TGF-Beta receptor type II fusion protein and its use

Abstract

The present invention provides compositions and methods for modulating TGFβ superfamily signaling. [Solution] In some embodiments, the present disclosure provides a transforming growth factor β-receptor II (TβRII) fusion polypeptide comprising a) an extracellular domain of a TβRII portion, b) a heterologous portion, and c) a linker portion having a linker of at least 10 amino acids in length, wherein the TβRII extracellular domain portion comprises an amino acid sequence that is at least 80% identical to i) a sequence beginning at any position 23 to 35 of a particular sequence and ending at any position 153 to 159, or ii) a sequence beginning at any position 23 to 60 of a different particular sequence and ending at any position 178 to 184.

Description

[Technical Field]

[0001] Cross-references to related applications This application is U.S. Provisional Application No. 62 / 501,229, filed on May 4, 2017. From; from U.S. Provisional Application No. 62 / 510,422, filed on 24 May 2017 ; and U.S. Provisional Application No. 62 / 578,674 filed on October 30, 2017 They claim the benefit of priority from the aforementioned applications. The aforementioned applications, in their entirety, are referenced herein. It will be used in the context of the project. [Background technology]

[0002] Background of the Invention Members of the transforming growth factor beta (TGFβ) superfamily include: Proliferation, differentiation, apoptosis, motility, extracellular matrix production, tissue remodeling, angiogenesis, immunology It is a multifunctional cytokine involved in essential cellular functions such as disease response and cell adhesion, and in chronic inflammation. It also plays an important role in the pathophysiology of disease conditions that differ as much as cancer. The family members are TGFβ, bone morphogenetic protein (BMP). Genetic protein, osteogenic protein (OP) Protein), growth and differentiation factor (GDF: growth and different Activation factor, inhibin / activin, Müllerian duct inhibitor (MIS) (Mullerian inhibitory substance) and glial cells Glyal-derived neurotrophic factor (GDNF) It is classified into major family groups, including factor.

[0003] TGFβ superfamily members cross the plasma membrane by inducing the formation of heteromeric complexes of specific type I and type II serine / threonine kinase receptors, transmit those signals, and also, through this transmission, activate a specific subset of SMAD proteins (some inhibitory and some excitatory). SMAD molecules relay the signal into the nucleus where they coordinate with other proteins to direct transcriptional responses. Dysfunctional TGFβ superfamily signaling has been associated with several clinical disorders, including cancer, fibrosis, bone disease, diabetic

[0004] nephropathy, and chronic vascular diseases such as atherosclerosis.

[0005] Therefore, an object of the present disclosure is to provide compositions and methods for modulating TGFβ superfamily signaling. SUMMARY OF THE INVENTION

[0006] MEANS FOR SOLVING THE PROBLEM GIST OF THE INVENTION In part, the present disclosure provides a TβRII polypeptide fusion protein and the use of such a fusion protein as a selective antagonist of TGFβ 1 or TGFβ3. As described herein, polypeptides that include some or all of the extracellular domain (ECD) of TβRII, with or without additional mutations, are variously affinity and bind to TGFβ1 or TGFβ3 and / or inhibit TGFβ1 or T GFβ3. In particular, polypeptides that include a heterologous moiety (e.g., an Fc immunoglobulin domain) and at least Linkers with a length of 10 amino acids (for example, linkers having the amino acid sequence of SEQ ID NO: 6) TβRII polypeptides containing (-) have shorter linkers. It exhibits remarkably superior TGFβ1 and TGFβ3 binding properties compared to tide. Therefore In certain aspects, this disclosure relates to the selective inhibition of TGFβ superfamily-related disorders. We provide a TβRII polypeptide for use.

[0007] In some embodiments, this disclosure relates to transforming growth factor β receptor II (TβR II) A fusion polypeptide comprising a) an extracellular domain of the TβRII portion and b) a heterogeneous portion. The Tβ i) The RII extracellular domain portion starts at any of positions 23-35 of Sequence ID No. 1, ii) A sequence ending at any of positions 153-159 of sequence number 1, or ii) Sequences 23-6 of sequence number 2 A sequence that starts at position 0 and ends at position 178-184 of sequence number 2, and a small number of others. The present invention provides a polypeptide containing at least 80% identical amino acid sequences.

[0008] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-35 of SEQ ID NO: 1 A sequence that begins with one of the following and ends at one of the positions 153-159 of sequence number 1, and at least They also contain amino acid sequences that are 80% identical.

[0009] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-35 of SEQ ID NO: 1 A sequence that begins with one of the following and ends at one of the positions 153-159 of sequence number 1, and at least They also contain amino acid sequences that are 90% identical.

[0010] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-35 of SEQ ID NO: 1 A sequence that begins with one of the following and ends at one of the positions 153-159 of sequence number 1, and at least They also contain amino acid sequences that are 95% identical.

[0011] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-35 of SEQ ID NO: 1 A sequence that begins with one of the following and ends at one of the positions 153-159 of sequence number 1, and at least It also contains amino acid sequences that are 97% identical.

[0012] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-35 of SEQ ID NO: 1 Contains an amino acid sequence that begins with one of the following and ends at any of the positions 153-159 of sequence number 1. nothing.

[0013] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-60 of SEQ ID NO: 2 A sequence that starts with one of the following and ends at one of the positions 178-184 of sequence number 2, and at least They also contain amino acid sequences that are 80% identical.

[0014] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-60 of SEQ ID NO: 2 A sequence that starts with one of the following and ends at one of the positions 178-184 of sequence number 2, and at least They also contain amino acid sequences that are 90% identical.

[0015] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-60 of SEQ ID NO: 2 A sequence that starts with one of the following and ends at one of the positions 178-184 of sequence number 2, and at least They also contain amino acid sequences that are 95% identical.

[0016] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-60 of SEQ ID NO: 2 A sequence that starts with one of the following and ends at one of the positions 178-184 of sequence number 2, and at least It also contains amino acid sequences that are 97% identical.

[0017] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-60 of SEQ ID NO: 2 Contains an amino acid sequence that begins with one of the following and ends at any of the positions 178-184 of sequence number 2. nothing.

[0018] In some embodiments, the TβRII extracellular domain portion is sequence number 18 and at least 8 Contains 0% identical amino acid sequences.

[0019] In some embodiments, the TβRII extracellular domain portion is sequence number 18 and at least 9 Contains 0% identical amino acid sequences.

[0020] In some embodiments, the TβRII extracellular domain portion is sequence number 18 and at least 9 Contains 5% identical amino acid sequences.

[0021] In some embodiments, the TβRII extracellular domain portion is sequence number 18 and at least 9 Contains 7% identical amino acid sequences.

[0022] In some embodiments, the extracellular domain portion of TβRII is the amino acid sequence of SEQ ID NO: 18. Includes.

[0023] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-35 of SEQ ID NO: 1 A sequence that begins with one of the following and ends at one of the positions 153-159 of sequence number 1, and at least They also consist of amino acid sequences that are 80% identical.

[0024] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-35 of SEQ ID NO: 1 A sequence that begins with one of the following and ends at one of the positions 153-159 of sequence number 1, and at least They also consist of amino acid sequences that are 90% identical.

[0025] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-35 of SEQ ID NO: 1 A sequence that begins with one of the following and ends at one of the positions 153-159 of sequence number 1, and at least They also consist of amino acid sequences that are 95% identical.

[0026] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-35 of SEQ ID NO: 1 A sequence that begins with one of the following and ends at one of the positions 153-159 of sequence number 1, and at least They also consist of amino acid sequences that are 97% identical.

[0027] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-35 of SEQ ID NO: 1 Starting with one of the amino acids and ending at any of the positions 153-159 of sequence number 1, Yes.

[0028] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-60 of SEQ ID NO: 2 A sequence that starts with one of the following and ends at one of the positions 178-184 of sequence number 2, and at least They also consist of amino acid sequences that are 80% identical.

[0029] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-60 of SEQ ID NO: 2 A sequence that starts with one of the following and ends at one of the positions 178-184 of sequence number 2, and at least They also consist of amino acid sequences that are 90% identical.

[0030] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-60 of SEQ ID NO: 2 A sequence that starts with one of the following and ends at one of the positions 178-184 of sequence number 2, and at least They also consist of amino acid sequences that are 95% identical.

[0031] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-60 of SEQ ID NO: 2 A sequence that starts with one of the following and ends at one of the positions 178-184 of sequence number 2, and at least They also consist of amino acid sequences that are 97% identical.

[0032] In some embodiments, the extracellular domain portion of TβRII is located at positions 23-60 of SEQ ID NO: 2 Starting with any of the amino acids and ending at any of the positions 178-184 in SEQ ID NO: 2 Yes.

[0033] In some embodiments, the TβRII extracellular domain portion is sequence number 18 and at least 8 They consist of 0% identical amino acid sequences.

[0034] In some embodiments, the TβRII extracellular domain portion is sequence number 18 and at least 9 They consist of 0% identical amino acid sequences.

[0035] In some embodiments, the TβRII extracellular domain portion is sequence number 18 and at least 9 They consist of 5% identical amino acid sequences.

[0036] In some embodiments, the TβRII extracellular domain portion is sequence number 18 and at least 9 They consist of 7% identical amino acid sequences.

[0037] In some embodiments, the extracellular domain portion of TβRII is the amino acid sequence of SEQ ID NO: 18. It consists of.

[0038] In some embodiments, the polypeptide includes an N-terminal leader sequence.

[0039] In some embodiments, the N-terminal leader sequence is one of the sequences 22-24. Contains a mino acid sequence.

[0040] In some embodiments, the N-terminal leader sequence includes the amino acid sequence of SEQ ID NO: 23.

[0041] In some embodiments, the heterogeneous portion is an immunoglobulin Fc domain.

[0042] In some embodiments, the immunoglobulin Fc domain is a human immunoglobulin Fc domain. It is.

[0043] In some embodiments, the heterogeneous portion is an amino acid that is at least 80% identical to SEQ ID NO: 20. Contains acid sequences.

[0044] In some embodiments, the heterogeneous portion is an amino acid that is at least 90% identical to SEQ ID NO: 20. Contains acid sequences.

[0045] In some embodiments, the linker is less than 25 amino acids long.

[0046] In some embodiments, the linker is between 10 and 25 amino acids in length.

[0047] In some embodiments, the linker is between 15 and 25 amino acids in length.

[0048] In some embodiments, the linker is between 17 and 22 amino acids in length.

[0049] In some embodiments, the linker is 21 amino acids long.

[0050] In some embodiments, the heterogeneous portion is an amino acid that is at least 95% identical to SEQ ID NO: 20. Contains acid sequences.

[0051] In some embodiments, the heterogeneous portion is an amino acid that is at least 97% identical to SEQ ID NO: 20. Contains acid sequences.

[0052] In some embodiments, the heterogeneous portion includes the amino acid sequence of SEQ ID NO: 20.

[0053] In some embodiments, the linker includes (GGGGS)n (where n = ≥ 2).

[0054] In some embodiments, the linker includes (GGGGS)n (where n = ≥ 3).

[0055] In some embodiments, the linker includes (GGGGS)n (where n = ≥ 4).

[0056] In some embodiments, the linker is (GGGGS) n (The formula includes n≠≧5.)

[0057] In some embodiments, the linker includes the amino acid sequence of SEQ ID NO: 21.

[0058] In some embodiments, the linker contains one of the amino acid sequences of SEQ ID NOs: 4-7. nothing.

[0059] In some embodiments, the linker includes the amino acid sequence of SEQ ID NO: 6.

[0060] In some embodiments, the polypeptide has at least 80 amino acids from the amino acid sequence of SEQ ID NO: 11. Contains amino acid sequences that are % identical.

[0061] In some embodiments, the polypeptide has at least 90 amino acids from the amino acid sequence of SEQ ID NO: 11. Contains amino acid sequences that are % identical.

[0062] In some embodiments, the polypeptide has at least 95 amino acids from the sequence of SEQ ID NO: 11. Contains amino acid sequences that are % identical.

[0063] In some embodiments, the polypeptide comprises the amino acid sequence of SEQ ID NO: 11.

[0064] In some embodiments, the polypeptide has at least 80 amino acids from the amino acid sequence of SEQ ID NO: 13. Contains amino acid sequences that are % identical.

[0065] In some embodiments, the polypeptide has at least 90 amino acids from the amino acid sequence of SEQ ID NO: 13. Contains amino acid sequences that are % identical.

[0066] In some embodiments, the polypeptide has at least 95 amino acids from the amino acid sequence of SEQ ID NO: 13. Contains amino acid sequences that are % identical.

[0067] In some embodiments, the polypeptide comprises the amino acid sequence of SEQ ID NO: 13.

[0068] In some embodiments, the polypeptide has the amino acid sequence of SEQ ID NO: 50 and at least 80 Contains amino acid sequences that are % identical.

[0069] In some embodiments, the polypeptide has at least 90 amino acids in the amino acid sequence of SEQ ID NO: 50. Contains amino acid sequences that are % identical.

[0070] In some embodiments, the polypeptide has at least 95 amino acids from the sequence of SEQ ID NO: 50. Contains amino acid sequences that are % identical.

[0071] In some embodiments, the polypeptide includes the amino acid sequence of SEQ ID NO: 50.

[0072] In some embodiments, the polypeptide has at least 80 amino acids from the amino acid sequence of SEQ ID NO: 51. Contains amino acid sequences that are % identical.

[0073] In some embodiments, the polypeptide has at least 90 amino acids from the amino acid sequence of SEQ ID NO: 51. Contains amino acid sequences that are % identical.

[0074] In some embodiments, the polypeptide has at least 95 amino acids from the amino acid sequence of SEQ ID NO: 51. Contains amino acid sequences that are % identical.

[0075] In some embodiments, the polypeptide includes the amino acid sequence of SEQ ID NO: 51.

[0076] In some embodiments, the polypeptide has the amino acid sequence of SEQ ID NO: 53 and at least 80 Contains amino acid sequences that are % identical.

[0077] In some embodiments, the polypeptide has at least 90 amino acids from the amino acid sequence of SEQ ID NO: 53. Contains amino acid sequences that are % identical.

[0078] In some embodiments, the polypeptide has at least 95 amino acids from the amino acid sequence of SEQ ID NO: 53. Contains amino acid sequences that are % identical.

[0079] In some embodiments, the polypeptide includes the amino acid sequence of SEQ ID NO: 53.

[0080] In some embodiments, the polypeptide has at least 80 amino acids from the amino acid sequence of SEQ ID NO: 56. Contains amino acid sequences that are % identical.

[0081] In some embodiments, the polypeptide has at least 90 amino acids from the amino acid sequence of SEQ ID NO: 56. Contains amino acid sequences that are % identical.

[0082] In some embodiments, the polypeptide has at least 95 amino acids from the sequence of SEQ ID NO: 56. Contains amino acid sequences that are % identical.

[0083] In some embodiments, the polypeptide includes the amino acid sequence of SEQ ID NO: 56.

[0084] In some embodiments, the polypeptide has at least 80 amino acids from the amino acid sequence of SEQ ID NO: 15. Contains amino acid sequences that are % identical.

[0085] In some embodiments, the polypeptide has at least 90 amino acids from the amino acid sequence of SEQ ID NO: 15. Contains amino acid sequences that are % identical.

[0086] In some embodiments, the polypeptide has at least 95 amino acids from the amino acid sequence of SEQ ID NO: 15. Contains amino acid sequences that are % identical.

[0087] In some embodiments, the polypeptide comprises the amino acid sequence of SEQ ID NO: 15.

[0088] In some embodiments, the TβRII polypeptide is amino acids 185-59 of SEQ ID NO: 2 Does not include 2.

[0089] In some embodiments, the TβRII polypeptide contains amino acids 1-22 of SEQ ID NO: 2 No.

[0090] In some embodiments, the polypeptide has a) the amino acid sequence of SEQ ID NO: 18 and at least It contains amino acid sequences that are 85%, 90%, 95%, 97%, or 99% identical, and 10, 9 , containing 8, 7, 6, 5, 4, 3, 2 or 1 or fewer additional amino acids, TβRII poly The peptide portion and b) the amino acid sequence of SEQ ID NO: 6, at least 85%, 90%, 95%, Contains amino acid sequences that are 97% or 99% identical, with 5, 4, 3, 2, or 1 or fewer additional amino acids. The linker portion contains additional amino acids, and c) the amino acid sequence of SEQ ID NO: 20 and at least 8 Contains amino acid sequences that are 5%, 90%, 95%, 97%, or 99% identical, and 25, 20 , a heterogeneous portion containing 15, 10, 5, 4, 3, 2 or 1 or fewer additional amino acids, d ) Consists of, if necessary, a leader array (e.g., sequence number 23) and, or derived from these. It becomes qualitative.

[0091] In some embodiments, the polypeptide comprises a) the amino acid sequence of SEQ ID NO: 18, and 10 , containing 9, 8, 7, 6, 5, 4, 3, 2 or 1 or fewer additional amino acids, TβRII The polypeptide portion and b) containing the amino acid sequence of SEQ ID NO: 6, in quantities of 5, 4, 3, 2, or 1. The linker portion includes the following additional amino acids, and c) the amino acid sequence of SEQ ID NO: 20 , containing 25, 20, 15, 10, 5, 4, 3, 2 or 1 or fewer additional amino acids, It consists of a seed portion and, if necessary, a leader sequence (e.g., sequence number 23), or These are the essential components.

[0092] In some embodiments, the polypeptide is a) the extracellular domain of the TβRII portion. , and is at least 85%, 90%, 95%, 97%, or 99% identical to the sequence of sequence number 18. an extracellular domain containing a certain amino acid sequence, and b) the sequence of SEQ ID NO: 20 and at least 85% , heterogeneous portions containing amino acid sequences that are 90%, 95%, 97%, or 99% identical, and c) The linker is at least 85%, 90%, 95%, and 97% of the amino acid sequence of SEQ ID NO: 6 A linker that connects an extracellular domain and a heterogeneous region, containing amino acid sequences that are 99% identical. —Includes the part.

[0093] In some embodiments, the polypeptide is a) the extracellular domain of the TβRII portion. a) an extracellular domain containing the amino acid sequence of SEQ ID NO: 18, and b) the amino acid sequence of SEQ ID NO: 20 A heterogeneous portion containing a column, and an extracellular domain in which the linker contains the amino acid sequence of SEQ ID NO: 6. It includes a linker section that connects different parts.

[0094] In some embodiments, the polypeptide has at least 85 amino acids from the amino acid sequence of SEQ ID NO: 48. Contains amino acid sequences that are identical by %, 90%, 95%, 97%, or 99%.

[0095] In some embodiments, the polypeptide includes the amino acid sequence of SEQ ID NO: 48.

[0096] In some embodiments, the polypeptide does not contain a leader sequence, or the leader sequence It was removed.

[0097] In some embodiments, polypeptides are glycosylated amino acids, pegylated amino acids, and Renesylated amino acids, acetylated amino acids, biotinylated amino acids, and condylated amino acids in the lipid substructure. Select from conjugated amino acids and amino acids conjugated with organic derivatizers. It contains one or more modified amino acid residues that are selected.

[0098] In some embodiments, the polypeptide is glycosylated.

[0099] In some embodiments, the polypeptide is specific to polypeptide expression in CHO cells. It has a glycosylation pattern.

[0100] In some embodiments, polypeptides are human with an equilibrium dissociation constant (KD) of less than 100 pM. Binds to TGFβ1.

[0101] In some embodiments, the polypeptide is human T with an equilibrium dissociation constant (KD) of less than 75 pM. It binds to GFβ1.

[0102] In some embodiments, the polypeptide is human T with an equilibrium dissociation constant (KD) of less than 60 pM. It binds to GFβ3.

[0103] In some embodiments, the polypeptide is human T with an equilibrium dissociation constant (KD) of less than 50 pM. It binds to GFβ3.

[0104] In some embodiments, polypeptides are determined using a reporter gene assay. It inhibits TGFβ1 with an IC50 of less than 1.0 nM.

[0105] In some embodiments, polypeptides are determined using a reporter gene assay. It inhibits TGFβ1 with an IC50 of less than 0.25 nM.

[0106] In some embodiments, polypeptides are determined using a reporter gene assay. It inhibits TGFβ1 with an IC50 of less than 0.1 nM.

[0107] In some embodiments, polypeptides are determined using a reporter gene assay. It inhibits TGFβ1 with an IC50 of less than 0.05 nM.

[0108] In some embodiments, polypeptides are determined using a reporter gene assay. It inhibits TGFβ3 with an IC50 of less than 0.3 nM.

[0109] In some embodiments, polypeptides are determined using a reporter gene assay. It inhibits TGFβ3 with an IC50 of less than 0.1 nM.

[0110] In some embodiments, polypeptides are determined using a reporter gene assay. It inhibits TGFβ3 with an IC50 of less than 0.05 nM.

[0111] In some embodiments, polypeptides are determined using a reporter gene assay. It inhibits TGFβ3 with an IC50 of less than 0.04 nM.

[0112] In some embodiments, the reporter gene assay is a CAGA reporter assay. ru.

[0113] In some embodiments, this disclosure uses any two of the polypeptides disclosed herein. This provides homodimers containing the compound.

[0114] In some embodiments, this disclosure relates to any of the polypeptides disclosed herein. Provides isolated polynucleotides containing the coding sequence. In some embodiments, This disclosure is operably linked to any of the polynucleotides disclosed herein. The present invention provides recombinant polynucleotides containing a promoter sequence. In some embodiments, Polynucleotides are at least 80% of one of sequence numbers 10, 12, or 14. , containing nucleotide sequences that are 85%, 90%, 95%, 97%, or 100% identical.

[0115] In some embodiments, this disclosure is made using any of the polynucleotides disclosed herein. The transformed cells are provided. In some embodiments, the cells are mammalian cells. In this embodiment, the cells are CHO cells or human cells.

[0116] In some embodiments, this disclosure includes pharmaceutically acceptable excipients and the information disclosed herein. A medical We provide medicines.

