotrophic lateral sclerosis, congestive obstructive pulmonary disease, cancer, AIDS, renal impairment, organ atrophy, androgen deprivation, and rheumatoid arthritis. Overexpression of myostatin may contribute to the cachexia, a syndrome of loss of fat and severe muscle. In one example, serum and intramuscular concentrations of the myostatin immunoreactive protein were found to be increased in men who exhibit AIDS-related muscle loss and were inversely related to the fat-free mass (Gonzalez-Cadavid et al., PNAS USA 95 : 14938-14943 (1998)). Myostatin levels have also been shown to increase in response to burn injuries, resulting in a catabolic effect of the muscle (Lang et al., PHASE B J 15, 1807-1809 (2001)). Additional conditions resulting from muscle loss can result from inactivity due to disability, such as confinement in a wheelchair, prolonged bed rest due to infarction, illness, spinal cord injury, a fracture or bone trauma, and muscle atrophy in a microgravity environment (space flight). For example, the plasma myostatin immunoreactive protein was found to increase after prolonged bed rest (Zachwieja et al, J Gravit Physiol 6 (2): 11 (1999).) It was also found that the muscles of rats exposed to a microgravity environment during a launch flight into space expressed an increasing amount of myostatin compared to the muscles of rats that were not exposed (Lalani et al., LEndocrin 167 (3): 417-28 (2000)).

In addition, age-related increases in muscle-to-fat relationships and age-related muscle atrophy appear to be related to myostatin. For example, the immunoreactive protein of seromyrostatin increased with age in the groups of young (19-35 years of age) intermediate age (36-75 years of age), and older (76-92 years of age) men and women , whereas the average of muscle mass and fat-free mass declined with age in these groups (Yarasheski et al., J Nutr Aging 6 (5): 343-8 (2002)). In addition, myostatin has now been found to be expressed at low levels in the heart muscle and expression is overregulated after cardiomyocytes after infarction (Sharma et al., J Cell Physiol 180 (l): 1-9 (1999 )). Therefore, the reduction of myostatin levels in the heart muscle can improve heart muscle recovery after infarction. Myostatin also occurs by influencing metabolic disorders including type 2 non-insulin dependent diabetes mellitus, hyperglycemia, and obesity. For example, the lack of myostatin has been shown to improve the obese and diabetic phenotypes of two mouse models (Yenes et al., Supra). It has been shown in the examples below that administering the AAV-ActRIIB5 vectors increases muscle relative to fat in an animal, in particular for models of obese animals. Therefore, the composition decreasing composition Fat administering the compositions of the present invention will improve diabetes, obesity, and hyperglycemic conditions in animals. In addition, the examples below and Figure 4B show that compositions containing ActRIIB5 can decrease food intake in obese individuals. In addition, increasing muscle mass by reducing myostatin levels can improve bone strength and reduce osteoporosis and other degenerative bone diseases. It has been found, for example, that mice deficient in myostatin showed an increase in the mineral content and density of the humerus of the mouse and an increase in the mineral content of trabecular and cortical bone in the regions of union of the muscles, as well as an increase in mass muscle (Hamrick et al., Calcif Tissue Int 71 (1): 63-8 (2002)). In addition, the ActlIBR compositions of the present invention can be used to treat the effects of androgen deprivation such as androgen deprivation therapy used for the treatment of prostate cancer. The present invention also provides methods and compositions for increasing muscle mass in fed animals by administering an effective dose of ActRIIB5 proteins to the animal. Since the mature terminal C myostatin polypeptide is identical to all tested species, ActRIIB5 proteins were expected to be effective for muscle mass gain and fat reduction in any species important agricultural including cattle, chicken, turkeys, and pigs. The ActRIIB5 proteins and the compositions of the present invention also antagonize the activity of activin A. An Activin A is known to be expressed in certain types of cancers, particularly gonadal tumors such as ovarian carcinomas, and to cause severe cachexia. (Ciprano et al., Endocrinol 141 (7): 2319-27 (2000), Shou et al., Endocrinol 138 (11): 5000-5 (1997); Coerver et al., Mol Endocrinol 10 (5): 534-43 (1996), Ito et al British J Cancer 82 (8): 1415-20 (2000), Lambert-Messerlian, et al., Gynecologic Oncology 74 91): 93-7 (1999). Example 4 below shows that the expression of activin A in the results of the models of animals in a severe cachexia. The expression of ActRIIB5 / Fc in the animals counteracts that cachexia, as shown in examples 5 and 6. Overexpression of myostatin is also thought to contribute to cachexia, as described above. Therefore, the compositions can be used to treat conditions related to overexpression of activin A, as well as the overexpression of myostatin, such as cachexia of certain cancers and the treatment of certain gonadal-like tumors. The compositions of the present invention can be used alone or in combination with other therapeutic agents to improve their therapeutic effects or to decrease potential side effects. These properties include activity, increased solubility, reduced degradation, increased period, reduced toxicity, and reduced immunogenicity. Thus, the compositions of the present invention are useful for extended treatment regimens. In addition, the hydrophilicity and hydrophobicity properties of the compounds of the invention are well balanced, in such a way that they improve their usefulness for both, in vitro and especially in in vivo uses. Specifically, the compounds of the invention have an appropriate degree of solubility in aqueous media that allows absorption and bioavailability in the body, while also having a degree of lipid solubility that allows the compounds to cross the cell membrane to a site putative action, such as a particular muscle mass. In addition, the ActRIIB5 proteins and the polypeptides of the present invention are useful for detecting and quantifying myostatin, activin A, or GDF-11 in any number of assays. In general, the ActRIIB5 polypeptides of the present invention are useful as capture agents to bind and immobilize myostatin, activin A, or GDF-11 in a variety of assays, similar to those described, for example, in Asai, ed., Methods in Cell Biology, 37, Antibodies in Cell Biology, Academic Press, Inc., New York (1993). The polypeptides can be labeled in a certain way or can react with a third molecule such as an antibody that is labeled to allow myostatin to be detected and quantified. For example, a polypeptide or a third molecule can be modified with a perceptible fraction, such as biotin, which can then be linked by a fourth molecule, such as streptavidin labeled by an enzyme, or other proteins. (Akerstrom, J Immunol 135: 2589 (1985); Chaubert, MOD Pathol 10: 585 (1997)). The invention has been described, the following examples are offered by way of illustration, and without limit. Example I: Isolation of cDNA and expression in cells The human novel activin type IIB receptor cDNA was isolated from a cDNA library of human testicular origin (Clontech, Inc.) according to the following protocol. Primers for the N-terminal and C-terminus of human activin receptor IIB (SEQ ID NO: 4) were generated and PCR was performed using these primers against templates for human cDNA libraries. PCT was performed using the GC-RICH PCR System (Roche, cat # 2140306). Both PCR terminal products N and C P were digested with Pvull / EcoRI and subcloned into the vector pcDNA3.1 -HisA (Invitrogen, Carlsbad, CA.) to make a full-length clone. After ordering a number of PCR products, a cDNA clone from the human cDNA test library was identified as a novel variant binding receptor with the N-terminus. The polynucleotide sequence of this receptor, designated type IIB5 human activin receptor (ActRIIB5). The cDNA clone was missing 152 nucleotide bases that correspond to complete Exon-4 in the type IIB receptor gene of human wild-type activin. The truncation of exon-4 in the variant binding resulted in the elimination of the amino acid sequence crossing the transmembrane region as well as in a frame change leading to a termination of the translator. The amino acid sequence of the variant binding receptor contains the largest extracellular domain, encoded by exons 1, 2 and 3 of the wild-type human activin type IIB receptor, and an additional tail region of 36 amino acids resulting from frame change. The amino acid sequence is set forth in SEQ ID NO: 