[0117] In some embodiments, this disclosure relates to cells against TGFβ superfamily members. A method for modulating a response, wherein any of the polypeptides disclosed herein The invention provides a method comprising exposing cells to one of the homodimers disclosed herein. In some embodiments, this disclosure describes how TGFβ superintensifies in patients who require it. Methods for treating diseases or conditions related to family members, as disclosed herein. Effective of any polypeptide or any homodimer disclosed herein The present invention provides a method comprising administering a certain amount to the patient. In some embodiments, TGFβ- The family members are TGFβ1 or TGFβ3. In some embodiments, The disease or condition is cancer. In some embodiments, cancer can be stomach cancer, intestinal cancer, skin cancer, etc. The cancers selected are breast cancer, melanoma, bone cancer, and thyroid cancer. In some embodiments, The disease or condition is a fibrous or sclerotic disease or condition. In some embodiments, Fibrous or sclerotic diseases or conditions include scleroderma, lupus erythematosus, pulmonary fibrosis, and atherosclerosis. Arteriosclerosis, hepatic fibrosis, diffuse systemic sclerosis, glomerulonephritis, nerve scarring, dermal scarring, The following are selected from radiation-induced fibrosis, hepatic fibrosis, idiopathic pulmonary fibrosis, and myelofibrosis. In some embodiments, the disease or condition is myelofibrosis. In some embodiments, the disease or The conditions include primary myelofibrosis, myelofibrosis after polycythemia euna, and essential thrombocythemia. Selected from the group consisting of later myelofibrosis. In some embodiments, the disease or condition is national According to the International Prognostic Prediction Scoring System (IPSS), or Dynamic IPSS (DIPSS). According to S), low-risk, intermediate-1 risk, intermediate-2 risk, or high-risk myelofibrosis Selected from the group consisting of. In some embodiments, the disease or condition is a heart disease. In some embodiments, the disease or condition is hereditary hemorrhagic telangiectasia (HHT), Marf Ahn syndrome, Loeys-Dietz syndrome, familial thoracic aortic aneurysm syndrome, arterial tortuosity syndrome, child Choose from preeclampsia, atherosclerosis, restenosis, and hypertrophic cardiomyopathy / congestive heart failure. In some embodiments, the disease or condition is pulmonary hypertension. Pulmonary hypertension is classified as Class I, Class II, or Class II by the World Health Organization. It is Class I or Class IV pulmonary hypertension. In some embodiments, the disease or condition involves the kidneys. It is a related disease or condition. In some embodiments, a kidney-related disease or condition is chronic kidney disease. Kidney disease (or failure), acute kidney disease (or failure), primary kidney disease, non-diabetic kidney disease Glomerulonephritis, interstitial nephritis, diabetic kidney disease, diabetic nephropathy, glomerulosclerosis, rapidly progressing Glomerulonephritis, renal fibrosis, Alport syndrome, IDDM nephritis, mesangial proliferative glomerulonephritis Nephritis, membranoproliferative glomerulonephritis, crescentic glomerulonephritis, interstitial renal fibrosis, focal segmental glomerulonephritis Spherosclerosis, membranous nephropathy, minimal change nephrotic syndrome, microimmune rapidly progressive glomerulonephritis IgA nephropathy, polycystic kidney disease, Dent's disease, nephrocytinosis, Hyman Nephritis, autosomal dominant (adult) polycystic kidney disease, autosomal recessive (child) polycystic kidney disease, acute kidney disease Organ damage, nephrotic syndrome, renal ischemia, glomerular epithelial cell disease or disorder, proteinuria, glomerular disease Diseases, membranous glomerulonephritis, focal segmental glomerulonephritis, pre-eclampsia, eclampsia, renal disorders, collagen disease vascular disorders Diseases, benign orthostatic (postural) proteinuria, IgM nephropathy, membranous nephropathy, sarcoidosis, glucose vera Diabetes, drug-induced kidney damage, Fabry disease, aminoaciduria, Fanconi syndrome, hypertension Stress nephrosclerosis, interstitial nephritis, sickle cell anemia, hemoglobinuria, myoglobinuria, we Gener granulomatosis, glycogen storage disease type 1, chronic kidney disease, chronic renal failure, low glomerular filtration rate (GFR), Renal vascular sclerosis, lupus nephritis, ANCA-positive microimmune crescentic glomerulonephritis, chronic glomerulonephritis Seed transplant nephropathy, nephrotoxicity, kidney necrosis, kidney injury, glomerular and tubular injury, renal function Renal failure, nephritis syndrome, acute renal failure, chronic renal failure, proximal fallopian tube dysfunction, acute kidney transplant rejection Chronic kidney transplant rejection, non-IgA mesangial proliferative glomerulonephritis, post-infectious glomerulonephritis, Vasculitis associated with all types of kidney disorders, all hereditary kidney diseases, all types of interstitial nephritis, renal translocation Related to kidney failure, kidney cancer, and other conditions (e.g., hypertension, diabetes, and autoimmune diseases) Kidney disease, Dent's disease, nephrosis, Hymann's nephritis, primary kidney disease, collapsed glomerulostomy, Dense deposit disease, cryoglobulinemia-associated glomerulonephritis, Noch-Schönlein disease Post-infectious glomerulonephritis, bacterial endocarditis, microscopic polyangiitis, Churg-Strauss syndrome Group, anti-GBM antibody-mediated glomerulonephritis, amyloidosis, monoclonal immunoglobulin deposition Syndrome, fibrillary glomerulonephritis, immunotactoid glomerulopathy, ischemic tubular injury, drug-induced tubular injury Tubulointerstitial nephritis, toxic tubulointerstitial nephritis, infectious tubulointerstitial nephritis, bacterial pyelonephritis, Viral tubulointerstitial infection resulting from riomavirus infection or HIV infection Nephritis, metabolic disease-induced tubulointerstitial disease, mixed connective tissue disease, cylindrical nephropathy, urate or sulfate Crystalline nephropathy, acute cellular tubulointerstitial nephropathy, which can result from uric acid or drug-induced crystal deposition. Tumors resulting from allograft rejection, lymphoma, or post-transplant lymphoproliferative disorders. Invasive disease, obstructive kidney disease, vascular disease, thrombotic microangiopathy, renal artery sclerosis, atheroma Embolistic diseases, mixed connective tissue disease, polyarteritis nodosa, calcineurin inhibitor-induced vascular diseases Patients: acute cellular vascular allograft rejection, acute humoral allograft rejection, early renal dysfunction End-stage renal disease (ERFD), end-stage renal disease (ESRD), renal vein thrombosis, acute tubular necrosis, acute interstitial Nephritis, pre-existing chronic kidney disease, renal artery stenosis, ischemic nephropathy, uremia, drug and toxin-induced chronic conditions. Tubulointerstitial nephritis, reflux nephropathy, kidney stones, Goodpasture syndrome, normocytic normochromic anemia Hematologic dysregulation, renal anemia, diabetic chronic kidney disease, IgG4-related disease, von Hippel-Lindau disease syndrome, tuberous sclerosis, nephronophthisis, renal medullary cystopathy, renal cell carcinoma, adenocarcinoma, nephroblastoma, lymphoma, Leukemia, hyposialylation disorder, chronic cyclosporine nephropathy, renal reperfusion injury, renal dysplasia, hyperurea Blood pressure, bilateral arterial occlusion, acute uric nephropathy, hypovolemia, acute bilateral obstructive urinary tract disease, hypercalcemia Umumemic nephropathy, hemolytic uremic syndrome, acute urinary retention, malignant nephrosclerosis, postpartum glomerulosclerosis, Scleroderma, non-Goodpasture anti-GBM disease, microscopic polyarteritis nodosa, allergic meat Blastomatosis, acute radiation nephritis, post-streptococcal glomerulonephritis, Waldenström macroglomerulonephritis Bulinemia, analgesic nephropathy, arteriovenous fistula, arteriovenous graft, dialysis, ectopic kidney, cavernous kidney, renal bone Dystrophy, mononephrosis, hydronephrosis, microalbuminuria, uremia, hematuria, hyperlipidemia, hypoalbuminuria Select from the group consisting of buminemia, lipiduria, acidosis, hyperkalemia, and edema. In some embodiments, the kidney-related disease or condition is a chronic kidney disease. [Brief explanation of the drawing]

[0118] [Figure 1] Figure 1 shows the amino acid sequence of the native precursor of the B (short) isoform of the human TGFβ receptor type II (hTβRII) (NP_003233.4). The solid underline indicates the mature extracellular domain (ECD) (residues 23-159), and the double underline indicates the valine that is replaced in the A (long) isoform. The dotted underline indicates the leader (residues 1-22).

[0119] [Figure 2] Figure 2 shows the amino acid sequence of the native precursor of the A (long) isoform of human TβRII (NP_001020018.1). The solid underline indicates the mature ECD (residues 23-184), and the double underline indicates isoleucine substitutions resulting from splicing. The dotted underline indicates the leader (residues 1-22).

[0120] [Figure 3] Figure 3 shows a comparison of the linker sequences of five different TβRII constructs.

[0121] [Figure 4A] Figures 4A and 4B show in tabular form the binding affinity between TGFβ1 and TGFβ3 and one of several different TβRII-Fc fusion protein constructs. [Figure 4B] Same as above.

[0122] [Figure 5A]Figures 5A and 5C graph the results from a reporter gene assay testing the affinity of TGFβ1 to one of several different TβRII-Fc fusion protein constructs. Figures 5B and 5D graph the results from a reporter gene assay testing the affinity of TGFβ3 to one of several different TβRII-Fc fusion protein constructs. Figures 5E and 5F provide IC50 data from these same experiments in tabular format. [Figure 5B] Same as above. [Figure 5C] Same as above. [Figure 5D] Same as above. [Figure 5E] Same as above. [Figure 5F] Same as above. [Modes for carrying out the invention]

[0123] Detailed description of the invention 1.Summary The proteins described herein are in human form unless otherwise specified. The NCBI reference for the substance is as follows: human TβRII isoform A(h TβRII ロング ), NP_001020018.1, and human TβRII isopho 'B(hTβRII ショート ), (NP_003233.4). Native from humans The sequences of the TβRII protein are shown in Figures 1 and 2. In some embodiments, T βRII proteins are derived from non-human animals such as mice, rats, dairy cows, or monkeys. That is the case.

[0124] The TGFβ superfamily shares common sequence elements and structural motifs, and is diverse in its composition. It contains growth factors. These proteins are highly effective in both vertebrates and invertebrates. It is well known that it exerts biological effects on a wide variety of cell types. Superfamily The members play important roles in pattern formation and tissue specificity during embryonic development. Various processes including adipogenesis, myogenesis, chondrogenesis, cardiogenesis, hematopoiesis, neurogenesis, and epithelial cell differentiation. It can influence the differentiation process. It can manipulate the activity of members of the TGFβ family. By doing so, it is often possible to cause significant physiological changes in living organisms. For example Piedmontese and Belgian Blue cattle breeds exhibit a significant increase in muscle mass due to GD. It carries a loss-of-function mutation in the F8 gene (also known as myostatin). Grobet et al. Nat Genet. 1997, Vol. 17 (No. 1): pp. 71-74. Similarly, in humans, GDF8 The inactive allele is thought to be involved in muscle mass increase and, where it is believed, in exceptional strength. Related. Schuelke et al., N Engl J Med 2004, Vol. 350: pp. 2682-2688.

[0125] TGFβ signaling is classified into type I (e.g., TβRI) and type II (e.g., TβRII). The serine / threonine kinase receptor heteromer complex mediated ligand stimulation It then phosphorylates and activates downstream SMAD proteins (Massague, 2000, Na (t. Rev. Mol. Cell Biol. Vol. 1: pp. 169-178). These type I and type II receptors The substance consists of a ligand-binding extracellular domain with a cysteine-rich region and a transmembrane domain. It consists of a transmembrane and a cytoplasmic domain having predictive serine / threonine specificity. It is a protein. Type I receptors are essential for signal transduction, and type II receptors are rigorous. It is required for ligament binding and for the expression of type I receptors. Type I and type II receptors are ligament binding. After binding, a stable complex is formed, and as a result, the type I receptor is phosphorylated by the type II receptor. It will be oxidized. TGFβ is divided into three types: TGFβ1, TGFβ2, and TGFβ3. It has three mammalian isoforms, each of which has distinctly different mechanisms in vivo. It has the ability. The binding of TGFβ to TβRII opens the TGFβ signaling pathway. This is an extremely important step in getting things started, and this activation causes SMAD2 to be phosphorylated. As a result, the activated SMAD2 / SMAD4 complex is translocated to the nucleus and gene generation occurs. This will modulate the current state.

[0126] Therefore, in certain embodiments, this disclosure relates to various TGFβ1 or TGFβ3-related As an antagonist of TGFβ1 or TGFβ3 for use in treating disorders, Tβ We provide RII polypeptides that are not constrained by any particular mechanism of action. However, such polypeptides bind to TGFβ1 or TGFβ3 and these ligands It is expected to act by inhibiting the ability of the ternary signaling complex to form. .

[0127] This disclosure relates to a fusion tan comprising a TβRII polypeptide and a heterogeneous portion (e.g., an Fc portion). Provides protein. In certain embodiments, the TβRII portion and the heterogeneous portion are linked to the linker. Therefore, they are fused. As will be explained in more detail below, this disclosure is a linker of a certain length. TβRII-Fc fusion protein containing (for example, a linker with 21 amino acids) However, surprisingly, the TβRII-Fc fusion protein has a linker consisting of only four amino acids. We demonstrated that it was possible to bind to TGFβ-1 and TGFβ-3 with stronger affinity than the original substance. To testify.

[0128] The terms used herein are used in the context relating to the present invention, and in the context in which each term is used. In a particular context, these generally have their usual meanings in the art. Additional guidance in describing compositions and methods, as well as their manufacture and use. In order to provide the implementers with information, certain terms are discussed below or elsewhere in this specification. The scope or meaning of any use of a term depends on the specific context in which that term is used. This becomes clear.

[0129] All of its grammatical forms and spelling variations of "homonymy" refer to the same species of living organism. Proteins from superfamilies, as well as homologous proteins from different species of organisms. This refers to a relationship between two proteins that share a "common evolutionary origin," including such relationships. Proteins (and the nucleic acids that encode them) are identified by the similarity of their sequences. They possess such sequence homology, and such similarity may also be expressed in terms of identity percentage. This may be due to the presence of stones, or specific residues or motifs and conserved locations. stomach.

[0130] The term "sequence similarity" in all its grammatical forms refers to the sharing of a common evolutionary origin. The degree of similarity or correspondence between nucleic acid or amino acid sequences, which may or may not be shared. This refers to [something]. However, in general usage and in this application, the term "homonymous" means "to a high degree" When modified by any adverb, it can refer to sequence similarity and relating to a common evolutionary origin. It may or may not be related.

[0131] Sequence identity percentage (%) for a reference polypeptide (or nucleotide) sequence )" or "percent (%) identical" refers to the same value within the reference polypeptide (nucleotide) sequence. Amino acid residues (or nucleic acids) within a candidate sequence that are identical to an amino acid residue (or nucleic acid) It is a percentage, and in order to achieve the maximum sequence identity percentage, as needed. , after aligning the sequences and introducing gaps, and any conservative substitution maintains sequence identity It is defined as the percentage that is not considered part of the amino acid sequence identity percentage. Alignment for determination is performed in various ways within the scope of skills in the relevant technical field. For example, BLAST, BLAST-2, ALIGN, or Megalign(DNAS Achieved using publicly available computer software such as TAR software. It is possible. A person skilled in the art can determine the maximum alignment over the full length of the sequences being compared. Appropriate parameters for array alignment, including any algorithm necessary to achieve this. The meter can be determined. However, in this specification, amino acid (nucleic acid) sequence identity % The values ​​are generated using the sequence comparison computer program ALIGN-2. The N-2 sequence comparison computer program is a copyrighted work of Genentech, Inc. The source code is licensed to the U.S. Copyright Office, Washington DC, 20559. It was filed together with the other documents and is registered under U.S. Copyright Registration No. TXU510087. The ALIGN-2 program is developed by Genentech, Inc., South San It is publicly available from Francisco, Calif., or can be compiled from the source code. It can be done. The ALIGN-2 program is Digital UNIX (registered trademark) Compiled for use with UNIX® operating systems, including V4.0D. It is necessary to set this up. All sequence comparison parameters are set by the ALIGN-2 program. It's set that way and doesn't change.

[0132] In all its grammatical forms, "agonize" refers to proteins and / or Or, to activate or amplify the gene expression of that protein (for example, to activate or amplify the gene expression of that protein) Activation (by means of, or by inducing an inactive protein to enter an active state) A process that increases the activity of proteins and / or genes. To point.

[0133] In all its grammatical forms, "antagonize" refers to proteins and / or genes (e.g., For example, by inhibiting or reducing the gene expression of that protein, or by activating the protein The process of inhibiting a protein (by inducing it to enter an inactive state), or the process of inhibiting a protein. This refers to a process that reduces the activity of proteins and / or genes.

[0134] The terms “approximately” and “about” refer to numerical values ​​throughout this specification and these claims. When used in conjunction, it refers to a precise section that is well known and recognized by those skilled in the art.

[0135] Numerical ranges disclosed herein include numbers that define the range.

[0136] The terms "a" and "an" depend on the context in which they are used. Unless otherwise clearly indicated, it includes multiple referents. The term "one (a)" (or " The terms "one (an)", as well as "one or more" and "at least one", are used in this context. In the specification, they may be used synonymously. Furthermore, "and / or" as used herein In some cases, two or more specified features, with or without other features, This shall be considered a specific disclosure for each of the components. Therefore, the term "and" When used herein in expressions such as "A and / or B", the term "and / or" means This includes "A and B", "A or B", "A" (alone), and "B" (alone). It is intended to mean "A, B, and / or C". Similarly, the term "and / or" means "A, B, and / or C". When used in expressions such as the following, it is intended to encompass each of the following aspects: A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0137] Throughout this specification, the word "comprise" or its inflected forms, for example, "comprise," is used. "rises" or "comprising" refers to the integer or group of integers being described. It is understood that this implies inclusion, but does not imply the exclusion of any other integer or any group of integers. As used herein, the term "contains" is used in a narrower sense than the term "consisting of". This also includes the use of "to essentially become."

[0138] The term "essentially derived from" refers to the materials or steps specified herein. This is limited to those that do not significantly affect the fundamental novel features of the disclosed invention.

[0139] The term "perceptible affinity," as used herein, refers to a dissociation constant less than 50 nM. (K DThis means joining at ).

[0140] The terms "polypeptide," "oligopeptide," "peptide," and "protein" are, In this specification, the terms are used synonymously to refer to chains of amino acids of any length. The chains are linear in shape. It may be present, or it may be branched, and it may contain modified amino acids. Non-amino acid molecules may be interspersed in the chain. These terms are used in nature or through intervention. Amino acid chains modified by the addition of, for example, disulfide bond formation, glycosylation, lipid attachment. Addition, acetylation, phosphorylation, or any other operation or modification, such as with a labeling component. This definition also includes amino acid chains having densification. Polypeptides containing one or more analogues (e.g., including unnatural amino acids) This also includes polypeptides containing other modifications known in the art. Peptides can exist as single chains or as associated chains. This is how it is understood.

[0141] 2. TβRII polypeptide The naturally occurring TβRII protein is the protein portion located on the outside of the cell ( A transmembrane has an extracellular portion and a protein portion located inside the cell (intracellular portion). It is a common protein. Aspects of this disclosure include mutations in the extracellular domain of TβRII and / or containing a mutant TβRII polypeptide that includes a shortened portion of the extracellular domain. As noted, human TβRII undergoes alternative splicing in the extracellular domain (ECD). At least two isoforms are generated by - A (long) and B (short) -naturally present (Figure 1 and 2 and SEQ ID NO: 1 and 2). Residue 2 of SEQ ID NO: 1 SEQ ID NO: 27 corresponding to 3 to 159 represents the native full-length extracellular domain of the short isoform of TβRII SEQ ID NO: 18 corresponding to residues 23 to 184 of SEQ ID NO: 2 represents the native full-length extracellular domain of the long isoform of TβRII represents the native full-length extracellular domain of the long isoform of TβRII. Unless otherwise stated, the numbering of amino acid positions for mutants based on the TβRII short and long isoforms refers to the corresponding positions in SEQ ID NO: 1 and SEQ ID NO: 2, which are the native precursors respectively

[0142] In certain embodiments, the present disclosure provides mutant TβRII polypeptides. The TβRII polypeptides of the present disclosure can bind to TGFβ superfamily members such as, but not limited to, TGFβ1 or TGFβ3, and inhibit their functions The TβRII polypeptide may consist of, or include, an amino acid sequence that is at least 80% identical, and optionally at least 85%, 90%, 95% 96%, 97%, 98%, 99%, or 100% identical, to the truncated ECD domain of a naturally occurring TβRII polypeptide whose C-terminus is present at any of amino acids 153 to 1 59 of SEQ ID NO: 1 The TβRII polypeptide may consist of, or include, an amino acid sequence that is at least 80% identical, and optionally at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical, to the truncated ECD domain of a naturally occurring TβRII polypeptide whose C-terminus is present at any of amino acids 178 to 184 of SEQ ID NO: 2 or include such an amino acid sequence The C-terminus of the TβRII polypeptide is present at any of amino acids 178 to 184 of SEQ ID NO: 2, and is at least 80% identical to the truncated ECD domain of a naturally occurring TβRII polypeptide and optionally at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical, and consists of, or includes, such an amino acid sequence ​ It may contain a polypeptide. In certain embodiments, the TβRII polypeptide has an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO: 1 and optionally at least 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical, and includes an amino acid sequence. Optionally, the TβRII polypeptide is from the sequence consisting of amino acids 160 - 567 of SEQ ID NO: 1, or from the sequence consisting of amino acids 185 - 592 of SEQ ID NO: 2 and does not include more than 5 consecutive amino acids, or 10, 20, 30, 40, 50, 52, 60, 70, 80, 90, 100, 150, or 200 or more consecutive amino acids. In some embodiments, the TβRII polypeptide does not include amino acids 160 - 567 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 and 160 - 567 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 185 - 592 of SEQ ID NO: 2. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 of SEQ ID NO: 2. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 and 185 - 592 of SEQ ID NO: 2. The unprocessed TβRII polypeptide may or may not include any signal sequence and any sequence on the N-terminal side of the signal sequence. As detailed herein, the N-terminus of the mature (processed) TβRII polypeptide may be at either amino acids 23 - 35 of SEQ ID NO: 1 or 23 - 60 of SEQ ID NO: 2. more consecutive amino acids. In some embodiments, the TβRII polypeptide does not include amino acids 160 - 567 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 and 160 - 567 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 185 - 592 of SEQ ID NO: 2. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 of SEQ ID NO: 2. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 and 185 - 592 of SEQ ID NO: 2. The unprocessed TβRII polypeptide may or may not include any signal sequence and any sequence on the N-terminal side of the signal sequence. As detailed herein, the N-terminus of the mature (processed) TβRII polypeptide may be at either amino acids 23 - 35 of SEQ ID NO: 1 or 23 - 60 of SEQ ID NO: 2. It may contain a polypeptide. In certain embodiments, the TβRII polypeptide has an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO: 1 and does not include more than 5 consecutive amino acids, or 10, 20, 30, 40, 50, 52, 60, 70, 80, 90, 100, 150, or 200 or more consecutive amino acids. In some embodiments, the TβRII polypeptide does not include amino acids 160 - 567 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 and 160 - 567 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 185 - 592 of SEQ ID NO: 2. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 of SEQ ID NO: 2. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 and 185 - 592 of SEQ ID NO: 2. The unprocessed TβRII polypeptide may or may not include any signal sequence and any sequence on the N-terminal side of the signal sequence. As detailed herein, the N-terminus of the mature (processed) TβRII polypeptide may be at either amino acids 23 - 35 of SEQ ID NO: 1 or 23 - 60 of SEQ ID NO: 2. It may contain a polypeptide. In certain embodiments, the TβRII polypeptide has an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO: 1 and does not include more than 5 consecutive amino acids, or 10, 20, 30, 40, 50, 52, 60, 70, 80, 90, 100, 150, or 200 or more consecutive amino acids. In some embodiments, the TβRII polypeptide does not include amino acids 160 - 567 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 and 160 - 567 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 185 - 592 of SEQ ID NO: 2. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 of SEQ ID NO: 2. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 and 185 - 592 of SEQ ID NO: 2. The unprocessed TβRII polypeptide may or may not include any signal sequence and any sequence on the N-terminal side of the signal sequence. As detailed herein, the N-terminus of the mature (processed) TβRII polypeptide may be at either amino acids 23 - 35 of SEQ ID NO: 1 or 23 - 60 of SEQ ID NO: 2. It may contain a polypeptide. In certain embodiments, the TβRII polypeptide has an amino acid sequence that is at least 80% identical to the amino acid sequence of SEQ ID NO: 1 and does not include more than 5 consecutive amino acids, or 10, 20, 30, 40, 50, 52, 60, 70, 80, 90, 100, 150, or 200 or more consecutive amino acids. In some embodiments, the TβRII polypeptide does not include amino acids 160 - 567 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 and 160 - 567 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 185 - 592 of SEQ ID NO: 2. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 of SEQ ID NO: 2. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 and 185 - 592 of SEQ ID NO: 2. The unprocessed TβRII polypeptide may or may not include any signal sequence and any sequence on the N-terminal side of the signal sequence. As detailed herein, the N-terminus of the mature (processed) TβRII polypeptide may be at either amino acids 23 - 35 of SEQ ID NO: 1 or 23 - 60 of SEQ ID NO: 2. and may or may not include any signal sequence and any sequence on the N-terminal side of the signal sequence. As detailed herein, the N-terminus of the mature (processed) TβRII polypeptide may be at either amino acids 23 - 35 of SEQ ID NO: 1 or 23 - 60 of SEQ ID NO: 2. and may or may not include any signal sequence and any sequence on the N-terminal side of the signal sequence. As detailed herein, the N-terminus of the mature (processed) TβRII polypeptide may be at either amino acids 23 - 35 of SEQ ID NO: 1 or 23 - 60 of SEQ ID NO: 2. 70, 80, 90, 100, 150, or 200 or more consecutive amino acids. In some embodiments, the TβRII polypeptide does not include amino acids 160 - 567 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 and 160 - 567 of SEQ ID NO: 1. In some embodiments, the TβRII polypeptide does not include amino acids 185 - 592 of SEQ ID NO: 2. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 of SEQ ID NO: 2. In some embodiments, the TβRII polypeptide does not include amino acids 1 - 22 and 185 - 592 of SEQ ID NO: 2. The unprocessed TβRII polypeptide may or may not include any signal sequence and any sequence on the N-terminal side of the signal sequence. As detailed herein, the N-terminus of the mature (processed) TβRII polypeptide may be at either amino acids 23 - 35 of SEQ ID NO: 1 or 23 - 60 of SEQ ID NO: 2.

[0143] An example of a mature TβRII polypeptide is amino acids 23-159 of sequence number 1 (sequence number 1). (Shown as number 27), amino acids 29-159 of SEQ ID NO: 1 (Shown as number 28) , amino acids 35-159 of SEQ ID NO: 1 (shown in SEQ ID NO: 29), amino acids of SEQ ID NO: 1 Acids 23-153 (shown in SEQ ID NO: 30), amino acids 29-153 of SEQ ID NO: 1 (sequence) (Shown as number 31), amino acids 35-153 of SEQ ID NO: 1 (Shown as number 32) , amino acids 23-184 of SEQ ID NO: 2 (shown in SEQ ID NO: 18), amino acids of SEQ ID NO: 2 Acids 29-184 (shown in SEQ ID NO: 33), amino acids 60-184 of SEQ ID NO: 2 (sequence) (shown in number 29), amino acids 23-178 of SEQ ID NO: 2 (shown in SEQ ID NO: 34) , amino acids 29-178 of SEQ ID NO: 2 (shown in SEQ ID NO: 35), and of SEQ ID NO: 2 Examples include amino acids 60-178 (shown in SEQ ID NO: 32), but are not limited to these. The corresponding mutant based on the long isoform of TβRII has a 25-amino acid insertion. It codes and also encodes a conservative Val-Ile substitution at the adjacent position on the C-terminal side of the insertion. It is expected that those skilled in the art will understand that this will include a nucleotide sequence. Therefore, TβRII polypeptide has short isoforms and long isoforms. TβRII polypeptides containing both, and their variants (e.g., amino acid 2 of SEQ ID NO: 1) 2, 3, 4, 5 in the sequence corresponding to amino acids 3-159 or 23-184 of SEQ ID NO: 2 Includes variants containing 10, 15, 20, 25, 30, or 35 or fewer amino acid substitutions. (m), those fragments, and the fusion protein containing any of the above, isolated extracellularly It may include a portion, but in any case, preferably any of the aforementioned TβRII polypeptides Both are substantial parents of at least one or both of TGFβ1 or TGFβ3. This will preserve the compatibility. Generally, TβRII polypeptides are biologically appropriate at the appropriate temperature. It is designed to be soluble in aqueous solutions at the given pH level and molar osmotic pressure concentration. ru.

[0144] In some embodiments, the mutant TβRII polypeptide of the Disclosure has a ligand-binding profile The extracellular domain contains one or more mutations that result in a change in the TβRII poly(TβRII) The peptide has a different amino acid distribution compared to the corresponding portion of the naturally occurring TβRII polypeptide. This may include changes to 1, 2, 5 or more columns. In some embodiments, mutations are This results in a substitution, insertion, or deletion at the position corresponding to position 70 of sequence number 1. In this embodiment, the mutation is a substitution or insertion at the position corresponding to position 110 of SEQ ID NO: 1. This results in deletions. For example, at positions corresponding to positions 70 and 110 of sequence number 1. Examples include substitutions from N to D or from D to K, respectively, but are not limited to these. No. Examples of such mutated TβRII polypeptides include SEQ ID NOs. 36-39. The sequences shown are, but are not limited to, those listed. The TβRII polypeptide is a sequence Number 10, 12, 14, or 16, or their silent variants, or stringer Either of the nucleic acids that hybridize to its complement under specific hybridization conditions. A polypeptide or a portion thereof, coded by one of the following in a particular embodiment. TβRII polypeptide is sequence number 12, or its silent variant, or A nucleic acid that hybridizes to its complement under stringent hybridization conditions, may contain a polypeptide or a portion thereof encoded by any one of

[0145] In some embodiments, the mutant TβRII polypeptide of the present disclosure is the human TβRII iso naturally present in form C (Konrad et al., BMC Genomics 8:318, 2007), a 36 - amino acid insertion (SEQ ID NO: 41), between a pair of glutamic acid residues (positions 151 and 152 of SEQ ID NO: 1 or positions 176 and 177 of SEQ ID NO: 2) located near the C - terminus of the human TβRII ECD.

[0146] The present disclosure further demonstrates that the TβRII polypeptide can be modified to selectively antagonize the TβRII ligand. The N70 residue is representative of a potential glycosylation site. In some embodiments, the TβRII polypeptide is not glycosylated. In some embodiments, the TβRII polypeptide has no glycosylation or reduced glycosylation at position Asn157. In some embodiments, the TβRII polypeptide has no glycosylation or reduced glycosylation at position Asn73.

[0147] In certain embodiments, the TβRII polypeptide binds to TGFβ1 and the TβRI I polypeptide does not show substantial binding to TGFβ3. In certain embodiments, the TβRII polypeptide binds to TGFβ3 and the TβRII polypeptide does not show substantial binding to TGFβ 1. Binding is determined using purified proteins in solution or B ​​​​​​​Purified proteins in surface plasmon resonance systems such as the iacore(trademark) system Quality can be used to make an assessment.

[0148] In certain embodiments, TβRII polypeptides enhance TGFβ1 cell signaling. Inhibits TβRII polypeptide, which has a moderate or severe effect on TGFβ3 signaling. It has a significant inhibitory effect. In a particular embodiment, the TβRII polypeptide is TGF By inhibiting β3 cell signaling, TβRII polypeptide inhibits TGFβ1 signaling. In contrast, it has a moderate or slight inhibitory effect. The inhibitory effect on cell signaling is The assay can be performed by methods known in the art.

[0149] In summary, the active portion of the TβRII polypeptide is amino acid sequence 23- of SEQ ID NO: 1. 153, 23-154, 23-155, 23-156, 23-157, or 23-15 These sequences may contain 8 and begin with any of amino acids 24-35 of SEQ ID NO: 1. It may also include variants of the TβRII polypeptide. Similarly, the active portion of the TβRII polypeptide is SEQ ID NO: 2 The amino acid sequences are 23-178, 23-179, 23-180, 23-181, and 23-18 2, or may contain 23-183, and any of amino acids 24-60 of SEQ ID NO: 2 These may also include variants of these sequences that begin with . Exemplary TβRII polypeptides are distributed Amino acids 29-159, 35-159, 23-153, 29-153 and 3 in row number 1 5-153, or amino acid sequence 29-184, 60-184, 23-17 of SEQ ID NO: 2 Includes 8, 29-178 and 60-178. Variants within these ranges, in particular, SEQ ID NOs. At least 80%, 85%, 90%, 95% for the corresponding portion of 1 or 2. Alternatively, it is possible that some have 99% identity. Amino acids 160-56 of SEQ ID NO: 1. TβRII polypeptide that does not contain the sequence consisting of amino acids 185-592 of sequence 7 or SEQ ID NO: 2 A peptide can be selected. In a particular embodiment, the TβRII polypeptide is sequence The amino acid sequence of number 18 is at least 80% identical, and at least 8 if necessary. 5%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical, mesh Contains an acid sequence.

[0150] As explained above, this disclosure identifies the naturally occurring TβRII polypeptide. Provides a TβRII polypeptide that shares the same or similar sequence as two amino acids. To determine the percentage of acid sequence identity, the sequences were aligned for optimal comparison. (For example, for optimal alignment, the first and second amino acid or nucleic acid sequences A gap can be introduced in one or both sequences, allowing for the ignorance of non-homologous sequences for comparison. (and so on). Then, the amino acid residues at the corresponding amino acid positions are compared. First sequence If the position in the middle is occupied by the same amino acid residue as the corresponding position in the second sequence. In this context, those molecules are identical at their respective positions (wherein used herein, amino acids " "Identity" is synonymous with amino acid "homology." The percentage of identity between two sequences is 2 The number of gaps that need to be introduced for optimal alignment of the sequence and each gap It is a function of the number of identical positions shared by those arrays, taking their lengths into account.

[0151] Sequence comparison and the determination of the percentage of identity and similarity between two sequences are performed using a mathematical algorithm. Zoom (Computational Molecular Biology, edited by Lesk, AM, Oxford University Press) , New York, 1988; Biocomputing: Informatics and Genome Projects, Smit h, DW (ed.), Academic Press, New York, 1993; Computer Analysis of Sequence ce Data, Part 1, edited by Griffin, AM and Griffin, HG, Humana Press, New Jersey , 1994; Sequence Analysis in Molecular Biology, von Heinje, G., Academ. ic Press, 1987; and Sequence Analysis Primer, Gribskov, M. and Devere This can be done using (edited by J. ux, M., Stockton Press, New York, 1991). ru.

[0152] In one embodiment, the percentage of identity between two amino acid sequences is determined by Needleman and Wunsch (J Using the algorithm (Mol.Biol. (Vol. 48): pp. 444-453 (1970)) This algorithm is defined and is used in the GAP program of the GCG software package (http It is incorporated in (available at : / / www.gcg.com). In certain embodiments, the following parameters However, the Blosum 62 matrix or PAM250 matrix used in the GAP program Either, and the weights of the gaps are 16, 14, 12, 10, 8, 6 or 4, and 1 , weights of length 2, 3, 4, 5, or 6. In yet another embodiment, a 2-nucleotide sequence The percentage of identity between them is the GAP program (Devereux) of the GCG software package. , J. et al., Nucleic Acids Res. Vol. 12 (No. 1): p. 387 (1984) (http: / / www. Determined using (available at gcg.com). Exemplary parameters are NWSgapdna .CMP matrix and weights of 40, 50, 60, 70 or 80 gaps and 1, 2, Includes the use of weights of length 3, 4, 5, or 6. Unless otherwise specified, a 2-amino acid sequence. The percentage of identity between them is calculated using the GAP program, with a Blosum 62 matrix of 10 The weights of the GAP and the length of 3 are used to determine the algorithm, and such an algorithm is determined. If the rhythm cannot calculate the desired identity percentage, the appropriate method disclosed herein An alternative method needs to be chosen.

[0153] In another embodiment, the percentage of identity between two amino acid sequences is determined by the ALIGN program ( The E. Myers and W. Miller algorithm (CAB) is incorporated into version 2.0. Using IOS, Vol. 4: pp. 11-17 (1989), PAM120 residue mass table, 12 The gap length penalty and a 4-point gap penalty are used to determine the result.

[0154] Another embodiment for determining the best overall alignment between two amino acid sequences is Br The algorithm by utlag et al. (Comp. App. Biosci. Vol. 6: pp. 237-245 (1990) This can be determined using the FASTDB computer program based on ). In column alignment, both the query sequence and the target sequence are amino acid sequences. The results of the comprehensive sequence alignment are presented as an identity percentage. In this context, amino acid sequence identity is determined by the algorithm developed by Brutlag et al. (Comp. App. Biosci. Vol. 6). Using the FASTDB computer program based on pages 237-245 (1990) This is done using [a specific method]. In certain embodiments, the identity and similarity of amino acid alignments are [a specific method]. The parameters used in calculating the cents are matrix = PAM 150, k-set = 2, mismatch Chi penalty = 1, combined penalty = 20, randomization group length = 0, cutoff score = 1. Includes a gap penalty of 5 and a gap size penalty of 0.05.

[0155] The TβRII polypeptide further contains one of various leader sequences at its N-terminus. There are also such sequences that target peptide expression and secretion pathways in eukaryotic cells. This will make it possible. For example, Ernst et al., U.S. Patent No. 5,082,783 (199 See (2nd year). Alternatively, use the native TβRII signal sequence in the cell It can perform the role of pushing from. Possible leader arrays include native leaders and organizational leaders. Plasminogen activator (TPA) and honeybee melittin (sequence numbers, respectively) Includes (numbers 22-24). Examples of TβRII-Fc fusion proteins containing TPA reader sequences. Examples include SEQ ID NOs. 11, 13, 15, and 17. Among the many variables involved in sensing, the most important are the selected leader sequence, the cell type used, and This can vary depending on the culture conditions, and therefore, the actual results of mature TβRII polypeptides. The N-terminal start site is either 1, 2, 3, 4 or 1 in the N-terminal direction or the C-terminal direction. A shift of 5 amino acids can occur. An example of a TβRII-Fc fusion protein is the sequence number. Examples include 11, 13, 15, and 17, based on the long isoform of TβRII. The corresponding mutant includes a 25-amino acid insertion, and a conservative value is located adjacent to the C-terminal side of the insertion. Those skilled in the art will understand that this will involve Val-Ile substitution.

[0156] In some embodiments, any of the TβRII polypeptides disclosed herein is sequence Number 18, 27, 30, 34, 36, 37, 38, 39, 48, 49, or 51 or one amino acid sequence and at least 80%, 85%, 90%, 92%, 94%, 95%, They are 97%, 99%, or 100% identical, but sequence numbers 18, 27, 30, 34, 36, Compared to the amino acid sequence of 37, 38, 39, 48, 49, or 51, one or more N It lacks terminal amino acids. In some embodiments, the TβRII polypeptide is SEQ ID NO: 18, 27, 30, 34, 36, 37, 38, 39, 48, 49, or 51 It lacks an amino acid corresponding to one of the first amino acids (threonine). In some embodiments, TβRII polypeptide is represented by sequence numbers 18, 27, 30, 34, 36, 37, 38, 39, 48, 49, or 51, one of the first and second amino acids (each, It lacks the amino acids corresponding to rheonine and isoleucine. In some embodiments, TβRII polypeptide is represented by SEQ ID NOs. 18, 27, 30, 34, 36, 37, 38, 39 , 48, 49, or 51, one of the first, second, and third amino acids (each, It lacks the amino acids corresponding to threonine, isoleucine, and proline. In terms of application, TβRII polypeptide is used in sequence numbers 18, 27, 30, 34, 36, and 37. , one of the first, second, third and fourth A of 38, 39, 48, 49 or 51 amino acids (threonine, isoleucine, proline, and proline, respectively) correspond to amino acids It lacks acid.

[0157] In some embodiments, any of the TβRII polypeptides disclosed herein is sequence One amino acid sequence with either number 18 or 51 and at least 80%, 85%, or 90% , 92%, 94%, 95%, 97%, 99%, or 100% identical, but sequence number 18 Alternatively, it lacks the amino acid corresponding to the first amino acid (threonine) of the 51st amino acid. In the application form, the TβRII polypeptide is the first and second A of SEQ ID NOs. 18 or 51. It lacks the amino acids corresponding to the amino acids (threonine and isoleucine, respectively). In some embodiments, the TβRII polypeptide is the first, second of SEQ ID NOs. 18 or 51. and the third amino acid (threonine, isoleucine, and proline, respectively) It lacks the amino acid. In some embodiments, the TβRII polypeptide is SEQ ID NO: 1 8 or 51 first, second, third, and fourth amino acids (threonine, isolol, respectively) It lacks the amino acids corresponding to isine, proline, and proline.

[0158] In some embodiments, the present disclosure is a composition comprising a mixture of TβRII polypeptides. Each of the TβRII polypeptides in the composition corresponds to SEQ ID NOs: 18, 27, 30, 34, 36 , one of the amino acid sequences 37, 38, 39, 48, 49, or 51 and at least 80%, 85%, 90%, 92%, 94%, 95%, 97%, 99%, or 100% identical It contains the amino acid sequence, but at least a portion of the TβRII polypeptide in the composition ( For example, at least 1%, 3%, 4%, 5%, 10%, 15%, 20%, 25%, 30% , 40%, 50%, 60%, 70%, 80%, 90%, 95%) are represented by sequence numbers 18 and 27. , one of the first, 30, 34, 36, 37, 38, 39, 48, 49 or 51 The second, third, and fourth amino acids (threonine, isoleucine, and proline, respectively) It contains amino acids corresponding to biproline, and at least one TβRII polypeptide in the composition Also a portion (for example, at least 1%, 3%, 4%, 5%, 10%, 15%, 20%, 25%) %, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%) are array numbers. 18, 27, 30, 34, 36, 37, 38, 39, 48, 49, or 51 The first, second, third, and fourth amino acids (threonine, isoleucine, and prudent, respectively) Compositions lacking one or more amino acids corresponding to roline and proline. Provided. In some embodiments, the present disclosure is a composition comprising a TβRII polypeptide. The TβRII polypeptide has one amino acid sequence from either SEQ ID NO: 18 or 51. At least 80%, 85%, 90%, 92%, 94%, 95%, 97%, 99%, or 1 Although 00% identical, at least 30% to 80% of the TβRII polypeptide in the composition , lacking the amino acid corresponding to the first amino acid (threonine) of sequence number 18 or 51 The present invention provides a composition.

[0159] In certain embodiments, the disclosure provides a method for altering the glycosylation of polypeptides. The aim is to induce specific mutations in the TβRII polypeptide. Such mutations are O Introducing one or more glycosylated sites, such as linked or N-linked glycosylated sites. It may be selected for the purpose of, or to eliminate. Asparagine linkage glycosylation recognition The site is a tripeptide sequence that is specifically recognized by the appropriate cellular glycosylation enzyme. , asparagine-X-threonine (or asparagine-X-serine) (where "X" is) It generally contains one or more serine or treonine amino acids. By the addition of a transcript residue, or by the addition of one or more serine or threonine residues Through this exchange, the sequence of the wild-type TβRII polypeptide (relative to the O-linked glycosylation site) You can also make changes. Either the first or third amino acid at the glycosylation recognition site. Or substitution or deletion of various amino acids in both (and / or at the 2nd position) The deletion of certain amino acids results in the deglycosylation of the modified tripeptide sequence. Another means of increasing the number of glycosylated substructures on the TβRII polypeptide is glycosides and T This is achieved by chemical or enzymatic coupling with βRII polypeptide. Depending on the coupling method, the sugars are (a) arginine and histidine; (b) free (c) Carboxyl group; (d) Free sulfhydryl group such as that of cysteine; (c) Serine (e) free hydroxyl group such as threonine or hydroxyproline; (e) pheny Aromatic residues such as rualanine, tyrosine, or tryptophan; or (f) These methods can be bonded to the amide group of lutamine. Issued in WO87 / 05330, and in the Aplin and Wriston (1981) CRC. These references are listed in Crit. Rev. Biochem., pp. 259-306, and are provided by: Incorporated herein by reference: one or The removal of multiple sugar chain substructures can be carried out chemically and / or enzymatically. Chemical deglycosylation is a process that involves, for example, the chemical deglycosylation of a compound such as trifluoromethanesulfonic acid or an equivalent compound. This may involve exposure of the compound to TβRII polypeptide. This procedure preserves the amino acid sequence. While retaining the linked sugars (N-acetylglucosamine or N-acetylgalactosamine) This results in the cleavage of most or all sugars except for (n). Chemical deglycosylation This is according to Hakimuddin et al. (1987) Arch. Biochem. Biophys. 259: 52. Furthermore, according to Edge et al. (1981) Anal. Biochem. Vol. 118: p. 131, It is listed that the enzymatic cleavage of the glycosylated substructure on TβRII polypeptide was performed by Thotakura et al. As described in (1987) Meth. Enzymol. Vol. 138: p. 350, various This can be accomplished by the use of endoglycosidases. Mammals, yeast, Insect and plant cells are all different types that can be influenced by the amino acid sequence of peptides. Since a lycosylation pattern can be introduced, it depends on the type of expression system used. The sequence of the TβRII polypeptide can be adjusted as appropriate. Generally, in humans TβRII polypeptides for use in mammalian cell lines that provide appropriate glycosylation For example, it will be expressed in HEK293 or CHO cell lines, but in other mammals. Animal-expressing cell lines, yeast cell lines with modified glycosylation enzymes, and insect cells are also available. It is expected to be useful.

[0160] This disclosure relates to mutants, particularly combinatorial mutants of TβRII polypeptides. We are further planning to develop a set of and methods for generating shortened mutants, combinatorial A pool of al-mutants is particularly useful for identifying functional mutant sequences. The purpose of screening a national library is, for example, to identify agonists or an TβRII polypeptide variants that can be used as either tagagonists, Alternatively, a TβRII polypeptide variant possessing all the novel activities together is generated. This is possible. Various screening assays are provided below, and such assays Mutants can be evaluated using this method. For example, the TβRII polypeptide mutant can be evaluated using Ability to bind to TβRII ligand, and the relationship between TβRII ligand and TβRII polypeptide. The ability to prevent binding, or to interfere with signaling induced by TβRII ligands. The ability to cross can be screened. TβRII polypeptide or The activity of the mutants can be evaluated using cell-based assays or in vivo assays, particularly in the examples. The test can also be performed using any of the assays disclosed herein.

[0161] A mutant induced by combinatorial chemistry, and naturally occurring TβRI A selective or Generally, mutants with increased efficacy can be generated. Similarly, mutations The induction has a variant with a dramatically different serum half-life compared to the corresponding wild-type TβRII polypeptide. It is possible to create different forms. For example, denatured proteins can be subjected to proteolytic degradation. Alternatively, it may cause the breakdown, removal, or inactivation of the native TβRII polypeptide. This makes it more stable for other processes that would otherwise cause problems, or it can be made less stable. Such mutants, and they By using a gene that modulates the half-life of the TβRII polypeptide, This can alter TβRII polypeptide levels. For example, a short half-life can be used to change the TβRII polypeptide level. It can produce short-term biological effects, and recombinant TβRII polypeptide in the patient's body This allows for more precise control at the cytoplasm level. In Fc fusion proteins, Mutations that alter the half-life of the protein are linked to the linker (if any) and / or F It can be added to part c.

[0162] The combinatorial library contains fewer possible TβRII polypeptide sequences. A degenerate library of genes encoding polypeptide libraries, each containing a portion of the same molecule. It can be created by Lee. For example, a mixture of synthetic oligonucleotides enzymatically It can ligate to the gene sequence, and as a result, the potential TβRII polypeptide A degenerate set of nucleotide sequences can be expressed as individual polypeptides, and Alternatively, as a larger set of fusion proteins (e.g., phage display), It can be expressed for the purpose of [reason].

[0163] From degenerate oligonucleotide sequences, the potential TβRII polypeptide variants There are many ways to generate ibraries. This involves the chemical synthesis of degenerate gene sequences. This is done using an automated DNA synthesis machine, and then the synthesized gene is transferred to a suitable vector for expression. It can be gated. The synthesis of degenerate oligonucleotides is a circumstance in the art. This is knowledge (for example, Narang, SA (1983) Tetrahedron Vol. 39: p. 3; Itakura et al. (1981) Recombinant DNA, Proc. 3rd Cleveland Symposium. Macromolecules, A G. Walton (ed.), Amsterdam: Elsevier, pp. 273-289; Itakura et al., (1984) An nu. Rev. Biochem. Vol. 53: p. 323; Itakura et al., (1984) Science 198 See Volume 1056; Ike et al., (1983) Nucleic Acid Res. Vol. 11, p. 477. (I want to be able to do that). Such methods have been used for the directional evolution of other proteins (for example, Scott et al., (1990) Science Vol. 249: pp. 386-390; Roberts et al., (199 (2nd year) PNAS USA Vol. 89: pp. 2429-2433; Devlin et al., (1990) Science Volume 249: pp. 404-406; Cwirla et al., (1990) PNAS USA Volume 87: 6378 ~Page 6382; and U.S. Patent Nos. 5,223,409 and 5,198,346 See also Nos. 5,096,815.

[0164] Alternatively, combinatorial libraries can be generated using other forms of mutagenesis. This can be done. For example, by generating a TβRII polypeptide variant and extracting it from a library. For example, a script that uses alanine scanning mutagenesis and similar methods. Wang (Ruf et al., (1994) Biochemistry Vol. 33: pp. 1565-1572; Wang) et al., (1994) J. Biol. Chem. Vol. 269: pp. 3095-3099; Balint et al., (1 993) Gene Vol. 137: pp. 109-118; Grodberg et al., (1993) Eur. J. Biochem. Vol. 218: pp. 597-601; Nagashima et al., (1993) J. Biol. Chem. Volume 268: pp. 2888-2892; Lowman et al., (1991) Biochemistry Volume 30: 1 pp. 0832-10838; and Cunningham et al., (1989) Science Vol. 244:1 (pp. 081-1085) Linker scanning mutagenesis (Gustin et al., (199 (3rd year) Virology Vol. 193: pp. 653-660; Brown et al., (1992) Mol. Cell B Vol. 12: pp. 2644-2652; McKnight et al., (1982) Science Vol. 232: (p. 316) Saturated mutagenesis (Meyers et al., (1986) Science Vol. 232: (Page 613) by PCR mutagenesis (Leung et al., (1989) Method Cell Mol According to Biol Vol. 1: pp. 11-19; or random mutations including chemical mutagenesis. Natural mutagenesis (Miller et al., (1992) A Short Course in Bacterial Genetics, C SHL Press, Cold Spring Harbor, NY; and Greener et al., (1994) Strategies It can be isolated using the method described in *in Mol Biol*, Vol. 7, pp. 32-34. In particular, combina In a trial setting, linker scanning mutagenesis is performed on the shortened (bioactive) form of Tβ. This is an attractive method for identifying RII polypeptides.