2. The C-terminal sequence is set forth in SEQ ID NO: 3. Due to the lack of the transmembrane region, ActRIIB5 encodes a soluble form of the receptor type IIB of activin. The transfection of ActRBII5 cDNA into cells led to the expression of secreted, rather than to membrane binding, form of the receptor protein. Example 2: Expression of ActRIIB5 The cDNA encoding ActRIIB5 was cloned into the mammalian pDC323 or pDC324 vectors (Bianchi et al., Biotech and Bioengineering, Vol 84 (4): 439-444 (2003)) and expressed in a 293T cell line. To generate the Fe fusions, the polynucleotides encoding ActRIIB5 (SEQ ID NO: 1) were cloned adjacent to the polynucleotides encoding the sequence linker (Gly) 8 adjacent to the polynucleotides encoding human IgGI Fe in a pDSRa vector ( described in WO / 9014363, incorporated herein by reference). The polynucleotides encoding ActRIIB-ECD (amino acids 1-124 of SEQ ID NO: 5) were cloned adjacent to the polynucleotides encoding human IgGI Fe in a pDSRa vector (no linker). These builders were transfected into a stable CHO cell line. The expressed soluble Fc-Fc fusions were used for the in vitro test side by side described below. For the in vivo animal experiments described in Example 4 below, the PCR products generated as described above were digested with Nhel / Sall and subcloned into an AAV-Fc vector at the same sites. The AAV-Fc vector allows the transfer of the ActRIIB5 gene in an animal for expression in vivo. Example 3: In vitro activities HuActRIIB5 / Fc and H or ActR 11 B-EC D / Fc were generated as described above. The ability of the ActRIIB5 receptor to inhibit the binding of each of the three ligands, myostatin, activin A, and GDF-II to the activin receptor IIB was tested using a cell-based activity assay as described below. Cell-based Activity Assay C2C12 A myostatin / activin / GDF-11 responsive indicator cell line generated by the transfection of myoblast cells C2C12 (ATCC No: CRL-1772) with a pMARE-luc construct. The constructor pMARE-luc was made cloning twelve repeats of the CAGA sequence, representing the myostatin / activin response elements (Dennler et al., EBO 17: 3091-3100 (1998)) in a pLuc-MCS indicator vector (Stratagene cat # 219087) upstream of the TATA box. C2C12 myoblast cells naturally express the myostatin / activin / GDF-11 receptor IIB activin receptor on their cell surface. When myostatin / activin A / GDF-11 binds to cellular receptors, the Smad path is activated, and the phosphorylated Smad binds to the response element (Macias-Silva et al., Cell 87: 1215 (1996)), resulting in expression of the luciferase gene. The luciferase activity is then measured using a commercial luciferase reporter kit (cat # E4550, Promega, Madison, Wl) according to the manufacturer's protocol. A stable line of C2C12 cells that has been transfected with pMARE-luc (C2C12 / pMARE clone # 44) was used to measure the activity according to the following procedure. The indicator cells were plated in 96 wells. Dilutions using screens of each type of soluble indicator were carried out with the fixed concentration in 4 nM of myostatin, activin 20 nM, and 4 nM GDF-11. Myostatin, activin and GDF-II, each were pre-incubated with the soluble receptors in various concentrations. The myostatin / activin / GDF-11 activity was measured by determining the luciferase activity in the treated cultures. IC50 values were determined by each soluble indicator as specified in table 1 below.

TABLE 1 The table above shows that soluble receptors can block myostatin signaling through its receptor but also activin A and GDF-11 signaling. BIAcore® Assay Blocking of the assays was performed using immobilized human ActRIIB-ECD / Fc (R &D Systems, Minneapolis, Mn.) On a CM5 chip (Biacore, Inc., Piscataway, NJ) in the presence and absence of each of the two soluble receptors ActRIIB-ECD / Fc and ActRIIB5 / Fc using the BIAcore® assay system according to the manufacturer's instructions. The signal that binds 100% myostatin was determined in the absence of the receptor in the solution. Several concentrations of the soluble receptors were diluted in sample buffer and incubated with 4 nM myostatin before being injected into the recipient surface. Since only myostatin-free molecules would be able to bind to the chip, a decreased binding response with increased receptor concentration indicated the binding of the receptors to myostatin in