[0165] Gene products of combinatorial libraries created by point mutation and shortening To screen for, incidentally, gene products that possess certain characteristics A wide range of methods for screening cDNA libraries are available in this technology field. This is publicly known. Such methods involve combinatorial mutations of TβRII polypeptides. Generally applicable to rapid screening of gene libraries generated by gene induction. It is expected to be effective. It is most widely used for screening large gene libraries. The method involves cloning a gene library into a replicable expression vector. The resulting vector library is used to transform appropriate cells. The relatively easy isolation of the vector encoding the detected gene is a desirable activity. Under conditions facilitated by the detection of [specific element], combinatorial gene expression is usually performed. Includes. Preferred assays include the TβRII ligand binding assay and the ligand-mediated cell assay. Includes a Gunnar signaling assay.

[0166] In certain embodiments, the TβRII polypeptide of the Disclosure is In addition to everything that naturally exists within the word, it may also include post-translational modifications. Modifications include acetylation, carboxylation, glycosylation, phosphorylation, lipid addition, and pegylation (Po This includes, but is not limited to, ethylene glycol and acylation. As a result, Modified TβRII polypeptides are polyethylene glycol, lipids, monosaccharides or polysaccharides. and may contain non-amino acid elements such as phosphates. TβRII polypeptide The effect of such non-amino acid elements on functionality is compared with other TβRII polypeptides. The variants can be tested as described herein. TβRII polyp Butido is produced in cells by cleaving the newly formed TβRII polypeptide. If so, post-translational processing also ensures proper protein folding and / or function. It is important for function. Various cells (e.g., CHO, HeLa, MDCK, 293, WI38) , NIH-3T3 or HEK-293) are specific cells for such post-translational activity. It possesses internal mechanisms and characteristic mechanisms, and involves the proper modification of TβRII polypeptides and processing It may be chosen to secure the .

[0167] 3. Linker This disclosure provides TβRII fusion proteins, and in these embodiments, the TβRII portion The components are connected to dissimilar parts (e.g., part Fc) by a linker. In some embodiments, The linker is a glycine and serine-rich linker. Other nearly neutral amino acids For example, but not limited to these, Thr, Asn, Pro, and Ala are also linkers. It can be used in arrays. In some embodiments, the linker uses Gly and Ser It contains various permutations of amino acid sequences. In some embodiments, the linker is 10Am. It is longer than the acid. In a further embodiment, the linker is at least 12, 15, It has a length of 20, 21, 25, 30, 35, 40, 45, or 50 amino acids. In the embodiment, the linker has 40, 35, 30, 25, 22, or fewer than 20 amino acids. In some embodiments, the linker is 10-50, 10-40, 10-30, 10-2 5, 10-21, 10-15, 10, 15-25, 17-22, 20, or 21 amino acids This is the length of the acid. In a preferred embodiment, the linker is the amino acid sequence GlyGlyGly GlySer(GGGGS)(sequence number 19), or its iteration (GGGGS)n(here) This includes n≧2). In certain embodiments, n≧3, or n=3~10. This application is (G The TβRII portion and the heterogeneous portion are fused together by the GGGS)4 linker. Compared to TβRII fusion proteins with n<4, TGFβ1 and TGFβ It reveals the surprising discovery that it has a strong affinity for 3. Therefore, In this embodiment, n≧4, or n=4~10. This application relates to (GGGGS)n linker(ko In this case, the protein containing n>4) is the protein having the (GGGGS)4 linker. It is also shown that it had similar inhibitory properties. Therefore, in some embodiments, (GGG In the GS)n linker, n is 4 or less. In some embodiments, n = 4 to 10, 4 ~9, 4~8, 4~7, 4~6, 4~5, 5~8, 5~7, or 5~6. Some implementation forms In this configuration, n=3, 4, 5, 6, or 7. In certain embodiments, n=4. In some embodiments... (GGGGS) n The linker containing the sequence also contains the N-terminal threonine. (Some embodiments) So, the linker is one of the following: GGGGSGGGGS (Sequence No. 21) TGGGGSGGGGS (Sequence 4) TGGGGSGGGGSGGGGS(Sequence 5) TGGGGSGGGGSGGGGSGGGGS(Sequence 6) TGGGGSGGGGSGGGGSGGGGSGGGGS(ICS25) TGGGGSGGGGSGGGGSGGGGSGGGGSGGGGS(ICS26) Taha TGGGGSGGGGSGGGGSGGGGSGGGGSGGGGSGGGGS (Sequence number No. 40)

[0168] In some embodiments, the linker is an amino acid compound of TGGGPKSCDK (SEQ ID NO: 7). Includes a column. In some embodiments, the linker lacks the N-terminal threonine (SEQ ID NO: 2). It is one of 1, 4-7, 25-26, or 40. In some embodiments, the linker - This does not include the amino acid sequence of SEQ ID NO: 26 or SEQ ID NO: 40.

[0169] 4. Different parts In certain embodiments, functional variants or modified forms of the TβRII polypeptide are Tβ A fusion tan having at least a portion of an RII polypeptide and one or more heterogeneous portions. It contains protein. Well-known examples of such heterogeneous parts include polyhistidine and Glu-Gl u, glutathione S transferase (GST), thioredoxin, protein A, p Rothein G, immunoglobulin heavy chain constant region (Fc), maltose-binding protein (MBP) Examples include, but are not limited to, human serum albumin, or the heterogeneous portion. You can select to assign the desired characteristics. For example, some heterogeneous parts are affiliate It is particularly useful for isolating fusion proteins by tide chromatography. For purification, glutathione conjugate resin, amylase conjugate resin, and affinity chromatography such as nickel or cobalt conjugate resins. A suitable matrix for the fee is used. Many such matrices are (H IS6) Useful Pharmacia GST purification system and fusion partner Available in kit form, such as the QIAexpress (trademark) system (Qiagen). It is possible. As another example, to facilitate the detection of heterologous parts of the TβRII polypeptide. , can be selected. Examples of such detection domains include various fluorescent proteins. Examples include (GFP, for example) and "epitope tags," where epitope tags are typically, It is a short peptide sequence for which a specific antibody can be used. A specific monoclonal antibody is readily available. Well-known epitope tags available for use include FLAG and influenza virus red. Examples include hemagglutination (HA) and c-myc tags. In some cases, heterogeneous parts are Protease cleavage region enables partial digestion of the fusion protein by the appropriate protease. It has a position, for example, a protease cleavage site of factor Xa or thrombin, thereby The recombinant protein is released from the fusion protein. Then, the released protein is... Subsequent chromatographic separation allows for isolation from heterogeneous regions. In one embodiment, the TβRII polypeptide is used in vivo. It is fused with a domain that stabilizes it ("stabilizer" domain). "and" refers to something that prolongs the serum half-life, and whether this prolongation is due to a decrease in disintegration, or kidney The question is whether this is due to decreased clearance by organs or to other pharmacokinetic effects. It means that fusion with the Fc portion of immunoglobulins is desired for a wide range of proteins. It is well known that this imparts specific pharmacokinetic properties. Similarly, fusion with human serum albumin is also possible. This can impart desirable characteristics. Other types of heterogeneous parts that can be selected include: Examples include polymerization domains (e.g., dimerization, tetramerization) and functional domains. .

[0170] As a specific example, this disclosure relates to TβRII fused to the Fc domain of Sequence ID No. 20. Provides a fusion protein containing polypeptide variants. If necessary, the Fc domain Asp-265, Lys-322, and Asn-434 (according to their corresponding full-length IgGs) It has one or more mutations in residues such as those that are numbered. In some cases, the patient has one or more of these mutations (e.g., the Asp-265 mutation). The mutant Fc domain has a greater ability to bind to the Fcγ receptor compared to the wild-type Fc domain. It has a decrease in [something]. In other cases, one or more of these mutations (e.g., Asn- The mutant Fc domain with the 434 mutation is more MH than the wild-type Fc domain. It has an increased ability to bind to Class C I-associated Fc receptors (FcRNs).

[0171] The different elements of the fusion protein can be arranged in any configuration that matches the desired functionality. It is understood that this is possible. For example, the TβRII polypeptide can be processed into the C-terminus of a heterogeneous domain. They may be placed at the end, or alternatively, a heterogeneous domain at the C-terminus of the TβRII polypeptide. It may be placed on the edge. The TβRII polypeptide domain and heterologous domain are fused together. They do not need to be adjacent within the protein; an additional domain or amino acid sequence can be either It may be included at the C or N-terminus of the domain, or between these domains. That's fine.

[0172] As used herein, the term “immunoglobulin Fc domain” or simply “Fc” This refers to the constant region of an immunoglobulin chain, preferably the constant region of a heavy immunoglobulin chain, or one of the same. It is understood to mean the carboxyl terminus of a part. For example, the immunoglobulin Fc region. The domains are: 1) CH1 domain, CH2 domain and CH3 domain, 2) CH1 domain and CH2 domain, 3) CH1 domain and CH3 domain, 4) CH2 domain and CH3 domain, or 5) two or more domains and immunoglobulin This may include a combination of hinge regions. In a preferred embodiment, the immunoglobulin Fc region is It includes at least the immunoglobulin hinge region, the CH2 domain, and the CH3 domain, and is preferable In some embodiments, the CH1 domain is missing. In some embodiments, the immunoglobulin Fc region This is the human immunoglobulin Fc region.

[0173] In one embodiment, the class of immunoglobulin from which the heavy chain constant region originates is IgG (Igγ ) (γ subclass 1, 2, 3 or 4). Other classes of immunoglobulins, IgA ( Even if you use Igα, IgD (Igδ), IgE (Igε), and IgM (Igμ), Good. The selection of an appropriate immunoglobulin heavy chain constant region is described in U.S. Patent No. 5,541,087. This is discussed in detail in publications No. 5,726,044. The weight of certain immunoglobulins from a particular immunoglobulin class and subclass. The selection of the chain constant region sequence is considered to be within the range of skill levels in this field. The portion of the DNA construct encoding the immunoglobulin Fc region is preferably a hinged May At least a portion of the IgA, preferably the CH3 domain of Fc gamma or IgA, IgD , including at least a portion of the homologous domain in either IgE or IgM.

[0174] Furthermore, amino acid substitutions or deletions within the constant region of immunoglobulin heavy chains are permitted in this specification. The methods and compositions shown may be useful in carrying out the work. One example is the Fc receptor. Amino acids in the upper CH2 region to create Fc mutants with reduced affinity for the target. This would be the introduction of substitution (Cole et al. (1997) J. Immunol. 159: 3613).

[0175] In some embodiments, this disclosure refers to Sequence IDs 11, 13, 15, and 17 or those. Any one amino acid sequence of the bioactive fragment and at least 80%, 85%, 90%, 95% TβRII The present invention provides polypeptide fusion proteins. In some embodiments, the TβRII polypeptide is used. The fusion protein is one of SEQ ID NOs. 11, 13, and 15 or any of their bioactive fragments. or one amino acid sequence and at least 80%, 85%, 90%, 95%, 96%, 97%, It contains amino acid sequences that are 98% or 99% identical. In some embodiments, TβRII Lipeptide fusion proteins are one of the following: SEQ ID NO: 13 or any one of its bioactive fragments. Mino acid sequence and at least 80%, 85%, 90%, 95%, 96%, 97%, 98% also It contains an amino acid sequence that is 99% identical. In some embodiments, the TβRII polypeptide The fusion protein is one amino acid sequence of either SEQ ID NO: 50 or its bioactive fragment. and at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% It contains a single amino acid sequence. In some embodiments, a TβRII polypeptide fusion protein The substance contains at least one amino acid sequence from either SEQ ID NO: 51 or its bioactive fragment. Also, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% are identical. It contains a amino acid sequence. In some embodiments, the TβRII polypeptide fusion protein is distributed At least 80% of the amino acid sequence of either sequence number 52 or any one of its bioactive fragments, Amino acid sequences that are 85%, 90%, 95%, 96%, 97%, 98%, or 99% identical. Includes. In some embodiments, the TβRII polypeptide fusion protein is SEQ ID NO: 53 or any one of the amino acid sequences of its bioactive fragments and at least 80%, 85%, 9 Contains amino acid sequences that are 0%, 95%, 96%, 97%, 98%, or 99% identical. In one embodiment, the TβRII polypeptide fusion protein is SEQ ID NO: 54 or Any one amino acid sequence of the bioactive fragment and at least 80%, 85%, 90%, 95% Contains amino acid sequences that are 96%, 97%, 98%, or 99% identical. Some implementations In this state, the TβRII polypeptide fusion protein is sequence number 55 or its bioactive cleavage. Any one amino acid sequence of the piece and at least 80%, 85%, 90%, 95%, 96% , containing amino acid sequences that are 97%, 98%, or 99% identical. In some embodiments, T The βRII polypeptide fusion protein is either SEQ ID NO: 56 or its bioactive fragment. or one amino acid sequence and at least 80%, 85%, 90%, 95%, 96%, 97%, It contains amino acid sequences that are 98% or 99% identical. In some embodiments, TβRII Lipeptide fusion proteins have the amino acid sequence of SEQ ID NO: 20 or its bioactive fragment and a small amount At the very least, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% are identical. It contains a certain amino acid sequence.

[0176] In some embodiments, the fusion protein described herein is TGFβ1 and TG It has improved binding affinity to Fβ3. In some embodiments, at least 10 amino acids Fusion proteins containing linkers of a certain length (e.g., SEQ ID NOs: 11, 13, 15, and 50) A fusion protein having any one amino acid sequence of ~56 is a reference fusion protein. Compared to (for example, a fusion protein having the amino acid sequence of SEQ ID NO: 9), TGFβ1 and has improved binding affinity to TGFβ3. In some embodiments, the fusion protein Quality levels are less than 200 pM, less than 150 pM, less than 100 pM, less than 75 pM, and less than 50 pM. , or K less than 25 pM D Then it binds to TGFβ1. In some embodiments, fusion tan Protein levels are less than 75 pM, less than 70 pM, less than 60 pM, less than 50 pM, less than 40 pM. K levels below 35 pM, below 25 pM, below 15 pM, below 10 pM, or below 5 pM D in It binds to TGFβ3.

[0177] In some embodiments, any of the polypeptides disclosed herein is measurable In some embodiments, it inhibits TGFβ1 and / or TGFβ3. Peptides are determined using the reporter gene assay to have an IC of less than 1.0, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, 0.1, 0.08, 0.09, 0 .07, 0.06, 0.05, 0.04, 0.03 or 0.02 nM to inhibit TGFβ1. In some embodiments, the polypeptide is determined using the reporter gene assay to have an IC of less than 1.0, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0 .3, 0.2, 0.1, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04 50 to inhibit TGFβ3. In some embodiments the reporter gene assay is a CAGA reporter assay. In some embodiments the CAGA assay is based on a human lung cancer cell line transfected with the pGL3(CAGA)12 reporter plasmid (Dennle r et al., 1998, EMBO 17:3091 - 3100) and a control Renilla reporter plasmid (p RLCMV) for transfection efficiency. The CAGA motif is present in the promoter of TGFβ-responsive genes (e.g., PAI-1), so this reporter is generally used for factors that signal through SMAD2 and SMAD3 50 . See, for example, Example 2. . 5. Fusion polypeptide In some embodiments, the disclosure provides a TβRII-containing fusion polypeptide. The fusion polypeptide can be prepared according to any of the methods disclosed herein or known in the art . . . . .

[0178] . . . .

[0179] In some embodiments, any of the fusion polypeptides disclosed herein contains the following components: Includes: a) any of the TβRII polypeptides disclosed herein ("A"), b) this Any of the linkers disclosed in the specification ("B"), c) any different parts disclosed herein One of the following ("C"), and a linker ("X") as needed. In such embodiments Fusion polypeptides are formed in the following manner (from N-terminus to C-terminus): ABC or C -BA may be arranged. In such embodiments, the fusion polypeptide is as follows: Sequence (N-terminus to C-terminus): Can be sequenced in XABC or XCBA format. In some embodiments, the fusion polypeptide is A, B, and C (and optionally the leader) - Includes each of the amino acid sequences (for example, the amino acid sequence of SEQ ID NO: 23), and 100, 90, 80 , 70, 60, 50, 40, 30, 20, 10, 5, 4, 3, 2 or 1 or less additional It contains amino acids (but may also include further post-translational modifications such as pegylation).

[0180] In some embodiments, the fusion polypeptide is structured as follows: (from N-terminus to C-terminus) Includes a leader sequence (e.g., sequence number 23) arranged in XABC, and a fused poly The peptide contains 1, 2, 3, 4, or 5 amino acids between X and A. Some embodiments So, the fusion polypeptide is in the following form: (from N-terminus to C-terminus) XCBA, The fusion polypeptide includes a leader sequence (e.g., sequence number 23) positioned as X and The C contains 1, 2, 3, 4, or 5 amino acids. In some embodiments, fusion polyp Petit is a Lee arranged in the following form: (from N-terminus to C-terminus) XABC. The fusion polypeptide contains a der sequence (e.g., sequence number 23), and the fusion polypeptide has alanine between X and A. This includes. In some embodiments, the fusion polypeptide is in the following form: (N-terminus to C-terminus). (to the end) XCBA, including a leader array (e.g., sequence number 23), The fused polypeptide contains alanine between X and C. In some embodiments, the fused polypeptide The leader is arranged in the following format: (from N-terminus to C-terminus) XABC. The fusion polypeptide contains a sequence (e.g., sequence number 23), and has glycine between X and A. Contains bialanine. In some embodiments, the fusion polypeptide is in the following form: (N-terminus) A leader sequence (e.g., sequence number 23) arranged in XCBA from end to C-terminus. The fusion polypeptide contains glycine and alanine between X and C. Morphologically, fusion polypeptides are structured as follows: (from N-terminus to C-terminus) XAB- The fusion polypeptide includes a leader sequence (e.g., sequence number 23) positioned at C, The fusion polypeptide contains threonine between X and A. In some embodiments, the fusion polypeptide is as follows: Format: (N-terminus to C-terminus) XCBA, leader sequence (e.g., The fusion polypeptide, including column number 23), contains threonine between X and C.

[0181] In some embodiments, the fusion polypeptide is the TβRII polypeptide disclosed herein. Any of the tide amino acid sequences (e.g., SEQ ID NO: 18) and at least 85%, 90%, 9 The Tβ of this fusion polypeptide contains amino acid sequences that are 5%, 97%, or 99% identical. The RII polypeptide portion contains 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer additional polynomials. It contains additional amino acids (but may also include further post-translational modifications such as pegylation). In some embodiments, the fusion polypeptide is any of the linker sequences disclosed herein. (For example, Sequence ID No. 6) is at least 85%, 90%, 95%, 97%, or 99% identical. The amino acid sequence of this fusion polypeptide is 5, 4, 3, 2 or containing one or fewer additional amino acids (but including further post-translational modifications such as pegylation) (In some embodiments, the fusion polypeptide may be a heterogeneous part disclosed herein.) One of the sub-sequences (e.g., sequence number 20) and at least 85%, 90%, 95%, 97% The heterogeneous portion of this fusion polypeptide contains amino acid sequences that are % or 99% identical, and 2 Contains 5, 20, 15, 10, 5, 4, 3, 2 or 1 or fewer additional amino acids (however (This may include further post-translational modifications such as pegylation.) In some embodiments, fused poly The peptide is any of the TβRII polypeptide amino acid sequences disclosed herein (e.g.) For example, including SEQ ID NO: 18), the TβRII polypeptide portion of this fusion polypeptide is Contains 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1 or fewer additional amino acids (however (This may include further post-translational modifications such as pegylation.) In some embodiments, fused poly The peptide comprises one of the linker sequences disclosed herein (e.g., SEQ ID NO: 6). Furthermore, the linker portion of this fusion polypeptide contains 5, 4, 3, 2, or 1 or fewer additional A Contains mino acids (but may also include further post-translational modifications such as pegylation). Some fruits In the application form, the fusion polypeptide is any of the heterologous subsequences disclosed herein (for example) For example, including sequence number 20), the heterogeneous portion of this fusion polypeptide is 25, 20, 15, 1 Contains 0, 5, 4, 3, 2 or 1 or fewer additional amino acids (but further pegylation, etc.) (This may include post-translation modifications.)

[0182] In some embodiments, the disclosure is a fusion polypeptide, and a) the fusion polypeptide The TβRII polypeptide group consists of 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer copies. It contains additional amino acids (but may also contain further post-translational modifications such as pegylation). , any of the TβRII polypeptide amino acid sequences disclosed herein (e.g., sequence Amino acids that are at least 85%, 90%, 95%, 97%, or 99% identical to (number 18). Acid sequence; b) Addition of 5, 4, 3, 2 or 1 or fewer linker segments in the fusion polypeptide. It contains amino acids (but may also include further post-translational modifications such as pegylation), At least 85% of any of the linker sequences disclosed in the details (e.g., sequence number 6), amino acid sequences that are 90%, 95%, 97%, or 99% identical; and c) fusion polypeptides The different parts of the chido have 25, 20, 15, 10, 5, 4, 3, 2 or 1 or fewer additional parts. This specification includes mino acids (but may also include further post-translational modifications such as pegylation). Any of the heterologous subsequences disclosed in (e.g., SEQ ID NO: 20) and at least 85%, 9 Amino acid sequences that are 0%, 95%, 97%, or 99% identical; and d) as necessary The leader array (e.g., sequence number 23), consists of or essentially consists of these ( The fusion polypeptide is provided (and not necessarily in the order listed above). In some embodiments, This disclosure relates to a fusion polypeptide, a) the TβRII polypeptide of the fusion polypeptide. The portion contains 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1 or fewer additional amino acids. (However, this may include further post-translational modifications such as pegging), as disclosed herein. Any of the TβRII polypeptide amino acid sequences (e.g., SEQ ID NO: 18); b) fusion molecule The linker portion of the lipeptide contains 5, 4, 3, 2, or 1 or fewer additional amino acids. However, the following may include further post-translational modifications such as pegylation, as disclosed herein. c) any of the syncer sequences (e.g., SEQ ID NO: 6); and the heterogeneous portion of the fusion polypeptide. However, it contains 25, 20, 15, 10, 5, 4, 3, 2 or 1 or fewer additional amino acids. However, the differences disclosed herein may include further post-translational modifications such as pegging. Any of the seed subsequences (e.g., sequence number 20); and d) leader array as needed A column (for example, sequence number 23), consisting of or essentially consisting of (and not necessarily) The fusion polypeptide is provided (not in the order listed above).

[0183] In some embodiments, this disclosure a) the amino acid sequence of SEQ ID NO: 18 and at least 85% , 90%, 95%, 97%, or 99% identical amino acid sequences and 10, 9, 8, 7 , consisting of 6, 5, 4, 3, 2 or 1 or fewer additional amino acids (however, such as PEGylation) (May include further post-translational modifications), TβRII polypeptide moiety; b) Sequence ID 6 The amino acid sequence of is at least 85%, 90%, 95%, 97%, or 99% identical to that of It consists of an amino acid sequence and 5, 4, 3, 2 or 1 or fewer additional amino acids (however, c) Linker portion (which may include further post-translational modifications such as ligation); and sequence number The 20 amino acid sequences are identical by at least 85%, 90%, 95%, 97%, or 99%. The amino acid sequence and 25, 20, 15, 10, 5, 4, 3, 2 or 1 or fewer additional A heterogeneous molecule consisting of amino acids (but sometimes including further post-translational modifications such as pegylation). The part; and d) a leader array (e.g., sequence number 23) as needed, consisting of Alternatively, a fusion polypeptide that is essentially derived from these (and not necessarily in the order listed above) Provided. In some embodiments, this disclosure provides a) the amino acid sequence of SEQ ID NO: 18 and 10 , consisting of 9, 8, 7, 6, 5, 4, 3, 2 or 1 or fewer additional amino acids (however, (May include further post-translational modifications such as pegylation), TβRII polypeptide moiety; b ) From the amino acid sequence of SEQ ID NO: 6 and 5, 4, 3, 2 or 1 or fewer additional amino acids The linker part (which may include further post-translational modifications such as pegging); c) Amino acid sequence of Sequence ID No. 20 and 25, 20, 15, 10, 5, 4, 3, 2 or consists of one or fewer additional amino acids (but does not include further post-translational modifications such as pegylation). (This may also be the case), heterogeneous parts; and d) leader arrays as needed (e.g., array arrays) 23) consisting of or essentially consisting of these (and not necessarily in the order above) , provides a fusion polypeptide.

[0184] In some embodiments, the fusion protein does not include a leader sequence. The fusion protein has at least 85%, 90%, and 92% of the amino acid sequence of SEQ ID NO: 48. Contains amino acid sequences that are identical by %, 95%, 96%, 97%, 99%, or 100%. .

[0185] [ka]

[0186] In some embodiments, the present disclosure relates to a TβRII fusion polypeptide, which is an antibody or so The present invention provides polypeptides that do not contain antigen-binding sites. In some embodiments, polypeptides This refers to transforming growth factor beta superfamily ligands (e.g., TGF). Affinity for cytokines other than β1, TGFβ2, and / or TGFβ3 They do not bind. In some embodiments, the polypeptide is TGFβ1, TGFβ2 and / Alternatively, it does not bind with an affinity that allows it to be sensed by cytokines other than TGFβ3. Some implementations In this state, polypeptides interact with cytokines other than TGFβ1 and / or TGFβ3. It does not bind with known affinity. In some embodiments, the polypeptide is CD4, CD8 , CD25, CTLA-4, IL-10, TGFβ receptor, PD-1, PD-L1, PD -L2, RANK, RANKL, HER2 / neu, EGFR1, CD20, VEGF, TNF-α, TNFR2, FoxP3, CD80, CD86, IFN-α, IFN-β, IFN-γ, GITR, 4-1BB, OX-40, TLR1-10, ErbB-1, HE R1, ErbB-3 / HER3, ErbB-4 / HER4, IGFR, IGFBP, IG F-1R, PDGFR, FGFR, VEGFR, HGFR, TRK receptor, ephrin receptor Body, AXL receptor, LTK receptor, TIE receptor, angiopoietin 1, 2, ROR receptor Body, DDR receptor, RET receptor, KLG receptor, RYK receptor, MuSK receptor, IL βR, IlαR, TNTRSF, TRAIL receptor, ARTC1, alpha-actinin -4, Bcr-abl, B-RAF, caspase, beta-catenin, fibronectin , GPNMB, GDP-L, LDLR, HLA-A2, MLA-A11, HSP70, K IAA205, MART2, MUM-1, 2, 3, PAP, neo-PAP, NFYC, OGT, OS-9, pml-RAR alpha fusion protein, PRDX5, PTPRK, KRAS2, NRAS, HRAS, RBAF600, SIRT2, SNRPD1, SYT -SSX1 or -SSX2 fusion protein, triose phosphate isomerase, BAG E, BAGE-1, BAGE-2, 3, 4, 5, GAGE-1, 2, 3, 4, 5, 6, 7 , 8, GnT-V, HERV-K MEL, KK-LC, KM-HN-1, LAGE, L AGE-1, CAMEL, MAGE-1, MAGE-A2, MAGE-A3, MAGE- A4, MAGE-AS, MAGE-A6, MAGE-A8, MAGE-A9, MAGE- A10, MAGE-A11, MAGE-A12, MAGE-3, MAGE-B1, MAG E-B2, MAGE-B5, MAGE-B6, MAGE-C1, MAGE-C2, Mucin 1 (MUC1), MART-1 / Melan A (MLANA), gp100, gp100 / P me117(S1LV), tyrosinase (TYR), TRP-1, HAGE, NA-88 , NY-ESO-1, NY-ESO-1 / LAGE-2, SAGE, Sp17, SSX- 1, 2, 3, 4, TRP2-1NT2, carcinoembryonic antigen (CEA), kallikrein in)4, Mammaglobm-A, OA1, Prostate-Specific Antigen (PSA), Prostate gland-specific membrane antigen, TRP-1 / , 75, TRP-2, AIM-2, BING-4, CPS F, Cyclin D1, Ep-CAM, EpbA3, FGF-5, gp250, iCE) AFP, M-CSF, mdm-2, MUCI, p53(TP53), PBF, FRAME PSMA, RAGE-1, RNF43, RU2AS, SOX10, STEAP1, Saba Ibin (BIRCS), hTERT, telomerase, WT1, SYCP1, BRDT, S PANX, XAGE, ADAM2, PAGE-5, LIP1, CTAGE-1, CSAG E, MMA1, CAGE, BORIS, HOM-TES-85, AF15q14, HCA 66I, LDHC, MORC, SGY-1, SPO11, TPX1, NY-SAR-35 , FTHLI7, NXF2 TDRD1, TEX 15, FATE, TPTE, Estro Estrogen receptor (ER), androgen receptor (AR), CD40, CD30, CD20, C D19, CD33, CD4, CD25, CD3, CA 72-4, CA 15-3, CA 27-29, CA 125, CA 19-9, Beta-human chorionic gonadotropin, 1 -2 microglobulin, squamous cell carcinoma antigen, nerve-specific enolase (enoJase), thermal shock Protein gp96, GM2, salgramostim, CTLA-4, 707-AP, AR T-4, CAP-1, CLCA2, Cyp-B, HST-2, HPV protein, EBV Proteins, hepatitis B or C virus proteins, and / or HIV proteins It does not bind with an affinity that can be detected as such.

[0187] In some embodiments, the present disclosure is a TβRII fusion polypeptide, and TβRII It provides polypeptides that do not contain additional ligand-binding domains other than the main one. In the application form, the polypeptide consists of a TβRII domain and a heterogeneous portion (e.g., the Fc portion). It contains a linear amino acid sequence including, but this linear amino acid sequence does not contain any additional ligans. It does not include a do-binding domain. In some embodiments, the polypeptide is a TβRII domain. It also contains a linear amino acid sequence including an Fc portion, but this linear amino acid sequence is It also does not include additional ligand-binding domains. In some embodiments, this disclosure does not include TβRII A fusion polypeptide comprising multiple ligand-binding domains within a single linear amino acid sequence. The present invention provides polypeptides that do not contain TβRII. In some embodiments, the present invention provides polypeptides that do not contain TβRII. Lipeptides, wherein one or more consecutive linker molecules are present within a single linear amino acid sequence. The present invention provides a polypeptide that does not contain columns. In some embodiments, the polypeptide is a single column. Multiple consecutive glycine and / or serine linkers within a chain-like amino acid sequence (e.g., (GGGGS)n (where n=≧4) does not include a linker. This disclosure states that the heterogeneous portion is the Fc domain, and that only one consecutive linker is the Fc domain. The present invention provides a TβRII fusion polypeptide covalently bonded to yin. In some embodiments, A single consecutive linker contains (GGGGS)n linkers (where n=≧4). In the past, it consisted of such (GGGGS)n linkers.

[0188] 6. Nucleic acids and methods of production In certain embodiments, this disclosure may refer to other proteins and / or other TβRII proteins. They are either isolated from lipeptide species, or otherwise they are practically absent (e.g., few At the very least, 80%, 90%, 95%, 96%, 97%, 98%, or even 99% of them (i) Provide isolated and / or purified forms of TβRII polypeptide fusion protein TβRII polypeptides are generally produced by expression from recombinant nucleic acids. It will become that.

[0189] In certain embodiments, this disclosure relates to the coding sequence of the extracellular portion of the TβRII protein. The nucleic acid comprises a soluble TβRII polypeptide containing the following: This disclosure also relates to host cells containing such nucleic acids. Host cells are either prokaryotic or Eukaryotic cells may also be used. For example, the polypeptide of this disclosure can be used in bacterial cells such as E. coli. It may be expressed in insect cells (for example, using a baculovirus expression system). It may be expressed in yeast or mammalian cells. The host cells are known to those skilled in the art. Therefore, some embodiments of this disclosure are TβR Further relating to methods for producing polypeptide II.

[0190] In certain aspects, this disclosure may refer to the TβRII polypeptide (fragment) disclosed herein. Isolated and encoding any of the following (including functional variants and fusion proteins): / or provides recombinant nucleic acids. Sequence IDs 10, 12 and 14 are IgG Fc domains. It encodes a variant of the TβRII extracellular domain fused to the target nucleic acid. It may be in a single strand or in a double-stranded form. Such nucleic acids are DNA molecules. These nucleic acids may be, for example, TβRII poly In methods for producing peptides or as direct therapeutic agents (e.g., antisense, RN) (It can be used in AI or gene therapy approaches.)

[0191] In a particular embodiment, the target nucleic acid encoding the TβRII polypeptide is SEQ ID NO: 10 It is further understood that this includes nucleic acids that are variants of 12 and 14. The column represents points of one or more nucleotide substitutions, additions, or deletions, such as allele variants. It contains different arrays.

[0192] In certain embodiments, this disclosure relates to Sequence IDs 10, 12, and 14 and at least 80 %, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical Provides an isolated or recombinant nucleic acid sequence. In certain embodiments, this disclosure provides , at least 80%, 85%, 90%, 95%, 96% of sequence number 12 or a fragment thereof, Isolated or recombinant nucleic acid compounds that are 97%, 98%, 99%, or 100% identical. Provides a sequence of nucleic acid sequences complementary to sequence numbers 10, 12, and 14, and sequence number 1 Those skilled in the art will understand that variants 0, 12, and 14 are also within the scope of this disclosure. It is expected. In further embodiments, the nucleic acid sequences of the present disclosure may be isolated and recombinant. It may be a and / or fused with a heterologous nucleotide sequence, It may be present in a DNA library.

[0193] In other embodiments, the nucleic acids of this disclosure are the nucleic acids specified in SEQ ID NOs: 10, 12, and 14. The rheotide sequence, the complementary sequences of sequence numbers 10, 12, and 14, or fragments thereof, and high This also includes nucleotide sequences that hybridize under trigger conditions, as discussed above. Changing the appropriate stringency conditions to promote DNA hybridization What can be done is expected to be easily understood by those skilled in the art. For example, 6.0 × at approximately 45°C. Hybridization with sodium chloride / sodium citrate (SSC), followed by 5 It will be possible to perform 2.0 × SSC cleaning at 0°C. For example, in the cleaning step The salt concentration was changed from a low stringency of approximately 2.0 × SSC at 50°C to approximately 0.2 × SSC at 50°C. You can choose from SSCs with high stringency. In addition, the washing step The temperature ranges from low stringency conditions at room temperature (approximately 22°C) to high stringency conditions at approximately 65°C. The conditions can be increased to lingency conditions. Both temperature and salt can be changed, The temperature or salt concentration may be kept constant while the other variable is changed. In embodiments, the present disclosure describes washing with 6×SSC at room temperature, followed by washing with 2×SSC at room temperature. This invention provides nucleic acids that hybridize under low stringency conditions.

[0194] Nucleic acids, such as those shown in Sequence IDs 10, 12, and 14, are due to the degeneracy of the genetic code. Different, isolated nucleic acids are also within the scope of this disclosure. For example, a number of amino acids, one of them It is specified by many triplets. The same amino acid, or synonyms (for example, CA U and CAC are codons that define histidine, and the codons that define the protein It can also result in "silent" mutations that do not affect the no-acid sequence. However, the target DNA sequence polymorphisms, which lead to changes in the amino acid sequence of proteins, exist between mammalian cells. It is expected that one or more nucleotides of nucleic acids that code for a specific protein will be found. These divergences (about 3-5% or less of nucleotides) contribute to natural allele variation. Those skilled in the art will understand that this can occur among individuals of a given species. This includes any and all such nucleotide diversity and the resulting a Mino acid polymorphism is within the scope of this disclosure.

[0195] A corresponding mutant based on the long isoform of TβRII has a 25-amino acid insertion. It codes and also encodes a conservative Val-Ile substitution at the adjacent position on the C-terminal side of the insertion. It is expected that those skilled in the art will understand that this will include a nucleotide sequence. TβR Corresponding transformation based on either the long (A) or short (B) isoform of II The variant performs a 108-nucleotide insertion (SEQ ID NO: 41) that encodes a 36-amino acid insertion. It contains the same position as the naturally occurring TβRII isoform C, but in a different location. It is also expected that this will involve the inclusion of heteronucleotide sequences.

[0196] In certain embodiments, recombinant nucleic acids are used in one or more expression constructs. It can be operably ligated to the nodal nucleotide sequence. The regulatory nucleotide sequence is Generally, it is expected to be suitable for host cells used in expression. A wide variety of suitable types. Suitable expression vectors and regulatory sequences are known in the art for various host cells. There is. Typically, the one or more regulatory nucleotide sequences are promoter sequences, Leader or signal sequence, ribosome binding site, transcription start and termination sequences, translation start and termination sequences, and may include enhancer or activator sequences, however these Not limited to, constitutive or inductive promoters known in the art. However, this disclosure intends for promoters to be naturally occurring promoters. Often, hybrid promotions combine elements of one or more promoters. It may also be a construct. Expression constructs reside on episomes such as plasmids in cells. Alternatively, the expression construct may be inserted into the chromosome. In a preferred embodiment, The current vector contains selection marker genes to enable the selection of transformed host cells. Contains. The selected marker gene is well known in the art and is used in host cells It will depend on the circumstances.

[0197] In certain embodiments disclosed herein, the nucleic acid in question is a TβRII polypeptide. A nucleotide sequence that controls, which is operably linked to at least one regulatory sequence. The expression vector contains the nucleotide sequence. The regulatory sequence is part of this technology. It is recognized in the field and selected to direct the expression of TβRII polypeptide. Therefore, the term "regulatory element" refers to promoters, enhancers, and other expression control elements. Includes. Exemplary regulatory sequences are found in Goddel; Gene Expression Technology: Methods i It is described in *Enzymology*, Academic Press, San Diego, CA (1990). For example, when operably ligated to a DNA sequence, various types of molecules control the expression of that DNA sequence. Using one of the diverse expression regulatory sequences in these vectors, TβRII poly It is possible to express DNA sequences that encode peptides. Such useful expression control The sequences include, for example, the early and late promoters of SV40, the tet promoter, and adeno. Virus or cytomegalovirus early promoter, RSV promoter, lac The system, trp system, TAC or TRC system, expression is directed by T7 RNA polymerase. The T7 promoter, the main operator and promoter region of the phage lambda, fd Regulatory regions of coat proteins, 3-phosphoglycerate kinase or other glycosulfating enzymes, Promoters of acid phosphatases, e.g., Pho5, promoter of yeast α-conjugation factor, Baculovirus polyhedron promoters, and prokaryotic or eukaryotic cells or their Other sequences known to control the expression of viral genes, as well as various combinations thereof. This includes the design of the expression vector, the selection of host cells to be transformed, and / or expression. It is understood that this may depend on factors such as the type of protein that is desired to be extracted. It should be so. Furthermore, the number of copies of the vector, the ability to control that number of copies, and the vector The expression of any other protein encoded by, for example, an antibiotic marker, This needs to be considered.

[0198] Recombinant nucleic acids included in this disclosure are cloned genes or portions thereof, derived from prokaryotic sources. In either or both of the following cells: Recombinant Tβ can be produced by ligating it into a vector suitable for expression. Expression vehicles for RII polypeptide production include plasmids and other vectors. For example, vectors suitable for expression in prokaryotic cells such as E. coli are of the following types. Plasmids include: pBR322-derived plasmid, pEMBL-derived plasmid, pEX Plasmid derived from pBTac, and plasmid derived from pUC.

[0199] Some mammalian expression vectors facilitate vector transmission in bacteria, particularly in prokaryotes. Both the sequence of elements and one or more eukaryotic transcription units expressed in eukaryotic cells. It contains: pcDNAI / amp, pcDNAI / neo, pRc / CMV, pSV2 gpt, pSV2neo, pSV2-dhfr, pTk2, pRSVneo, pMSG, pSVT7, pko-neo, and pHyg-derived vectors transfect eukaryotic cells Examples of mammalian expression vectors suitable for this purpose. Some of these vectors are prokaryotic cells. To promote replication and drug resistance selection in both eukaryotic and oocytes, pBR322, etc. Modified with sequences from bacterial plasmids. Alternatively, bovine papillomavirus (BPV) -1) or Epstein-Barr virus (pHEBo, derived from pREP and p205 Using derivatives of viruses such as ) for transient protein expression in eukaryotic cells This is possible. Examples of other virus (including retrovirus) expression systems are listed below for gene therapy delivery. It can be found in the system description. For plasmid preparation and transformation of host organisms. Various methods used are well known in this field. For other suitable expression systems and general recombination procedures, see Molecular C Loning A Laboratory Manual, 3rd edition, edited by Sambrook, Fritsch and Maniatis (Cold See Spring Harbor Laboratory Press, 2001. In some cases, It is desirable to express recombinant polypeptides using a baculovirus expression system. There are such baculovirus expression systems, for example, pVL-derived vectors ( (pVL1392, pVL1393 and pVL941), pAcUW-derived vectors ( For example, pAcUW1), and pBlueBac-derived vectors (e.g., β-gal-containing vectors). One example is the pBlueBac III.

[0200] In certain embodiments, a vector, for example, a Pcmv-Script vector (St ratagene, La Jolla, Calif.), pcDN4 vector (Invi (Trogen, Carlsbad, Calif.) and pCI-neo vector (Pr Omega, Madison, Wisc.) is a target TβRII poly(TβRII) It will be designed for peptide production. In a preferred embodiment, the vector is HEK -293 cells will be designed for the production of the target TβRII polypeptide. It is expected that the target gene construct will be used to produce cells grown in culture for purification. Inducing the expression of the target TβRII polypeptide in cells, for example, a fusion protein It can produce proteins that include mutant proteins.

[0201] This disclosure provides coding sequences (for example) for one or more target TβRII polypeptides. For example, a host transfected with a recombinant gene containing sequence numbers 10, 12, or 14. This also applies to cells. The host cell can be either a prokaryotic or eukaryotic cell. For example, The TβRII polypeptide disclosed in the specification is used in bacterial cells such as E. coli. It may be expressed in insect cells (for example, using a baculovirus expression system). It may be expressed in yeast or mammalian cells. Cells are known to those skilled in the art.

[0202] Therefore, this disclosure further relates to a method for producing the target TβRII polypeptide. For example, an expression vector encoding the TβRII polypeptide was transfected. The host cells are cultured under conditions suitable for inducing the expression of the TβRII polypeptide. It is possible to obtain TβRII from a mixture of cells and a culture medium containing TβRII polypeptide. The I polypeptide can be secreted and isolated. Alternatively, the TβRII polypeptide can be extracted. To retain the protein in the cytoplasm or membrane fraction, to collect the cells, to lyse them, and to isolate the protein. This can be done. Cell cultures include host cells and culture medium. Culture media suitable for cell culture are provided by this technology. This is well known in the field. Ion exchange chromatography, gel filtration chromatography Ultrafiltration, electrophoresis, and antibodies specific to specific epitopes of TβRII polypeptides are used. Immunoaffinity purification, as well as fusion with TβRII polypeptide, is performed by Domaine. Affinity purification using a drug that binds to protein (for example, using a protein A column) To purify proteins, including the TβRII-Fc fusion (which can be purified) Using methods known in the art, the target TβRII polypeptide is cultured in cell culture. It can be isolated from soil, host cells, or both. In preferred embodiments, TβRI Polypeptide I is a fusion protein containing a domain that facilitates its purification. For example, The purification process involves, for example, a series of ions containing three or more of the following in any order. The following steps can be performed by the Lamb chromatography process: Protein A chromatography Fee, Q Sepharose chromatography, Phenylen Sepharose chromatography Size exclusion chromatography and cation exchange chromatography. Viral filtration. Purification can also be carried out using buffer exchange.

[0203] In another embodiment, the poly(N-terminus) of a desired portion of the recombinant TβRII polypeptide (His) / Fusion gene encoding purified reader sequences such as enterokinase cleavage site sequences. Denshi, Ni 2+ Expression by affinity chromatography using metal resin This can enable the purification of the fusion protein. Then, the purified leader sequence is entered By removing it with kinase treatment, purified TβRII polypeptide can be obtained. (For example, Hochuli et al., (1987) J. Chromatography Vol. 411: p. 177; See also Janknecht et al., PNAS USA, Vol. 88, p. 8972.

[0204] The methods for creating fusion genes are well known. Essentially, it involves combining different polypeptide sequences. The ligation of various DNA fragments is performed according to conventional techniques, smoothing for ligation. Using terminal or offset terminals, restriction enzyme digestion is used to obtain the appropriate terminal. Then, by appropriately utilizing the embedding of the attached ends, alkali is used to avoid undesirable linkages. This is performed using phosphatase treatment and enzymatic ligation. In this embodiment, the fusion gene is synthesized using a conventional method including an automated DNA synthesis device. This can be done. Alternatively, PCR amplification of gene fragments can be performed between two consecutive gene fragments. This can be done using an anchor primer that creates a target overhang, and then, These overhangs can be annealed to generate chimeric gene sequences (example) For example, Current Protocols in Molecular Biology, edited by Ausubel et al., John Wiley & S. (See ons: 1992).

[0205] 7. Changes in Fc-fusion proteins This application further proposes TβRII-Fc fusion proteins having modified or mutated Fc regions. Such antibodies and Fc fusion proteins are used, for example, in antigen-dependent cell-mediated cytotoxicity (A). Modulation of effector functions such as DCC and complement-dependent cell injury (CDC) This may be useful. In addition, the modification improves the stability of antibodies and Fc fusion proteins. It is possible. Amino acid sequence variants of antibodies and Fc fusion proteins can be appropriate nuclei. It is prepared by introducing an ocidal change into DNA or by peptide synthesis. The unagi variant is, for example, the amino acid of the antibody and Fc fusion protein disclosed herein. Deletion of residues from a sequence, and / or insertion of residues into the amino acid sequence, and / or This includes substitution of residues within the aforementioned amino acid sequence, provided that the final construct has the desired properties. Any combination of deletions, insertions, and substitutions is performed to reach the final construct. (Amino acids) The changes include altering the number or location of glycosylation sites in antibody and Fc fusion. It is possible to alter the post-translational processes of proteins.

[0206] Antibodies and Fc fusion proteins with reduced effector function were described by Bluestone et al. The listed Ala-Ala mutations (WO94 / 28027 and WO98 / 47531) See also: Xu et al., 2000, Cell Immunol vol. 200; pp. 16-26. By introducing changes in the amino acid sequence, including but not limited to (the desired amino acid), the production of Therefore, in certain embodiments, the Ala-Ala mutation may be included. Using the Fc fusion protein of this disclosure, which has mutations within the constant region, effectors -The function can be reduced or eliminated. According to these embodiments, the antibody and F The c fusion protein is either a mutation to alanine at position 234, or at position 235. This may include mutations to alanine, or combinations thereof. In one embodiment, an antibody or The Fc fusion protein contains the IgG4 framework, in this case the Ala-Ala mutation. The mutation is a mutation from phenylalanine to alanine at position 234, and / or This represents a mutation from leucine to alanine at position 235. In another embodiment... The antibody or Fc fusion protein contains the IgG1 framework, in this case, Al a-Ala mutation is a mutation from leucine to alanine at position 234, and / Alternatively, it would represent a mutation from leucine to alanine at position 235. The Fc fusion protein contains a point mutation K322A in the CH2 domain, among other mutations. Natural mutations can be possessed as an alternative or additional mutation (Hezareh et al., 2001 J) Virol. 75:12161-8).

[0207] In certain embodiments, an antibody or Fc fusion protein is used to induce complement-dependent cell toxicology (CDC). ) can be modified to either enhance or inhibit it. Modulation of C activity involves one or more amino acid substitutions, insertions, or deletions in the Fc region. This can be achieved by introducing (see, for example, U.S. Patent No. 6,194,551). (Please be illuminated). Alternatively, or in addition, it is possible to introduce cysteine ​​residues into the Fc region. This enables the formation of interchain disulfide bonds within this region. The resulting homodimers exhibit improved or decreased internalization capacity and / or complement-mediated effects. It can result in an increase or decrease in cell death. Caron et al., J. Exp Med. 176: Pages 1191-1195 (1992) and Shopes, BJ Immunol. Vol. 148:29 Pages 18-2922 (1992), WO99 / 51642, Duncan and Winter Nature Volume 322: pp. 738-734 (1988); U.S. Patent No. 5,648,260, U.S. Special See Permit No. 5,624,821 and WO94 / 29351.

[0208] 8. Screening assay In certain embodiments, the present invention relates to the transmission of TGFβ1, TGFβ3, and TβRII signals. TβR for identifying compounds (drugs) that are agonists or antagonists in the drug pathway Regarding the use of II polypeptides (e.g., soluble TβRII polypeptide). The compounds identified by leaning were tested in vitro for TGFβ1 and Their ability to modulate TGFβ3 signaling activity can be assessed. If necessary, these compounds will be further tested in animal models to enable tissue formation in vivo. Their ability to modulate length can be assessed.

[0209] By targeting TGFβ1, TGFβ3, and TβRII polypeptides, tissue formation There are a great many approaches to screening for therapeutic agents that modulate length. In a particular embodiment, high-throughput screening of compounds is performed to TGF It is possible to identify drugs that disrupt β1, TGFβ3, or TβRII-mediated cell signaling. In certain embodiments, the assay involves TGFβ1 of the TβRII polypeptide. This involves screening compounds that specifically inhibit or reduce binding to TGFβ3 and This is done for determination. Alternatively, an assay is used to determine the TGF of TβRII polypeptide. Compounds that enhance binding to β1 or TGFβ3 can be identified. Further experiments In the application method, the compound is interacted with TGFβ1, TGFβ3, or TβRII polypeptides. They can be identified by the abilities they employ.