the solution. The graph of the binding signal against the concentration of the soluble receptor, ActRIIB-ECD / Fc and ActRIIB5 / Fc was calculated to have an EC50 of approximately 18 nM and 7 nM respectively. The comparison between the two receptors is shown in figure 1. Example 4: Expression of Activin A in C57BI / 6 mice To explore the postnatal role of activin in postnatal animals, activin A was over expressed in mice using the transfer of gene by AAV. Female C57B1 / 6 young adult mice of equal age (5 weeks-old) (Charles River laboratories, Wilmington, Mass) were separated into two balanced weight groups (n = 6 / group), which were subsequently injected via the portal vein with AAV-activin A or the empty vector AAV (control) in IX 10 3 pfu / mouse. The effects on body weight and body composition were analyzed. The AAV-activin A transduces to the group shown a drastic reduction in body weight compared to the control mice transduced with the AAV empty vector. Within 2 weeks of the AAV injection, the group transduced with A activin became severely cachexic since its average weight body was only about 1/2 of the control group transduced by the empty vector. Autopsy revealed that administration of AAV-activin A resulted in dramatic exhaustion by approximately 60% of lean body mass, skeletal muscle mass and fat mass. In addition, mice transduced with activin also showed severe organ loss as indicated by significantly reduced organ weights such as liver and heart. An additional experiment using a reduced amount (I X 1012 pfu / mouse) of the AAV-Activin A virus was performed. The results showed a reduction in body weight and lean body mass resulting from the activin transduction but the effects were less dramatic with respect to the initial experiment using AAV-activin A at a higher dose (I X 10 3 pfu / mouse). This shows that the postnatal cachectic effect of activin A is dose dependent. Example 5: Anabolic effect of AAV-ActRIIB5 in C57B1 / 6 mice Male C57B1 / 6 mice of equal age (5-weeks of age) were divided into 5 groups (n = 10 per group). The AAV viral particles were packaged and titrated before injection as follows: AAV-vacuum, AAV-activin A, AAV-ActRIIB5 / Fc, AAV-ActRI I B-ECD / Fc, and AAV-ProMyo / Fc, where AAV -ProMyo is stopped by the myostatin propeptide. Each one of the above AAV viruses was injected in 8x10 2pfu / mouse except for AAV-activin A, of which a reduced amount of viral particles at 1 x 10 12 pfu / mouse was injected (n = 1 O / group). Viral particles were injected via the portal vein. The body weights were determined every third day. The results are shown in Figure 2. The group of AAV-ActRIIB5 / Fc and the group AAV-ActRI I B-ECD / Fc developed increased body weights compared to the control group of the AA V vector, as well as an increase in body weight compared to the AAV-ProMyo / Fc group. Comparing the two soluble receptor groups, the AAV-ActR 11 B5 / Fc group showed the greatest amount of gain in body weight gained. In contrast, the control group of the AAV vector showed a dramatic decrease in body weights compared to the control group of the AAV vector. Changes in body weight, by post-viral injection in seven weeks, of individual groups were plotted as a percentage of the control group (empty vector group AAV). The AAV-ActRI IB5 / Fc group showed the highest average body weight gain over the control, approximately 25%, compared to the 21% increase in body weight for the ActRIIB-ECD / Fc group. The AAV-ActRI I B5 / Fc group and the AAV-ActRIIB-ECD / Fc group showed greater body weight increases than those produced by the ProMyo / Fc group of approximately 16%. In contrast, the AAV-activin group had a significant fall in body weight by 19%. A comparison of these changes is shown in Figure 3. Lean body mass by post-viral injection of one month, in each group of ten mice was determined using nuclear magnetic resonance (NMR) by measuring the body composition of live mice. At the same time, the body fat content of the mice in each group was determined. The measurements were taken in live mice using the EcoMRI 2003 (Eco Medical Systems, Houston, Tx). EchoMRI 2004 is a complete body composition analyzer that measures the masses of fat and lean tissues in live animals using NMR technology. The average percentage of lean and fat mass as a percentage of body weight for each group of 10 mice is presented in Table 2 below.