[0210] Various assay formats are expected to be sufficient and are clearly described herein in light of this disclosure. What is not described is nevertheless expected to be identified by those skilled in the art. As described in the document, the test compound (drug) of the present invention is used in any combinatorial chemistry It may also be produced by the Tree method. Alternatively, the target compound may be produced in vivo or in These may be naturally occurring biomolecules synthesized in vitro. The modulation of tissue growth. The compounds (drugs) tested for their ability to act as inhibitors include, for example, bacteria. It can be produced by yeast, plants, or other organisms (e.g., natural products), and is also produced chemically. This can also be done (for example, small molecules including peptide mimes), or by recombinant It may also be produced by this invention. The test compound intended by this invention is a non-peptidyl organic compound. This includes molecules such as seeds, peptides, polypeptides, peptide mimes, sugars, hormones, and nucleic acid molecules. In certain embodiments, the test drug is an organic molecule having a molecular weight of less than approximately 2,000 daltons. He is a child.

[0211] The test compound of the present invention can be provided as a single, individual entity, or as a combination. For more complex libraries, such as those created by natural chemistry... These libraries can be provided. These libraries include, for example, alcohols and halides. Organic compounds of the following classes: methyl, amine, amide, ester, aldehyde, ether, and other classes. This may include presenting the test compound to the test system in an isolated form. or, especially in the initial screening step, it may be a compound mixture. If necessary, the compound may be derivatized using other compounds as needed. However, it may have a derivatizing group that facilitates the isolation of the compound. A non-limiting example of a derivatizing group is... For example, biotin, fluorescein, digoxigenin, green fluorescent protein, isotopes, poly Histidine, magnetic beads, glutathione S transferase (GST), photoactivatable Examples include crosslinking agents, or combinations thereof.

[0212] Many drug screening programs test libraries of compounds and natural extracts. In this context, high-throughput assays maximize the number of compounds investigated within a given period. Desirable for conversion. Those that can be derived using purified or semi-purified proteins, etc. Assays performed in cell-free systems allow for the rapid development of changes in molecular targets mediated by the test compound. Because they can be generated in a way that allows for raw and relatively easy detection, It is often preferred as a "primary" screen. Furthermore, the cytotoxicity of the test compound or The effect of bioavailability can generally be ignored in in vitro systems. Instead, the assay is performed between TβRII polypeptide and TGFβ1 or TGFβ3. The focus is on the drug's effect on molecular targets, which can manifest as changes in binding affinity. It can happen.

[0213] For illustrative purposes only, the exemplary screening assay of the present invention uses the target compound, Isolated and purified TβRI that can normally bind to TGFβ1 or TGFβ3 The compound is brought into contact with the TβRII polypeptide. Then, the T Add a composition containing a βRII ligand. TβRII / TGFβ1 or TβRI Detection and quantification of the I / TGFβ3 complex involves TβRII polypeptide and TGFβ1 or The effectiveness of the compound will be determined in terms of inhibiting (or enhancing) complex formation with TGFβ3. This becomes a means to achieve this. The effectiveness of the compound is determined by data obtained using various concentrations of the test compound. Furthermore, it can be evaluated by generating a dose-response curve. In addition, a control assay is performed. This also allows us to obtain a baseline for comparison. For example, in a control assay, isolation A composition containing purified TGFβ1 or TGFβ3 and TβRII polypeptide. When added to a substance, the formation of TβRII / TGFβ1 or TβRII / TGFβ3 complexes is tested. The compound is quantified in the absence of the compound. Generally, the order in which the reactants are mixed may be changed, and they may be mixed simultaneously. It is expected that this may be understood. Furthermore, as an alternative to purified protein... Using cell extracts and lysates, a suitable cell-free assay system can be obtained. .

[0214] Complex formation between TβRII polypeptide and TGFβ1 or TGFβ3 is performed in various ways. It can be detected by various methods. For example, the modulation of complex formation can be detected, for example, Proteins that can be labeled, for example, radiolabeled (e.g., 32 P, 35 S, 1 4 C or 3 H), fluorescently labeled (e.g., FITC) or enzyme-labeled TβR Using the II polypeptide or TGFβ1 or TGFβ3, by immunoassay It can be quantified by chromatographic detection or other means.

[0215] In certain embodiments, the present invention relates to the TβRII polypeptide and its binding protein. When measuring the degree of interaction between two elements, either directly or indirectly, using a fluorescence polarization assay, Furthermore, we intend to use fluorescence resonance energy transfer (FRET) assays. In addition, optical waveguides (e.g., PCT Publication WO96 / 26432 and U.S. Patent No. 5,677,196) Based on surface plasmon resonance (SPR), surface charge sensors, and area force sensors. Other detection methods, such as those mentioned above, are compatible with many embodiments of the present invention.

[0216] Furthermore, the present invention interferes with the interaction between the TβRII polypeptide and its binding protein. Also known as a "two-hybrid assay" for identifying drugs that enhance or increase The use of interaction capture assays is intended. For example, U.S. Patent No. 5,283,31 No. 7; Zervos et al. (1993) Cell Vol. 72: pp. 223-232; Madura et al. (1993) ) J Biol Chem Vol. 268: pp. 12046-12054; Bartel et al. (1993) Biote Chniques, Vol. 14: pp. 920-924; and Iwabuchi et al. (1993), Oncogene, Vol. 8. (See pages 1693-1696). In certain embodiments, the present invention relates to TβRII Compounds that dissociate the interaction between polypeptides and their binding proteins (e.g., small molecules) We intend to use a reverse two-hybrid system to identify the offspring (or peptide). For example Vidal and Legrain, (1999) Nucleic Acids Res Vol. 27: 919-929 Pages; Vidal and Legrain, (1999) Trends Biotechnol Vol. 17: pp. 374-371 ;and U.S. Patent No. 5,525,490, No. 5,955,280 and No. 5, Please refer to issues 965 and 368.

[0217] In certain embodiments, the target compound is TβRII or TGFβ1 or They are identified by their ability to interact with TGFβ3 polypeptides. Compounds and TβR The interaction between TGFβ2 or TGFβ1 or TGFβ3 polypeptides is covalent. It may be, or it may be non-covalent. For example, such an interaction i It can be identified at the protein level using in vitro biochemical methods (Ja Koby WB et al., 1974, Methods in Enzymology, Vol. 46: p. 1). In certain cases This involves using compounds in mechanism-based assays, for example, TGFβ1 or TGFβ3 or T This assay is used to detect compounds that bind to βRII polypeptides, and for screening purposes. This can include solid-phase or fluid-phase coupling events. Alternatively, TGFβ1 Alternatively, a reporter system (e.g.,) can use a gene encoding TGFβ3 or TβRII polypeptide. For example, β-galactosidase, luciferase, or green fluorescent protein can be used to stimulate cells. Infect and preferably perform high-throughput screening on the library. Screening can be performed on a broader scale, or on individual members of the library. Other mechanism-based binding assays, such as those that detect changes in free energy. You may use (i). The binding assay involves using a label fixed to a well, bead, or tip. Targets captured by immobilized antibodies, or by capillary electrophoresis This can be done using segmented targets. The bound compounds are usually analyzed by colorimetric analysis or Detection can be performed using fluorescence or surface plasmon resonance.

[0218] In certain embodiments, the present invention relates to TGFβ1-mediated or TGFβ3-mediated cell signaling The present invention provides methods and agents for modulating (stimulating or inhibiting) these substances. Therefore, any identified compound can be used in vitro in whole cells or tissues. Alternatively, testing in vivo modulates TGFβ1 or TGFβ3 signaling. The ability to perform these actions can be confirmed. Various methods known in the art can be used to verify these capabilities. It can be used for this purpose.

[0219] 9. Exemplary Therapeutic Uses When used herein, a therapeutic agent that “prevents” a disorder or condition is a statistical sample. In the treatment, the occurrence of defects or conditions in the treated sample compared to the untreated control sample. A compound that reduces the rate, or one of the impairments or conditions compared to an untreated control sample. A compound that delays the onset of multiple symptoms or reduces the severity of the aforementioned symptoms. It refers to.

[0220] The terms “treatment,” “taking action,” “mitigation,” and similar terms are defined herein. The term is generally used to mean obtaining the desired pharmacological and / or physiological effect. to improve, alleviate, and / or improve one or more symptoms of the condition being treated. It is also sometimes used to refer to reducing the severity of the aforementioned symptoms. The effect is To completely or partially delay the onset or recurrence of a disease, condition, or its symptoms. From this perspective, it may also be preventative, as well as / or disease or condition and / if Is it from the perspective of partial or complete cure of adverse effects caused by a disease or condition? It may also be therapeutic. When used herein, "treatment" refers to mammals, especially rhinoceros beetles. (a) having a predisposition to the disease or condition However, in subjects who have not yet been diagnosed with the disease or condition, (b) To prevent the occurrence of a condition, (b) To suppress a disease or condition (for example, (c) to prevent its progression, or to alleviate a disease or condition (for example, disease To cause regression of an illness or condition, or to bring about improvement in one or more symptoms. ) includes.

[0221] The terms “patient,” “subject,” or “individual” are used synonymously in this specification and refer to humans or This refers to either a non-human animal or a mammal, e.g., a human or a non-human primate. Species, laboratory animals, domestic animals (including cattle, pigs, camels, etc.), companion animals (e.g., dogs, This includes cats, other domesticated animals, and rodents (e.g., mice and rats). In a particular embodiment, the patient, subject, or individual is a human being.

[0222] This disclosure describes diseases or conditions related to TGFβ superfamily members as described above. TβRII polypeptides containing TβRII-Fc fusion proteins (hereinafter referred to herein) By administering an effective dose of a "therapeutic agent" (collectively referred to as a "therapeutic agent") to the target, the treatment or prevention is performed. A method is provided. In some embodiments, the disease or condition is TGFβ1 or TGFβ3 It is related to abnormal regulation of signal transduction. To treat certain cardiovascular or vascular disorders. Methods and compositions for treating or preventing cancer are also provided. In addition, this disclosure is provided for treating or preventing cancer The present disclosure provides methods and compositions for treating or The present invention provides methods and compositions for prevention.

[0223] In particular, the polypeptide therapeutic agents of this disclosure are used for the treatment or prevention of chronic vascular or cardiovascular diseases. Useful. Examples of this type of disorder include heart disease (myocardial disease, myocardial infarction, angina pectoris, and This includes heart valve disease; kidney disease (chronic glomerulinitis, diabetic nephropathy, and lupus-related conditions). Disorders associated with atherosclerosis or other types of arteriosclerosis (including nephritis) Thrombotic disorders (including stroke, cerebral hemorrhage, subarachnoid hemorrhage, angina pectoris, and renal arteriosclerosis); brain Thrombosis, including thrombotic bowel necrosis; complications of diabetes (diabetes-related kidney disease, cataracts, diabetes). Related kidney disease, diabetes-related neuropathology, diabetes-related gangrene, and diabetes-related Including chronic infections; vascular inflammatory disorders (systemic lupus erythematosus, rheumatoid arthritis, joint inflammation) Arteritis, large cell arteritis, Kawasaki disease, Takayasu arteritis, Churg-Strauss syndrome, and Ch. Schönlein purpura; diabetic vascular disorders; and cardiac disorders, such as congenital Cardiac disease, cardiomyopathy (e.g., dilated, hypertrophic, and restrictive cardiomyopathy), and congestive heart failure are among the conditions that can be diagnosed. However, these are not the only examples. Further examples of disorders include hereditary hemorrhagic capillary syndrome. Hypertelangiectasia (HHT), Marfan syndrome, Loeys-Dietz syndrome, Familial thoracic aortic aorta Aneurysm syndrome, tortuosity syndrome, pre-eclampsia, and restenosis are examples, but are not limited to these. do not have.

[0224] It can be administered alone to target TβRII polypeptides, or to TGFβ-related heart blood One or more drugs or therapeutic agents useful for treating tubal disorders and / or conditions It can be administered in combination with a drug, for example, a therapeutic agent. In certain embodiments So, the second drug or treatment modality is angioplasty, beta-blockers, antihypertensive drugs, or cardiac stimulants. Drugs, antithrombotic drugs, vasodilators, hormone antagonists, endothelin antagonists, Luxium channel blockers, phosphodiesterase inhibitors, angiotensin type 2 antagonists Selected from one or more nistrogens and / or cytokine blockers / inhibitors ru.

[0225] In particular, the polypeptide therapeutic agents of this disclosure are useful for the treatment or prevention of cancer (tumors). The terms "cancer" and "cancerous" refer to mammals typically characterized by uncontrolled cell growth / proliferation. Refers to or describes a physical condition in an animal. Cancer, or neoplastic Examples of disorders include cancer, lymphoma, blastoma, sarcoma, and leukemia, but are not limited to these. It is not determined. More specific examples of such cancers include squamous cell carcinoma, peritoneal carcinoma, and hepatocyte cancer. Cancer, digestive cancer, pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, Patoma, breast cancer, colon cancer, colorectal cancer, endometrial or uterine cancer, salivary gland cancer, kidney cancer Hmm, prostate cancer, vulvar cancer, thyroid cancer, liver cancer, stomach cancer, intestinal cancer, skin cancer, bone cancer, Stomach cancer, melanoma, and various types of head and neck cancer (including head and neck squamous cell carcinoma) Examples include neoplastic disorders and related conditions such as esophageal cancer, theca cell tumor, and masculinization. Tumors, endometrial hyperplasia, endometriosis, fibrosarcoma, choriocarcinoma, nasopharyngeal carcinoma, pharyngeal carcinoma, hepatoblastoma, Posi sarcoma, skin cancer, hemangioma, cavernous hemangioma, hemangioblastoma, retinoblastoma, astrocytoma, collagen Scars, schwannomas, oligodendrogliomas, medulloblastomas, neuroblastomas, rhabdomyosarcomas, osteogenic sarcomas, leioblastomas Myosarcoma, urinary tract cancer, Wilms' tumor, renal cell carcinoma, prostate cancer, and abnormal angiogenesis associated with nevus. Examples include, and Meigs syndrome. Particularly suitable for treatment with the therapeutic agents described herein. Cancers that have elevated TβRII levels detectable in the tumor or serum This involves increasing the expression level of TGFβ1 or TGFβ3 or increasing their biological activity. To be metastatic, or at risk of becoming metastatic, or a combination of these. It may be characterized by one or more of the following:

[0226] In certain embodiments of such methods, one or more polypeptide therapeutic agents They can be administered together (simultaneously) or at different times (sequentially). In addition, polypeptide therapeutics can be used for the treatment of cancer or to inhibit angiogenesis. It can be administered together with other types of compounds.

[0227] In certain embodiments, the method covered by this disclosure can be used on its own. The target method is other conventional anticancer treatments aimed at treating or preventing proliferative disorders (e.g., tumors). It may be used in combination with therapeutic approaches. For example, such methods may be used to prevent the onset of the disease. Prevention of cancer, prevention of cancer recurrence and metastasis after surgery, and other conventional cancer treatments It can be used as an adjuvant. This disclosure expands the possibilities for conventional cancer treatment (e.g., chemical The effectiveness of (medical therapy, radiotherapy, phototherapy, immunotherapy, and surgery) is assessed by target polypeptide It is recognized that this can be enhanced by the use of cytotoxic agents.

[0228] A wide variety of conventional compounds have been proven to possess anti-tumor or anti-cancer activity. These compounds are used in chemotherapy to reduce solid tumors, metastases, and further growths. To prevent growth, or malignant cells in leukemic or myelopathic lesions It has been used as a pharmaceutical agent to reduce the number of tumors. Chemotherapy is used for various types of malignancies. While they remain effective in treating lesions, many anti-cancer compounds have undesirable side effects. It induces. When two or more different treatments are combined, those treatments They work synergistically, enabling a reduction in the dosage of each of these treatments, resulting in a higher It has been proven that the harmful side effects caused by each compound can be reduced by adjusting the dosage. In other cases, there are two or more malignant lesions for which a certain treatment is ineffective. It may also respond to combination therapy with different treatments.

[0229] The therapeutic agents disclosed herein may be used in combination with other conventional anti-cancer agents, either simultaneously or sequentially. Subsequently, when administered in either case, such therapeutic agents enhance the therapeutic effect of anti-cancer drugs. It can strengthen or overcome the cell's resistance to such anti-cancer agents. Yes, it is possible. This allows for a reduction in the dosage of anti-cancer drugs, thereby reducing undesirable side effects. Use is reduced, or the effectiveness of anti-cancer agents in resistant cells is restored.

[0230] According to this disclosure, polypeptide therapeutic agents described herein are used for the treatment of diseases and other purposes. It can be used in combination with the composition and procedure of TβRII polyp Surgical treatment, radiation therapy, or chemotherapy combined with cytoplasmic remedy (Cytoplasmic remedy) can be used to treat tumors as conventionally. And, thereafter, in order to prolong the dormancy of micrometastases and all residual primary tumors To stabilize the condition, TβRII polypeptide can be administered to the patient.

[0231] In certain aspects of the present invention, other useful in combination tumor therapy with TβRII polypeptides Therapeutic agents are used in other cancer treatments, such as surgery, cytotoxic drugs, and radiological procedures (including irradiation). (including the administration of radioactive materials), chemotherapy agents, anti-hormone agents, growth inhibitors, anti-cancer drugs Compositions, and treatments with anticancer agents listed herein and known in the art Placement, or combinations thereof.

[0232] The term "cytotoxic agent," as used herein, refers to an agent that inhibits or inhibits the function of cells. This refers to a substance that interferes with and / or causes cell destruction. This term is a radioactive isotope. (For example, At 211 , I 131 , I 125 , Y 90 Re 186 Re 188 Sm 1 53 , Bi 212 , P 32 , and radioactive isotopes of Lu); chemotherapeutic agents, e.g., methotrexate Lexart, Adriamycin, Vinca alkaloids (vincristine, vinblastine) , etoposide, doxorubicin, melphalan, mitomycin C, chlorambucil, Daunorubicin or other inserts; enzymes and their fragments, e.g., nucleolytic enzymes; anti Biomaterials; as well as toxins of bacterial, fungal, plant or animal origin, such as small molecule toxins or yeasts. Artificially active toxins (including their fragments and / or variants); as disclosed below It is intended to include a variety of antitumor or anticancer agents. Other cytotoxic agents are included below. This is explained as follows: Tumor-killing agents cause the destruction of tumor cells.

[0233] "Chemotherapy agents" are chemical substances useful in treating cancer. Examples of chemotherapy agents include A Lucilates, for example, thiotepa and CYTOXAN® cyclophosphamide; Alkyl sulfonates, such as busulfan, improsulfan, and biposulfan Aziridines, for example, benzodopa, carbocone, metured opa and uredopa; ethyleneimine and methylamelam ines) (altretamine, triethylenemelamine, triethylenephosphoramide, trieth (Contains lentiophosphoramide and trimethylolomelamine) ;Acetogenins (especially bratacin and bratacinone); Delta-9-tetrahydrocan Dronabinol (MARINOL®); Beta-Lapacon; La Pacol; Colchicine; Betulinic acid; Camptothecin (synthetic analog Topotecan (HYCA) MTIN (registered trademark), CPT-11 (irinotecan), CAMPTOSAR (registered trademark) )), acetylcamptothecin, scopoletin, and 9-aminocanthol Putothecin; bryostatin; calistatin; CC-1065 (its adzereci Podophyllotoxin; Podophyllotoxin Phosphate; Teniposide; Cryptophycin (especially cryptophycin 1 and cryptophycin) Syn-8); Dorastatin; Duocalmycin (synthetic analogues, KW-2189 and CB) Contains 1-TM1; erythrorobin; pancratistatin; sarcodicin; spongi Statins; nitrogen mustards, e.g., chlorambucil, chlornafadin, syl Clophosfamide, Estramustine, Ifosfamide, Mechloretamine, Mechloretamine Dioxide hydrochloride, melphalan, nobembitin, fenestrine, prednimustine, Trophosphamide, uracil mustard; nitrosoureas, for example, Lumustine, chlorozotosine, hotemustine, lomustine, nimustine, and ranimustine N; Antibiotics, for example, endothelial antibiotics (for example, calicheamicin, in particular, calichemicin). Amycin gamma II (gammall) and calicheamicin omega II (omegall) (for example) See Agnew, Chem Intl. Ed. Engl., Vol. 33: pp. 183-186 (1994). I want to be; dynemycin (including dynemycin A); esperamicin; and Ne Occultinostatin chromophore and related pigment protein enediin antibiotic chromof Aclasinomycin, actinomycin, anthramycin (authramycin), azacerin, bleomycin, kakutinomycin, carab icin), carminomycin, cardinophilin, chromomycinis, da Kutinomycin, daunorubicin, detorubicin, 6-diazo-5-oxo-L-norlo Ishin, ADRIAMYCIN (registered trademark) Doxorubicin (Molfolino-doxorubicin) , cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin and deoxy (Including doxorubicin), epirubicin, esorubicin, idarubicin, marcelomaicin Mitomycin, for example, mitomycin C, mycophenolic acid, nogaramycin, Ribomycin, Peplomycin, Porphyomycin, Puromycin, Queramicin Rhodolubicin, streptonigrin, streptozocin, tubercidine, ubenimec Su, dinostatin, zolubicin; antimetabolites, e.g., methotrexate and 5-fluxine; Oloracil (5-FU); folate analogs, e.g., denopterin, methotrexate, pr Telopterin, trimethrexate; purine analogs, e.g., fludarabine, 6-mercury Putopurines, thiamiprines, thioguanines; pyrimidine analogs, e.g., ancitabine, Azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, do Xyfluridine, enocitabine, phloxuridine; androgens, e.g., carsterol N, dromostanolone propionate, epithiostanol, mepitiostanol, testopropyl alcohol Tons; anti-adrenal agents, for example, aminoglutethimide, mi Tin, trilostane; folic acid supplements, e.g., folinic acid; acegraton; aldofosphate Amid glycoside; aminolevulinic acid; enyluracil; amsacrine, bestlovesil ;bisanthrene;edatraxate;demecolsin;diaziquan;e elfornithine; eriptinium acetate; epotilone; etoglucide; nitrate Gallium; hydroxyurea; lentinan; ronidamine; maytansinoids, for example, may Tansine and anthamitosine; mitogwazone; mitoxantrone; mopidamol (mo pidanmol; nitraerine; pentostatin; fenamet; pirarubici N; Rosoxantrone; 2-Ethylhydrazide; Procarbazine; PSK (Registered Trademark) Sugar complex (JHS Natural Products, Eugene, OR); Lazocine Sun; Rhizoxin; Schizophyllan; Spirogermanium; Tenuazonic acid; Triadicone; 2,2',2”-Trichlorotriethylamine; Trichothecene (especially T-2 toxin, Verracurin A, loridine A, and anguidin); urethane; binde Syn (ELDISINE®, FILDESIN®); Dacarbazine Mannomustine; Mitobronitol; Mitractol; Pipobroman; Gacitosine (ga cytosine; arabinoside ("Ara-C"); thiotepa; taxoid, e.g., TAX OL (Registered Trademark) Paclitaxel (Bristol-Myers Squibb Onc Philology, Princeton, NJ), ABRAXANE (trademark) plagiarism Cremofoll-free, albumin-modified nanoparticle formulation (American Pharm aceutical Partners, Schaumberg, Illinois) and TAXOTERE® Doxetaxel (Rhone-Poulenc R Orer, Antony, France; Chlorambucil; Gemcitabine (GEMZA) R(registered trademark); 6-thioguanine; mercaptopurine; methotrexate; platinum-like metal. For example, cisplatin and carboplatin; vinblastine (VELBAN (registered) Trademark); Platinum; Etoposide (VP-16); Ifosfamide; Mitoxanthrone; Bin Christine (ONCOVIN®); Oxaliplatin; Leucovorin (leucov ovin; Vinorelbin (NAVELBINE®); Novantron; Edatrex Sart; Daunomycin; Aminopterin; Ibandronate; Topoisomerase inhibitors RFS 2000; Difluoromethylornithine (DMFO); Retinoids, e.g., Tinoic acid; capecitabine (XELODA®); any of the above pharmaceuticals Salts, acids, or derivatives that are generally acceptable; as well as combinations of two or more of the above. For example, CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisolone) FOLFOX (an abbreviation for nisolone combination therapy) and oxaliplatin (ELOXATIN) Examples include (trademark)) an abbreviation for a treatment regimen using 5-FU and leucovorin in combination. .

[0234] To regulate, reduce, block, or inhibit the effects of hormones that can promote cancer growth. Antihormone agents, which act in this way and are often in the form of systemic or systemic treatment drugs, These are included in the definition. They may also be hormones themselves. An example is anti-estrogen. Drugs and selective esterogen receptor modulators (SERMs) [e.g., tamoxifen] tamoxifen (NOLVADEX®, containing tamoxifen), EVISTA® ) Raloxifene, Droloxifene, 4-Hydroxytamoxifene, Trioxyphen N, Keoxyfen, LYl 17018, Onapriston, and FARESTON ( [Registered Trademark] Toremifene included; Anti-progesterone agent; Estrogen receptor downregulator ERDs (Endocrine Disorders); drugs that work to suppress or block the ovaries, e.g., progesterone Lumon-releasing hormone (LHRH) agonists, for example, LUPRON® and ELIGARD® Leuprolide Acetate, Goserelin Acetate, Buserelin Acetate Salts and triptorelin; other antiandrogens, e.g., flutamide, nilutamide and bicalutamide; and aromatase, an enzyme that regulates estrogen production in the adrenal gland. Inhibitory aromatase inhibitors, such as 4(5)-imidazole, aminoglutethimide, MEGASE® megestrol acetate, AROMASIN® Exemestane, Holmesan, Fadrozol, RIVIS OR (registered trademark) Borozo Letrozole, FEMARA® (registered trademark), and ARIMIDEX® (registered trademark) Examples include nastrozole. In addition, such a definition of chemotherapeutic agents is bisphosphonic. Nate, for example, clodronate (e.g., BONEFOS® or OSTA) © (Registered Trademark), DIDROC AL (Registered Trademark), Etidrone, NE-58095 ZOMET A (registered trademark) Zoledronic acid / Zoledrone, FOSAMAX (registered trademark) Alendronate, AREDIA (registered trademark), Pamidronate, SKELID (registered trademark) (Trademark) Childronate, or ACTONEL (registered trademark) Risedronate; and Roxacitabine (1,3-dioxolanenucleoside cytosine analog); antisense Ligonucleotides, particularly in the signaling pathways associated with abnormal cell proliferation, are involved in genetics. Expression of offspring, such as PKC-alpha, Raf, and H-Ras, and epidermal growth factor receptor Substances that inhibit the expression of the body's (EGF-R); vaccines, for example, THERATOPE (registered (Trademark) Vaccines and gene therapy vaccines, e.g., ALLOVECTIN® (registered trademark) Vaccines, LEUVECTIN® vaccine and VAXID® vaccine Chin; LURTOTECAN® topoisomerase 1 inhibitor; ABARELIX (Registered Trademark) rmRH; Lapatinib Tosylate (also known as GW572016, Er Bubble tyrosine kinase small molecule inhibitors against bB-2 and EGFR; and the above It contains any pharmaceutically acceptable salt, acid, or derivative of the substance.

[0235] When used herein, "growth inhibitor" refers to in vitro or in viv This refers to a compound or composition that inhibits cell growth in either way. Therefore, growth inhibition The agent may significantly reduce the percentage of cells in the S phase. Examples of growth inhibitors. Examples include drugs that block cell cycle progression (outside of the S phase), such as G1 arrest and M Examples of drugs that induce M-phase arrest include vinca-based drugs (vincli). (stines and vinblastine), taxane-based drugs, and topoisomerase II inhibitors For example, doxorubicin, epirubicin, daunorubicin, etoposide and bleoma Contains isine. Drugs that stop G1, such as DNA alkylating agents, such as tamoxif. Prednisone, dacarbazine, mechloretamine, cisplatin, methotrexate 5-Fluorouracil and ara-C also have an impact on S-phase arrest. Further information is available at The Molecular Basis of Cancer, edited by Mendelsohn and Israel, Chapter 1, title "Cell “cycle regulation, oncogenes, and antiplastic drugs” by Murakami et al. (WB This can be found in Saunders (Philadelphia, 1995), particularly on page 13. Taxane-based drugs (paclitaxel and docetaxel) are both derived from the yew tree. It is an anticancer drug derived from the European yew (Taxoteres). (Registered trademark), Rhone-Poulenc Rorer) is a manufacturer of paclitaxel (TAXOL It is a semi-synthetic analog of (registered trademark, Bristol-Myers Squibb). Clitaxel and docetaxel promote the construction of microtubules from tubulin dimers. By preventing depolymerization, microtubules are stabilized, and as a result, mitosis of cells is inhibited. It will become that.

[0236] In yet another embodiment, TβRII polypeptide is useful for the treatment or prevention of fibrosis. It may be. As used herein, the term “fibrosis” refers to a condition in which cells in an organ or tissue are affected. This refers to the abnormal formation or development of excessive fibrous connective tissue. The processes involved in fibrosis are: These processes occur as part of normal tissue formation or repair, but dysregulation of these processes can affect cells. This can lead to changes in composition and excessive connective tissue deposition that progressively impairs tissue or organ function. It can occur. The formation of fibrous tissue can result from repair or reaction processes. The disability or condition is related to vascular diseases, such as heart disease, cerebral disease, and peripheral vascular disease. Furthermore, related to tissues and organ systems, including the heart, skin, kidneys, peritoneum, gastrointestinal tract, and liver. (For example, Wynn, 2004, Nat Rev. 4, incorporated herein by reference) This includes fibroproliferative disorders (as disclosed on pages 583-594), but these include Not limited to. Exemplary disorders that can be treated include kidney injury / fibrosis. Diseases such as chronic nephropathy associated with diabetes (e.g., diabetic nephropathy), lupus, scleroderma, Renal fibrosis, including glomerulonephritis, focal segmental glomerulosclerosis, and IgA nephropathy; gastrointestinal fibrosis For example, scleroderma and radiation-induced gastrointestinal fibrosis; hepatic fibrosis, for example, cirrhosis, arco Viral-induced liver fibrosis, bile duct injury, primary biliary cirrhosis, infection or virus-induced liver fibrosis Hepatitis, congenital liver fibrosis and autoimmune hepatitis; as well as other fibrotic conditions, such as cystic liver fibrosis. Fibrosis, endocardial cardiomyopathy, mediastinal fibrosis, sarcoidosis, scleroderma, spinal cord injury / fibrosis, Myelofibrosis, vascular restenosis, atherosclerosis, injection fibrosis (as a complication of intramuscular injection) Endocardial cardiomyopathy, retroperitoneal fibrosis, and renal systemic fibrosis (which can occur especially in children) Fibrosis is one example, but it is not limited to these.

[0237] As used herein, the terms “fibrous disorder,” “fibrous condition,” and “fibrous disease” are used. "Fibrosis" is used synonymously to refer to a disorder, condition, or disease characterized by fibrosis. Examples of disorders include lupus, sclerotic disorders (e.g., scleroderma, atherosclerosis, etc.) and systemic sclerosis (including, for example, diffuse systemic sclerosis and progressive systemic sclerosis), Vascular fibrosis, pancreatic fibrosis, hepatic fibrosis (e.g., cirrhosis), renal fibrosis, skeletal muscular fibrosis, heart Visceral fibrosis (e.g., endocardial myocardial fibrosis, idiopathic cardiomyopathy), cutaneous fibrosis (e.g., scleroderma, Skin scars, keloids, and keloid formation due to post-traumatic surgery, ocular fibrosis (for example, Glaucoma, ocular sclerosis, conjunctival and corneal scarring, as well as pterygium), myelofibrosis, chronic graft pairs Host disease, Peyronie's disease, post-cystoscopy urethral stricture, idiopathic and pharmacologically induced retroflank Membrane fibrosis, mediastinal fibrosis, proliferative fibrosis, neoplastic fibrosis, Dupuytren's disease, stenosis, nerve scarring Examples include, but are not limited to, scarring, dermal scarring, idiopathic pulmonary fibrosis, and radiation-induced fibrosis. It will not be done.

[0238] In some embodiments, any of the polypeptides disclosed herein (e.g., sequence number A polypeptide having one amino acid sequence of numbers 13 and 50-56, alone To be used on its own, or in combination with one or more adjunctive therapies or active agents. And to treat, prevent, or The rate of progression and / or severity can be reduced. In some embodiments, Interstitial lung disease is pulmonary fibrosis. In some embodiments, interstitial lung disease is one of the following: Caused by any one of the following: silicosis, asbestosis, beryllium pulmonary disease, hypersensitivity pneumonitis, drug-induced dysphagia. Use (e.g., antibiotics, chemotherapy drugs, antiarrhythmics, statins), systemic sclerosis, multiple Myositis, dermatomyositis, systemic lupus erythematosus, rheumatoid arthritis, infections (e.g., atypical pneumonia) , Pneumocystis pneumonia, tuberculosis, Chlamydia trachomatis, and (or respiratory syncytial virus), carcinomatous lymphangitis, smoking, or developmental disorders. In terms of treatment methods, interstitial lung diseases include idiopathic (e.g., sarcoidosis, idiopathic pulmonary fibrosis, etc.) This includes Man-Ricci syndrome and / or anti-synthetase syndrome. Therefore, interstitial lung disease is idiopathic pulmonary fibrosis. In some embodiments, idiopathic pulmonary fibrosis The treatment is administered in combination with additional therapeutic agents. In some embodiments, the additional therapeutic agent is Pirfenidone, N-acetylcysteine, prednisone, azathioprine, ninteda The selection is made from the group consisting of nibs, their derivatives, and combinations thereof.

[0239] In some embodiments, any of the polypeptides disclosed herein (e.g., sequence number A polypeptide having one amino acid sequence of numbers 13 and 50-56, alone To be used on its own, or in combination with one or more adjunctive therapies or active agents. To treat, prevent, or slow the progression of kidney-related diseases or conditions. It can reduce the severity of the condition. When used herein, "kidney-related" "Renal disease or condition" refers to any disease, disorder, or condition affecting the kidneys or renal system. This is possible. Examples of kidney-related diseases or conditions include chronic kidney disease (or failure), acute kidney disease. Organ diseases (or failures), primary kidney disease, non-diabetic kidney disease, glomerulonephritis, interstitial nephritis Diabetic kidney disease, diabetic nephropathy, glomerulosclerosis, rapidly progressive glomerulonephritis, renal fibrosis, Alport syndrome, IDDM nephritis, mesangial proliferative glomerulonephritis, membranoproliferative glomerulonephritis Inflammation, crescentic glomerulonephritis, interstitial renal fibrosis, focal segmental glomerulosclerosis, membranous nephropathy, micro Minimal change nephrotic syndrome, microimmune rapidly progressive glomerulonephritis, IgA nephropathy, polycystic cysts Kidney, Dent's disease, nephrogenic cystinosis, Hymann's nephritis, autosomal dominant (adult) polycystic kidney disease, erythrocyte Chromosomal recessive (childhood) polycystic kidney disease, acute kidney injury, nephrotic syndrome, renal ischemia, glomerular epithelium Cellular diseases or disorders, proteinuria, glomerular diseases, membranous glomerulonephritis, focal segmental glomerulonephritis, children Preeclampsia, eclampsia, renal disorders, collagen disease-related vascular disorders, benign orthostatic (postural) proteinuria, IgM nephropathy , membranous nephropathy, sarcoidosis, diabetes mellitus, drug-induced kidney injury, Fabry disease, A Minoaciduria, Fanconi syndrome, hypertensive nephrosclerosis, interstitial nephritis, sickle cell anemia, hemoglobin Globinuria, myoglobinuria, Wegener's granulomatosis, glycogen storage disease type 1, chronic kidney disease, chronic renal failure, low glomerular filtration rate (GFR), renal vascular sclerosis, lupus nephritis, ANCA positive trace Immunotransitional crescentic glomerulonephritis, chronic allogeneic transplant nephropathy, nephrotoxicity, kidney necrosis, kidney injury Wounds, glomerular and tubular injuries, renal dysfunction, nephritis syndrome, acute renal failure, chronic renal failure, Fallopian tube dysfunction, acute kidney transplant rejection, chronic kidney transplant rejection, non-IgA mesangial Proliferative glomerulonephritis, post-infectious glomerulonephritis, vasculitis associated with all types of kidney disorders, all types of kidney disease Transmissible kidney disease, all forms of interstitial nephritis, kidney transplant failure, kidney cancer, and other conditions (e.g., hypertension). Kidney diseases associated with diabetes and autoimmune diseases, Dent's disease, nephrogenic cystinosis, Hyman's kidney Inflammation, primary kidney disease, collapsed glomerulosynovitis, dense deposit disease, cryoglobulinemia-related conditions Glomerulonephritis, Noch-Schönlein disease, post-infectious glomerulonephritis, bacterial endocarditis, microscopic Polyangiitis, Churg-Strauss syndrome, anti-GBM antibody-mediated glomerulonephritis, amyloidosis Cis, monoclonal immunoglobulinemia, fibrillary glomerulonephritis, immunotactoid glomerulonephritis Physical symptoms, ischemic tubular injury, drug-induced tubulointerstitial nephritis, toxic tubulointerstitial nephritis, infectious As a result of tubulointerstitial nephritis, bacterial pyelonephritis, polyomavirus infection or HIV infection Viral tubulointerstitial nephritis resulting from this, metabolic disease-induced tubulointerstitial disease, mixed type It can result from nephropathy, cylindrical nephropathy, urate or oxalate or drug-induced crystal deposition. Crystalline nephropathy, acute cellular tubulointerstitial allograft rejection, lymphoma, or post-transplant lymphoma Tumor-invasive diseases resulting from tumor-proliferative disorders, obstructive kidney disease, vascular diseases, thrombotic Microangiopathy, renal arteriosclerosis, atherothrombotic disease, mixed connective tissue disease, polyarteriovenous nodule Inflammation, calcineurin inhibitor-induced vascular disease, acute cellular vascular allograft rejection, acute Humoral allograft rejection, early renal failure (ERFD), end-stage renal disease (ESRD), renal vascular Thrombosis, acute tubular necrosis, acute interstitial nephritis, pre-existing chronic kidney disease, renal artery stenosis, ischemic Nephropathy, uremia, drug and toxin-induced chronic tubulointerstitial nephritis, reflux nephropathy, kidney stones, etc. Dopasture syndrome, normocytic normochromic anemia, renal anemia, diabetic chronic kidney disease, IgG4 Related diseases: von Hippel-Lindau syndrome, tuberous sclerosis, nephronophthenia, renal medullary cystic disease Renal cell carcinoma, adenocarcinoma, nephroblastoma, lymphoma, leukemia, hyposialylation disorder, chronic cyclosporine Nephropathy, renal reperfusion injury, renal dysplasia, hyperuremia, bilateral arterial occlusion, acute uric nephropathy, hypovolemia Acute bilateral obstructive urinary tract disease, hypercalcemia nephropathy, hemolytic uremic syndrome, acute urinary retention Malignant nephrosclerosis, postpartum glomerulosclerosis, scleroderma, non-Goodpasture anti-GBM disease, microscopic Polyarteritis nodosa, allergic granulomatosis, acute radiation nephritis, post-streptococcal glomerulosclerosis Nephritis, Waldenström macroglobulinemia, analgesic nephropathy, arteriovenous fistula, arteriovenous glomerulonephrosis Phytophthous dystrophy, dialysis, ectopic kidney, cavernous kidney, renal osteodystrophy, mononephropathy, hydronephrosis, microalbumin Peeinguria, uremia, hematuria, hyperlipidemia, hypoalbuminemia, fatty urine, acidosis, hyperkalemia This includes, but is not limited to, bloody edema and edema.

[0240] In some embodiments, any of the polypeptides disclosed herein (e.g., sequence number A polypeptide having one amino acid sequence of numbers 13 and 50-56, alone To be used on its own, or in combination with one or more adjunctive therapies or active agents. And to treat chronic kidney disease (e.g., tissue damage, inflammation and / or fibrosis) It can prevent the disease or reduce its progression and / or severity. Chronic kidney disease (CKD), also known as chronic kidney disease, is a condition that can last for months or years. This is a gradual loss of kidney function over time. Symptoms of worsening kidney function include poor overall health. This may include feeling that and experiencing a loss of appetite. In many cases, chronic kidney disease is People with kidney disease, such as those with high blood pressure or diabetes, and those with a blood relative who has CKD. It is diagnosed as a result of screening of people who are known to be at risk. The patient may develop one of its recognized complications, such as cardiovascular disease, anemia, or pericarditis. It may be confirmed at that time. Recent expert guidelines classify the severity of CKD into 5 stages. It is classified into stages, with Stage 1 being mild and usually causing little to no symptoms. Stage 5 CKD is a serious illness with a short life expectancy if left untreated. Often referred to as end-stage renal disease or end-stage renal failure, this term is now considered outdated. This is generally synonymous with nephrotic renal failure or chronic renal failure, and may include forms of dialysis, but is ideal. In general, this usually means that the patient requires renal replacement therapy equivalent to a kidney transplant. (CKD) In its early stages, it often presents with no specific symptoms, and generally involves an increase in serum creatinine or urinary protein. It is detected only as an additive. As kidney function declines, various symptoms manifest as follows: It can manifest as fluid overload and is produced by the kidneys via the renin-angiotensin system. The production of vasoactive hormones can cause blood pressure to rise, resulting in hypertension. The risk of developing the disease and / or developing congestive heart failure increases. It can accumulate, resulting in azotemia and ultimately uremia (from lethargy to pericarditis). It can lead to various symptoms, including encephalopathy. Due to this high level of systemic circulation, the urine is highly concentrated and ductile. It is excreted in phosphate sweat and crystallizes on the skin as the sweat evaporates ("urea frost"). Potassium It can accumulate in the blood (causing various symptoms, including fatigue and potentially fatal arrhythmias). Symptomatic hyperkalemia. Hyperkalemia is usually characterized by a glomerular filtration rate of 20-25 mg / L. The onset of symptoms does not occur until potassium levels drop to less than l / min / 1.73m2, and at the time of onset, renal potassium excretion is impaired. Potassium excretion capacity is reduced. Hyperkalemia in CKD is due to acidic blood (potassium extracellular transfer). It can be exacerbated by (leading to) and insulin deficiency. Erythropoietin It can also reduce synthesis, leading to anemia. This can progress from mild edema to fatal pulmonary edema. Various symptoms of excess body fluid volume can occur. Hyperphosphatemia caused by decreased phosphate excretion is Generally, it can occur after a decrease in glomerular filtration. Hyperphosphatemia is associated with an increased cardiovascular risk. This directly stimulates vascular calcification. Hypocalcemia may also become apparent, which is... Generally, it is caused by stimulation of fibroblast growth factor 23. Osteocytes use the enzyme 1-al pha-hydroxylase (1,25-dihydroxy of 25-hydroxycholecalciferol) It is a potent inhibitor of (which is involved in the conversion to vitamin D3), and is involved in increasing the production of FGF23. It agrees. Later, this led to secondary hyperparathyroidism, renal osteodystrophy, and vascular It progresses to calcification, and vascular calcification further impairs cardiac function. Metabolic acidosis ( This occurs due to the accumulation of sulfates, phosphates, uric acid, etc., and the excessive action of acid on enzymes. This can cause changes in enzyme activity, and hyperpotassium can result from excessive acid (acidic blood). The promotion of mucinosis can also lead to increased excitability of the pericardium and neuronal membranes. Sidosis is caused by a decrease in the ability of cells in the proximal tubules to produce sufficient ammonia. Iron deficiency anemia, in particular, can occur as kidney function declines, and its prevalence increases with age. It is frequently seen in those requiring hydrodialysis. Iron deficiency anemia is multifactorial in terms of its cause. However, these include increased inflammation, decreased erythropoietin, and hyperuricemia, which contribute to bone marrow suppression. Connected. People with CKD suffer from rapidly progressive atherosclerosis, and the general public Patients with chronic kidney disease (CKD) and cardiovascular disease are more likely to develop cardiovascular disease than others. Some individuals tend to have a significantly worse prognosis than those who suffer from cardiovascular disease alone. In this embodiment, chronic kidney disease is a condition resulting from kidney disease, affecting the interrelated skeletal, cardiovascular, and It is a widespread syndrome of mineral metabolism disorders, including bone mineral metabolism disorders associated with chronic kidney disease. CKD-MBD is a type of bone dystrophy often referred to as renal bone dystrophy (ROD). Encompassing various pathological conditions, ROD is in any of the polypeptides disclosed herein, or Preferred embodiment of treatment in combination with one or more adjunctive therapies or active agents Therefore, depending on the relative contribution of different virulence factors, ROD is a diverse disease of bone remodeling. This manifests as a specific pattern (Rosen et al. (ed.) Primer on the Metabolic Bone Dis). eases and Disorders of Mineral Metabolism, 7th edition, American Society for Hruska, in Bone and Mineral Research, Washington DC, pp. 343-349 In 2008, Chronic kidney disease mineral bone disorders (CKD-MBD) were diagnosed. At one end of that range are uremic osteodystrophy and ROD with low bone turnover. This is characterized by a small number of active remodeling sites, significant suppression of bone formation, and low bone resorption. The other end is characterized by hyperparathyroidism, high bone turnover, and fibrous osteitis. There is a rod that is accompanied by this.

[0241] In certain embodiments, any of the polypeptides disclosed herein (e.g., sequence number Polypeptides having amino acid sequences 13 and 50-56 can be used alone. Or used in combination with one or more adjunctive therapies or active agents, for bone marrow fibers Diseases (e.g., primary myelofibrosis, myelofibrosis after polycythemia euna, and essential platelet disease) To treat, prevent, or control the rate of progression of myelofibrosis (post-hemorrhagic myelofibrosis) and / or if It can reduce the severity. In particular, using ActRIIB antagonists alone To use, or in combination with one or more adjunctive therapies or active agents. For example, ineffective hematopoiesis, anemia, inflammation, fibrosis (e.g., myelofibrosis, splenic fibrosis, and liver fibrosis) Visceral fibrosis), pancytopenia, thrombocytopenia, extramedullary hematopoiesis (e.g., splenic extramedullary hematopoiesis, hepatic extramedullary hematopoiesis, Extramedullary hematopoiesis in the lungs and lymph nodes, hepatomegaly, splenomegaly, osteosclerosis, myelofibrosis of the bone, deformities Red blood cell count, fatigue, weight loss, night sweats, fever, itching, bone pain, early satiety, abdominal pain or discomfort , including joint pain, muscle pain, parasthesia, cachexia, splenic infarction and hemorrhage, bone marrow Treating, preventing, or slowing the progression of one or more complications of fibrosis. It can reduce the degree and / or severity.

[0242] When used herein, inhibition of the fibrous response of cells is used within the liver (or liver tissue). One or more cells, one or more cells within the kidney (or kidney tissue), muscle One or more cells within a tissue, one or more cells within the heart (or cardiac tissue) Cells, one or more cells in the pancreas, one or more cells in the skin, one in the bone or multiple cells, one or more cells within a vascular structure, one or more stem cells, This includes, but is not limited to, the inhibition of the fibrous response of one or more cells within the eye. do not have.

[0243] In some embodiments, any of the TβRII polypeptides disclosed herein are used to treat autoimmune diseases. It can be used to treat or treat disorders. In some embodiments, autoimmune diseases or disorders This includes spondyloarthritis; ankylosing spondylitis, arthritis, psoriatic arthritis / spondylitis, enteritis-induced arthritis, and reactive arthritis. Arthritis, Reiter's syndrome, undifferentiated spondyloarthropathy; reactive arthritis, rheumatoid arthritis, inflammatory bowel syndrome Crohn's disease, rheumatoid arthritis, rheumatoid spondylitis, osteoarthritis, gouty arthritis, allergies Ghee, multiple sclerosis, autoimmune diabetes, autoimmune uveitis, and nephrotic syndrome It is selected from the group consisting of the following.

[0244] In some embodiments, any of the TβRII polypeptides of this disclosure are used to treat metabolic disorders. It can be used for obesity or diabetes (for example). In some embodiments, metabolic disorders are obesity or diabetes (for example). (Type 1 or Type 2 diabetes), fatty liver disease, diabetic neuropathy, peripheral neuropathy, glucose These include diabetic retinopathy, diabetic ulcer formation, retinopathy ulcer formation, and diabetic macrovascular disease.

[0245] In some embodiments, any of the TβRII polypeptides of this disclosure is used to treat infectious diseases. It can be used for monitoring the progress of implantation or organ or tissue transplantation.