TABLE 2 As can be seen from Table 2 above, the AAV-ActRIIB5 / Fc group of mice showed the smallest percentage of body fat, and the largest percentage of lean mass for all groups after one month. These data show that ActRIIB5 / Fc is effective in improving body weight, lean body mass and decreasing fat mass in tested animals. In a related experiment, the five groups of ten mice per group were tested by the absorbing force using a meter from Columbia Instruments, model 1027 dsm (Columbus, Ohio). The results were averaged by each group. The AAV-promyostatin / Fc group average of the absorption strength compared to the AAV-vacuum control mice was approximately 21% for the promiostatin / Fc group, approximately 31% for the ActRI I B-ECD / Fc group approximately 33% for the ActRIIB5 / Fc group of mice. The increase in measured absorbent strength was approximately 46% for the promiostatin / Fc group, approximately 56% for the ActRI I B-ECD / Fc group and approximately 60% for the ActRIIB5 / Fc group. Exe VI: Changes in body weight and composition in obese mice Two groups of Ay Obese mice (Jackson Laboratories, Bar Harbor, Maine) of 11 animals each (8 animals per group at the completion of the experiment) were injected with an empty vector of AAV and an AAV-ActRI I B5 / Fc vector respectively. The viruses were injected in the 8x10 pfu / mouse into the portal vein of each mouse. The mice were then monitored for changes in body weight, food intake, lean muscle mass and fat mass in a period of three months post injection. The food intake was determined by weighing the remaining non-eaten food in the mouse cage on a daily basis and calculating the weekly intake. Lean muscle mass and fat mass were determined by NMR as described above. The results of the experiments are shown in Figures 4 and 5. Figure 4A shows a decrease in body weight and Figure 4B shows a decrease in weekly food intake in the AAV-ActRI I B5 / Fc mice compared to the mice of control. Figure 5A shows increase in lean mass, as determined in NMR by AAV-ActR 11 B5 / Fc, while Figure 5B shows a large decrease in fat mass for the AAV-ActRI IB5 / Fc compared to the control mice , by approximately 50%. At the termination of the experiment, the mice were sacrificed and examined for internal changes. The livers of the treated AAV-ActR 11 B5 / Fc mice were compared with those treated with AAV-vacuum control. Visual inspection of the livers of mice treated with AAV-vacuum and mice treated with AAV-ActRIIB5 / Fc showed that the livers of the AAV-empty control mice contained fatty deposits within the livers, while the mice treated with AAV -ActRIIB5 / Fc were free of fatty deposits. Therefore, the expression of ActRIIB5 / Fc in A mice corrected the fatty livers that characterize the obese mice and, as well as caused a decrease in total body weight, a decrease in the amount of food consumed, an increase in lean muscle mass and great decrease in fat mass. The present invention should not be limited in scope by the specific embodiments described herein, which are intended as illustrations only of the individual aspects of the invention, and functionally equivalent methods and components are within the scope of the invention. In fact, the various modifications of the invention, in addition to those shown and described herein, will become apparent to those skilled in the art from the foregoing description and the accompanying drawings. Such modifications are desired to fall within the scope of the appended claims.

Claims

CLAIMS 1. Isolated nucleic acid molecule comprising a polynucleotide selected from the group consisting of: (a) a polynucleotide having the polynucleotide sequence set forth in SEQ ID NO: 1 or its complement; (b) a polynucleotide encoding a polypeptide consisting of the amino acid sequence set forth in SEQ ID NO: 2; and (c) polynucleotide sequence that hybridizes to either, (a) or (b) under conditions of moderate rigor in approximately 50% formamide, 6X SSC at approximately 42 ° C and washing conditions approximately 60 ° C, 0.5X SSC, 0.1% SDS, and wherein the encoded polypeptide comprises a C-terminus having an amino acid sequence set forth in SEQ ID NO: 3, and wherein the polypeptide is capable of binding to myostatin, activin A, or GDF-II. (d) polynucleotide sequence that hybridizes to either (a) or (b) under moderate stringency conditions in approximately 50% formamide, 6X SSC at approximately 42 ° C and washing conditions of approximately 60 ° C, 0.5X SSC, 0.1% SDS, and wherein the encoded polypeptide comprises a C-terminus having an amino acid sequence of at least 80% identical to SEQ ID NO: 3, and wherein the polypeptide is capable of binding to myostatin, activin A , or GDF-11.
2. Isolated nucleic acid molecule comprising a polynucleotide encoding a polypeptide having an amino acid sequence of at least 80% identical to the amino acid sequence set forth in SEQ ID NO: 2, wherein the polynucleotide comprises a C-terminal having the amino acid sequence set forth in SEQ ID NO: 3. Isolated nucleic acid sequence according to claim 2, wherein the polynucleotide comprises a C-terminal having an amino acid sequence of at least 80% identical to the SEQ ID NO: 3. The isolated nucleic acid molecule according to claim 1, wherein the nucleic acid molecule further comprises polynucleotides that encode at least one heterologous protein in frame with the polynucleotides that encode a receptor of type. Activin IIB5. 5. Isolated nucleic acid molecule according to claim 4, wherein the heterologous protein is a Fe. 6. Isolated nucleic acid molecule according to claim 5, wherein the Fe is linked by a peptide linker. 7. Isolated nucleic acid molecule comprising a polynucleotide consisting of the sequence set forth in SEQ ID NO: 1. 8. Nucleic acid molecule in accordance with any of claims 1 to 7, wherein the polynucleotide is operably linked to a translational or transcriptional regulatory sequence. 9. Nucleic acid molecule according to claim 8, wherein the transcriptional or translational sequence comprises a transcriptional promoter or enhancer. 10. Recombinant vector that directs the expression of the nucleic acid molecule of claim 1. 11. Isolated protein comprising an activin type IIB5 receptor polypeptide, wherein the polypeptide is selected from the group consisting of: (a) a polypeptide which consists of the amino acid sequence set forth in SEQ ID NO: 2; (b) a polypeptide consisting of an amino acid sequence having at least 80% identity to SEQ ID NO: 2, wherein the polypeptide is capable of binding myostatin, activin A or GDF-II; (c) the polypeptide of (b), wherein the C-terminus of the polypeptide comprises the amino acid sequence set forth in SEQ ID NO: 3; and wherein the polypeptide is capable of binding myostatin, activin A, or GDF-II; and (d) polypeptide of (b), wherein the C-terminus of the polypeptide comprises an amino acid sequence having at least 80% identity to SEQ ID NO: 3, and wherein the polypeptide is able to bind myostatin, activin A, or GDF-II. 12. Isolated protein comprising an activin type IIB5 receptor polypeptide, wherein the polypeptide consists of the amino acid sequence set forth in SEQ ID NO: 2. 13. Protein according to claim 12, wherein the amino acid residue 64 in SEQ ID NO: 2 is alanine. 14. Isolated protein comprising a polypeptide encoded by the polynucleotide set forth in SEQ ID NO: 1. 15. A protein according to claim 11, wherein the polypeptide is fused to at least one heterologous polypeptide. 16. Protein according to claim 15, wherein the heterologous protein is a Fe. Polypeptide (17. Protein according to claim 16, wherein the Fe polypeptide is linked via a sequence linker. 18. Host cell engineered to express the nucleic acid molecule of claim 1. 19. Host cell according to claim 18, wherein the host cell is a mammalian cell. 20. Host cell engineered to produce the protein of claim 11. 21. Host cell according to claim 20, wherein the host cell is a cell mammal 22. Method of producing an activin IIB5 receptor polypeptide comprising the host cell culture of claim 21, which promotes expression of the polypeptide, and recovery of the polypeptide. 23. Pharmaceutical composition comprising the activin type IIB5 receptor protein of claim 11, in admixture with a pharmaceutically acceptable carrier. 24. Method of inhibiting myostatin activity in a subject in need thereof comprising administering a therapeutically effective amount of the composition of claim 23 to the subject. 25. Method of increasing lean muscle mass in a subject in need thereof comprising administering a therapeutically effective amount of the composition of claim 23 to the subject. 26. Method of increasing the ratio of lean muscle mass to fat in a subject in need thereof comprising administering a therapeutically effective amount of the composition of claim 23 to the subject. 27. Method of treating a muscle wasting disease muscle in a subject suffering from such a disease comprising administering a therapeutically effective amount of the composition of claim 23 to the subject. 28. Method according to claim 27, in where the disease is cachexia of cancer. 29. Method according to claim 27, wherein the selected disease is muscular dystrophy, amyotrophic lateral sclerosis, congestive obstructive pulmonary disease, chronic cardiac arrest, cancer cachexia, AIDS, renal deficiency, uremia, rheumatoid arthritis, sarcopenia related to the age, organ atrophy, carpal tunnel syndrome, androgen deficiency, and muscle loss due to prolonged bed rest, spinal cord injury, myocardial infarction, bone fracture, and aging. 30. Method of treating a metabolic disorder in a subject in need thereof comprising administering a therapeutically effective amount of the composition of claim 23 to the subject. 31. The method of claim 30, wherein the metabolic disorder is selected from diabetes, obesity, hyperglycemia, and bone loss. 32. Method of treating a disease in which activin is over expressed in a subject comprising administering a therapeutically effective amount of the composition of claim 23 to the subject. 33. The method of claim 30, wherein the disease is cancer. 34. The method of claim 25, wherein the subject is a fed animal. 35. Method of treating a muscle loss disorder comprising administering the vector of claim 10, to a subject, wherein the vector is capable of directing the expression of the ActRIIB5 polypeptides in the subject. 36. Method according to claim 35, wherein the vector is an AAV vector.