[0246] In some embodiments, any of the TβRII polypeptides of this disclosure is used in chronic obstructive pulmonary disease Used in the treatment of COPD, chronic obstructive airway disorder, idiopathic pulmonary fibrosis, and / or asthma. It is possible. In part, this disclosure relates to pulmonary hypertension (e.g., pulmonary arterial hypertension). A method of treating ) and providing TβRII polypeptide (for example) to patients who need it. (a polypeptide containing one of the amino acid sequences of SEQ ID NOs. 13 and 50-56) The disclosure also relates to methods including administering an effective dose. In some embodiments, the disclosure relates to pulmonary hypertension. One or more complications of the disease (e.g., increased smooth muscle and / or endothelial cells in the pulmonary artery) Angiogenesis in the pulmonary arteries, dyspnea, chest pain, pulmonary vascular remodeling, right ventricular hypertrophy, and A method for treating pulmonary fibrosis, and for patients who require it, TβRII polypeptide. The present disclosure envisions a method that includes administering an effective dose of [the substance]. In some embodiments, the present disclosure involves the lung A method for preventing one or more complications of hypertension, which is used in patients who require it. The proposed method involves administering an effective dose of βRII polypeptide. In this context, the present disclosure is a method for reducing the rate of progression of pulmonary hypertension, which requires The proposed method involves administering an effective dose of TβRII polypeptide to the patient. In this embodiment, the present disclosure reduces the rate of progression of one or more complications of pulmonary hypertension. This method involves administering an effective amount of TβRII polypeptide to patients who require it. The present disclosure intends to include methods that include the following. In some embodiments, the present disclosure reduces the severity of pulmonary hypertension. This method involves administering an effective dose of TβRII polypeptide to patients who require it. The present disclosure intends to include methods that include doing so. In some embodiments, the present disclosure describes one of the conditions of pulmonary hypertension. Alternatively, a method to reduce the severity of multiple complications, provided to patients who require it (TβR) The proposed method involves administering an effective dose of polypeptide II, in the lungs as needed. To treat, prevent, or reduce the rate of progression and / or severity of hypertension. In order to treat, prevent, or treat one or more complications of pulmonary hypertension Disclosed herein for reducing the rate of progression and / or severity of those progressions The methods used include one or more adjunctive therapies or additional activities to treat pulmonary hypertension. This may further include administering sexually transmitted drugs to the patient.

[0247] This invention relates to the treatment of TβRII polyps in combination with one or more other therapeutic modalities. The use of butyl is intended. Therefore, in addition to the use of TβRII polypeptide, the line It may also be administered as part of one or more "standard" treatments for vascular disorders. For example, TβRII polypeptide can be used as a cytotoxin, immunosuppressant, radiotoxic agent, and / or It can be administered in combination with (i.e., together with) a therapeutic antibody. Specific concomitant therapeutic agents shown include steroids (e.g., corticosteroids, e.g., Prednisone), immunosuppressants and / or anti-inflammatory agents (e.g., gamma-interferon) Cyclophosphamide, azathioprine, methotrexate, penicillamine, cyclosporine Phosphorus, colchicine, antithymocyte globulin, mycophenolate mofetil, and hydro Xychloroquine), cytotoxic agents, calcium channel blockers (e.g., nifedipine), Angiotensin-converting enzyme (ACE) inhibitors, para-aminobenzoic acid (PABA), Dimethyl sulfoxide, transforming growth factor beta (TGFβ) inhibitor, Examples include tarleukin-5 (IL-5) inhibitors and pancaspase inhibitors, These are not the only options.

[0248] As an additional antifibrotic agent that can be used in combination with TβRII polypeptide. This refers to lectins (for example, those described in U.S. Patent No. 7,026,283). The entire contents of this reference are incorporated herein by reference)), and Wynn et al. 2007, J Clin Invest, Vol. 117: pp. 524-529 (The full contents of this reference are Examples include antifibrotic agents described in (which are incorporated herein by reference), but , but not limited to, additional antifibrotic agents and treatments include various anti-inflammatory agents. / Immunosuppressant / Cytotoxic drugs (colchicine, azathioprine, cyclophosphamide, prednisolone) (Contains zoon, thalidomide, pentoxifylline and theophylline), TGFβ signal Transduction modifiers (relaxin, SMAD7, HGF and BMP7, as well as TGFβ1, (including TβRIs, TβRIIs, EGR-Is and CTGF inhibitors), cytokines and Cytokine receptor antagonists (IL-1β, IL-5, IL-6, IL-13, I L-21, IL-4R, IL-13Rα1, GM-CSF, TNF-α, Oncostatin M, WlSP-I and PDGF inhibitors), cytokines and chemokines (IFN- γ, IFN-α / β, IL-12, IL-10, HGF, CXCL10 and CXCL1 1) Chemokine antagonists (CXCL1, CXCL2, CXCL12, CCL2, Inhibitors of CCL3, CCL6, CCL17, and CCL18, chemokine receptor antagonists Gonists (inhibitors of CCR2, CCR3, CCR5, CCR7, CXCR2 and CXCR4) (harmful agents), TLR antagonists (inhibitors of TLR3, TLR4, and TLR9), angiogenesis Live antagonist (VEGF-specific antibody and adenosine deaminase replacement therapy), antihypertensive Drugs (beta-blockers and inhibitors of ANG11, ACE, and aldosterone), blood vessels Agonists (ET-1 receptor antagonists and bosentan), collagen Inhibitors of enzymes that synthesize and process prolyl hydroxylase, B cell antagonist (rituximab), integrin / adhesion molecule antagonist (α1 Molecules that block β1 and αvβ6 integrins, as well as integrin-linked kinases Inhibitors (and antibodies specific to ICAM-I and VCAM-I) target myofibroblasts. Apoptosis-promoting drugs, MMP inhibitors (inhibition of MMP2, MMP9, and MMP12) Examples include drugs, as well as TlMP inhibitors (antibodies specific to TIMP-1), but these Not limited to this.

[0249] Administer the TβRII polypeptide and the concomitant therapeutic agent or combination therapy using the same formulation. They can be administered together or separately. In the case of separate administration, TβRII polypeptide It may be administered before, after, or concurrently with the combination therapy or combination treatment. Yes, it is possible. One drug may be administered before the other drug, even if the interval between doses ranges from a few minutes to several weeks. Often, or may continue. Two or more types of therapeutic agents may be applied separately to the subject. In this embodiment, these different types of drugs provide a beneficial combined effect to target tissue or cells. In contrast, a meaningful period exists between each delivery so that it can still be effective. This would generally guarantee that there were no interruptions.

[0250] 10. Pharmaceutical Compositions A therapeutic agent described herein (e.g., a TβRII fusion polypeptide) in a pharmaceutical composition It can be formulated into pharmaceutical compositions. One or more pharmaceutical compositions for use in accordance with this disclosure. It can be formulated in the conventional way using physiologically acceptable carriers or excipients. Such formulations comply with the requirements of most regulatory authorities and are essentially pyrogens. Become a free agent.

[0251] In certain embodiments, the therapeutic method of the present disclosure may involve systemic administration of the composition or implantation of an indwelling agent. or includes local administration as a device. When administered, therapeutic use for the purposes of this disclosure The therapeutic composition is in a pyrogen-free, physiologically acceptable form. It may be included as needed, in treatments other than TβRII signaling antagonists. A suitable agent can be used in the manner disclosed herein to target a compound (e.g., TβRII polypeptide). It can be administered simultaneously with or sequentially to (d).

[0252] Typically, the protein therapeutic agents disclosed herein are administered parenterally, in particular intravenously. Alternatively, it will be administered subcutaneously. Pharmaceutical compositions suitable for parenteral administration include antioxidants and buffers. The agent, bacteriostatic agent, solute to make the preparation isotonic with the blood of the intended recipient, or suspension. Or one or more pharmaceutically acceptable sterile isotonic waters that may contain a thickening agent or This refers to a non-aqueous solution, dispersion, suspension or emulsion, or a sterile injection solution prepared immediately before use. Alternatively, one or more TβRII particles can be combined with sterile powder that is reconstituted into a dispersion. May contain lipeptides. Suitable aqueous and non-aqueous materials may be used in the pharmaceutical compositions of this disclosure. Examples of aqueous carriers include water, ethanol, and polyols (e.g., glycerol, propylene). Polyglycols, polyethylene glycols and similar substances, and their equivalents A fine mixture, vegetable oil, such as olive oil, and injectable organic ester, such as olive oil. Ethyl iodide is one example. Proper fluidity is achieved, for example, by coating agents such as lecithin. Through use, in the case of dispersions, the required particle size is maintained, and through the use of surfactants, It is possible to possess it.

[0253] The composition and formulation may, if desired, be provided as one or more unit preparations containing the active ingredient. It can be provided in a pack or dispenser device that can contain the shape. This may include, for example, metal or plastic foil, such as blister packs. Instructions for administration can be attached to the dispenser device.

[0254] Furthermore, the composition can be encapsulated or injected into a form for delivery to a target tissue site. In certain embodiments, the composition of the present invention comprises one or more therapeutic compounds (e.g., For example, TβRII polypeptide is delivered to a target tissue site to give structure to developing tissue. It may include a matrix that is capable of being reabsorbed into the body, and optimally. For example, The matrix can result in the sustained release of TβRII polypeptide. Trix can be formed using materials currently used in other implantable medical applications. Cut.

[0255] The selection of matrix materials is based on biocompatibility, biodegradability, mechanical properties, aesthetic appearance, and Based on interfacial properties, the appropriate formulation will be determined by the specific application of the target composition. Possible matrix for compositions is a biodegradable, chemically defined calcium sulfate. It is um, tricalcium phosphate, hydroxyapatite, polylactic acid, and polyanhydride. Other possible materials, such as bone and dermal collagen, are biodegradable and biodegradable. It is clearly defined in terms of physical properties. Further matrices are pure proteins or cells. It is composed of exomatrix components. Other possible matrices include, for example, sintered hydroxyl Apatite, bioglass, aluminates, or other ceramics are non-biodegradable. It is chemically defined. The matrix is ​​one of the above types of materials. Combinations of these, for example, polylactic acid and hydroxyapatite, or collagen and tricalcium phosphate. It can also be composed of aluminium. Bioceramics have a composition, for example, aluminium The composition of calcium phosphate can be varied, as can the pore size and grain size in terms of processing. The diameter, particle shape, and biodegradability can be varied.

[0256] In a particular embodiment, the method of the present invention involves each of the following: It contains capsules, cashews, pills, tablets, and medicinal candies (flavoring base, usually Using sucrose and gum arabic or tragacanth, in powder or granular form, Alternatively, as a solution or suspension in an aqueous or non-aqueous liquid, or as an oil-in-water type or As a water-in-oil liquid emulsion, or as an elixir or syrup, or Roach (inert gelatin and glycerin, or sucrose and gum arabic) (using a base), as well as / or mouthwash and similar products. It can be administered orally. The drug can be administered as a pill, lick, or paste. It is also possible to do so.

[0257] Solid dosage forms for oral administration (capsules, tablets, pills, sugar-coated pills, powders, granules and similar) In the case of (that which), one or more therapeutic compounds of the present invention are one or more pharmaceutical Acceptable carriers include, for example, sodium citrate or calcium phosphate, and / or It may be mixed with any of the following: (1) a filler or bulking agent, for example, Starch, lactose, sucrose, glucose, mannitol and / or silicic acid, (2) Binders, for example, carboxymethylcellulose, alginate, gelatin, polyvinylcellulose (3) Moisturizers, such as nylpyrrolidone, sucrose and / or gum arabic, Glycerol, (4) Disintegrant, e.g., agar, calcium carbonate, potato or tapioca Oka starch, alginic acid, certain silicates, and sodium carbonate, (5) slow dissolution (6) an adsorbent, e.g., paraffin, (6) an absorption enhancer, e.g., a quaternary ammonium compound, 7) Humectants, such as cetyl alcohol and glycerol monostearate, (8 ) Adsorbents, for example, kaolin and bentonite clay, (9) Lubricants, for example, talc, Calcium stearate, magnesium stearate, solid polyethylene glycol, Sodium uryl sulfate, and mixtures thereof, and (10) coloring agents. Capsules, tablets In the case of drugs and pills, the pharmaceutical composition may also include buffering agents. Similar types of solid compositions The substances include lactose or milk sugar and high molecular weight polyethylene glycol and similar substances. Excipients such as those used are utilized as fillers in gelatin-based soft and hard capsules. It can also be used.

[0258] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions and microemulsions. It includes solutions, suspensions, syrups, and elixirs. In addition to the active ingredient, the liquid dosage forms include: Inert diluents commonly used in this art, such as water or other solvents, Solubilizers and emulsifiers, for example, ethyl alcohol, isopropyl alcohol, ethyl carbonate L, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1, 3-Butylene glycol, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, Olive oil, castor oil and sesame oil), glycerol, tetrahydrofurfuryl (tetrah ydrofuryl) alcohol, polyethylene glycol, and sorbitan fatty acid esters , and mixtures thereof may be included. In addition to inert diluents, the oral composition may contain adjuvant For example, humectants, emulsifiers and suspending agents, sweeteners, flavoring agents, colorants, fragrances, and / or may also contain preservatives.

[0259] The suspension contains the active compound, as well as a suspending agent, such as ethoxylated isostearyl alcohol. Cellulose, polyoxyethylene sorbitol, and sorbitan esters, microcrystalline cellulose Aluminum, metahydroxide, bentonite, agar, and tragacanth, and these It may contain a mixture of these.

[0260] The composition of the present invention also includes adjuvants, such as preservatives, humectants, emulsifiers, and dispersants. It is possible. Prevention of microbial action can be achieved with various antibacterial and antifungal agents, such as parabens and chloroforms. By including lobutanol, phenol, sorbic acid and similar substances It can be secured. By combining isotonic agents such as sugar, sodium chloride, and similar substances. It may also be desirable to include it in the product. In addition, drugs that slow down absorption, such as monos By including aluminum thearate and gelatin, the sustained efficacy of the injectable drug form It can lead to absorption.

[0261] The drug regimen affects the action of the target compound of the present invention (e.g., TβRII fusion polypeptide). It is understood that the decision will be made by the attending physician, taking into account various modifying factors. Various factors include the patient's age, sex and diet, disease severity, number of doses administered, and other clinical factors. This includes, but is not limited to, bed-related factors. If necessary, the dosage used for reconstitution is... This can vary depending on the type of matrix and the type of compounds in the composition. The addition of other known growth factors to the composition may also affect the dosage. Bone growth and / or This involves periodic assessment of the restoration, such as X-ray (including DEXA), tissue morphometric assessment, and tectonics. The tracycling markers allow for monitoring of its progress.

[0262] In certain embodiments, the present invention enables the in vivo use of TβRII fusion polypeptides. We also offer gene therapy for the production of [unclear]. Such therapies address the disorders listed above. The therapeutic effect is achieved by introducing the TβRII polynucleotide sequence into cells or tissues containing it. It is expected that this will be achieved. Delivery of the TβRII polynucleotide sequence is expected to be achieved by the chimeric virus. This can be achieved using recombinant expression vectors, such as colloidal dispersions. The use of targeted liposomes is preferred for the therapeutic delivery of TβRII polynucleotide sequences.

[0263] Various viral vectors that can be used in gene therapies as taught herein - is an adenovirus, herpesvirus, vaccinia, or preferably an RNA virus. A virus, for example, a retrovirus. Preferably, the retroviral vector is a mouse. It is a derivative of a triretrovirus. It is a retrovirus that can insert a single foreign gene. Examples of rovirus vectors include Moloney's mouse leukemia virus (MoMuLV), and - Bay mouse sarcoma virus (HaMuSV), mouse mammary cancer virus (MuMTV), and Bilous sarcoma virus (RSV) is one example, but is not limited to these. Numerous other viruses... These retroviral vectors can incorporate multiple genes. All of them can have genes for selection markers transferred or incorporated, and as a result, Transduced cells can be identified and generated, for example, sugars, glycolipids, or tannins. By attaching a protein, retroviral vectors can be made target-specific. The preferred targeting is performed by using antibodies. TβRII polynucleotide To enable targeted delivery of retroviral vectors containing cytoplasm, specific po Inserting a nucleotide sequence into the genome under a retrovirus or binding it to the viral capsule. Those skilled in the art will expect to see that it can be made to fit together. In a preferred embodiment, The vector is targeted to bone or cartilage.

[0264] Alternatively, plus, which encodes the retroviral structural genes gag, pol, and env. Mido was directly transfected into tissue culture cells using conventional calcium phosphate transfection. It can then be infected. Next, these cells are given a vector containing the target gene. The tarplasmid is transfected. The resulting cells are then subjected to a retroviral vector. Release into the culture medium.

[0265] Another targeted delivery system for TβRII polynucleotides is a colloidal dispersion system. Colloidal dispersion systems include polymer composites, nanocapsules, microspheres, beads, and This includes lipid-based systems, and the lipid-based systems include oil-in-water emulsions, micelles, and mixed micelles. The present invention includes liposomes. Somesomes are useful as delivery vehicles in vitro and in vivo, for humans These are membrane vesicles. They encapsulate RNA, DNA, and intact viral particles within an aqueous solution. It can be delivered to cells in a bioactive form (e.g., Fraley et al., Trends Biochem. Sci., See Volume 6, page 77, 1981. Efficient genetic research using liposome vehicles. The gene transfer method is publicly known in the art. For example, Mannino et al., Biotechniques, See Volume 6, page 682, 1988. The composition of liposomes is usually phospholipids. The combination is usually with steroids, especially cholesterol. Phospholipids or other lipids can also be used. The physical properties of liposomes are pH, It depends on the on-intensity and the presence of divalent cations.

[0266] Examples of lipids useful for liposome formation include phosphatidyl compounds, for example, phosphatidyl Tidylglycerol, phosphatidylcholine, phosphatidylserine, phosphatidyl Examples include tanolamines, sphingolipids, cerebrosides, and gangliosides. Exemplary phospholipids include egg phosphatidylcholine and dipalmitoyl phosphatidylcholine. Examples include distearoylphosphatidylcholine. For example, organ specificity, Liposome targeting based on cellular specificity and organelle specificity is also possible with this technology. It is common knowledge in the field.

[0267] This disclosure relates to acids and bases for adjusting pH and for maintaining pH within a narrow range. The present invention provides a formulation that can be modified to include a buffering agent. [Examples]

[0268] The present invention has now been described in general terms, but the present invention can be further described by referring to the following examples. It is expected that this will be easier to understand. The following examples are simply examples of a particular embodiment of the present invention. This is included for the purpose of illustrating the application method and is not intended to limit the present invention. do not have. (Example 1) Generation of receptor fusion protein mutants TβRII ECD mutant

[0269] A TβRII fusion protein containing the soluble extracellular portion and human Fc portion of human TβRII. This generated a TβRIIA with the amino acid sequence of SEQ ID NO: 18 for each fusion protein. The amino acid sequence is converted to the IgG Fc portion having the amino acid sequence of SEQ ID NO: 20, with several different values. They were fused by a single linker. Each of the fusion proteins is the amino acid of SEQ ID NO: 23. This also included TPA reader sequences containing acid sequences (see below). Organizational Plasminogen Activator (TPA): MDAMKRGLCCVLLLCG AVFVSP (Sequence ID 23)

[0270] Figure 3 provides illustrative outlines of some of the structures designed. A table detailing the sequences of different constructs tested in the experiment is provided below: [Table 1-1] [Table 1-2]

[0271] The amino acid sequences of the construct components and each construct are used to express these constructs. The nucleic acid sequences used are provided below. [ka] [ka] [ka] [ka] [ka] [ka] [ka] [ka]

[0272] We successfully expressed various constructs in CHO cells and performed size exclusion chromatography for analysis. The substance was purified to high purity by CG and SDS-PAGE. hTβRII(G 4S)2-hFc, hTβRII(G4S)3-hFc, hTβRII(G4S)4-h Fc, hTβRII(G4S)5-hFc and hTβRII(G4S)6-hFc tan The protein was analyzed by SDS-PAGE after being maintained in PBS for 13 days at 37°C. It consistently showed similarly strong stability. hTβRII(G4S)2-hFc, hTβRII (G4S)3-hFc and hTβRII(G4S)4-hFc proteins were also found in rats and mice. It was maintained in saturates or human serum and showed similarly strong stability.

[0273] TβRII ECD mutant The TβRII domain included in the fusion protein described above (for example, SEQ ID NO: 18) In addition, this disclosure also envisions fusion proteins containing alternative TβRII domains. The fusion protein is the wild-type hTβRII shown below. ショート (23~159) Arrangement (distribution) This may include column number 27), or any of the other TβRII polypeptides disclosed below: [ka]

[0274] (1) hTβRII shown below ショート (23~159 / D110K) Amino acid sequence (distribution) (Column number 36) (Underline the substituted residues in the sequence). [ka] [ka]

[0275] (2) The N-terminal shortened hTβRII shown below ショート (29~159) Amino acid sequence (sequence) Number 28). [ka]

[0276] (3) The N-terminal shortened hTβRII shown below ショート (35~159) Amino acid sequence (sequence) Number 29). [ka]

[0277] (4) C-terminal shortened hTβRII shown below ショート (23~153) Amino acid sequence (sequence) Number 30). [ka]

[0278] (5) C-terminal shortened hTβRII shown below ショート (23~153 / N70D) Amino Acids Sequence (SEQ ID NO: 38) (Substituted residues in the sequence are underlined). [ka]

[0279] The applicants have identified the wild-type hTβRII shown above and below. ロング (23~184) Five corresponding variants based on the sequence (sequence number 49) (sequence numbers 37, 33, 34, 39) (Underlining the 25 amino acid insertions in these sequences) is also being considered. Splicing However, this results in a conservative amino acid substitution (Val→Ile) at the adjacent position on the C-terminal side of the insertion. Please note the following. [ka] [ka]

[0280] (1) hTβRII shown below ロング (23~184 / D135K) Amino acid sequence (sequence) Number 37) (Double underline the substituted residue in the sequence). [ka]

[0281] (2) The N-terminal shortened hTβRII shown below ロング (29~184) Amino acid sequence (sequence number) No. 33). [ka]

[0282] (3) The N-terminal shortened hTβRII shown below ロング (60~184) Amino acid sequence (sequence number) (Same as No. 29). [ka]

[0283] (4) C-terminal shortened hTβRII shown below ロング (23~178) Amino acid sequence (sequence number) No. 34). [ka]

[0284] (5) C-terminal shortened hTβRII shown below ロング (23~178 / N95D) Amino Acid Blend Column (Sequence ID 39) (Double underline the substituted residues in the sequence). [ka] [ka] Additional TβRII ECD variants include the following:

[0285] (A) N-terminal and C-terminal shortened hTβRII shown below ショート (35~153) or hTβRII ロング (60-178) Amino acid sequence (SEQ ID NO: 32). [ka]

[0286] (B) N-terminal and C-terminal shortened hTβRII shown below ショート (29~153) Amino Acid sequence (SEQ ID NO: 31). [ka]

[0287] (C) N-terminal and C-terminal shortened hTβRII shown below ロング (29~178) Amino Acids Array (sequence number 35). [ka]

[0288] The above mutants (SEQ ID NOs: 36, 28, 29, 30, 38, 37, 33, 34, 39, 32 ,31 and 35) all of the hTβRII isoform C (Konrad et al., BMC Genom ICS Vol. 8: p. 318, 2007) contains 36 amino acid insertions (adaptations) that are naturally occurring. Column number 41) is a pair of glutamic acid residues located near the C-terminus of hTβRII ECD. (The content between position 151 and 152 of sequence number 1, or position 176 and 177 of sequence number 2) I was able to do it. GRCKIRHIGS NNRLQRSTCQ NTGWESAHVM KTPGFR (Sequence ID 41)

[0289] For example, hTβRII ショート (29~159) Regarding the mutant (sequence number 28) The pairs of glutamic acid residues adjacent to the insertion site, as needed, are shown below (underlined). [ka]

[0290] Fc domain variant The above construct was generated having the amino acid sequence of SEQ ID NO: 20 in the Fc domain, but this disclosure This is the human IgG2 Fc domain (SEQ ID NO: 42, see below) or the full-length human IgG1 F c(hG1Fc) (SEQ ID NO: 43, see below), containing an alternative Fc domain, hTβRI We are aiming for an I-hFc fusion protein. If necessary, we will add a port unrelated to the Fc domain. We were able to bind the lipeptide to the Fc domain. [ka]

[0291] Leader sequence variant The generated construct above included a TPA reader array, but also an alternative reader array, for example, Native leader sequence (sequence number 22 - see below) or honeybee melittin (sequence number 24 -The leader array shown below may be used. Native: MGRGLLRGLWPLHIVLWTRIAS (Sequence ID 22) Honeybee melittin (HBML):MKFLVNVALVFMVVYISYIYA(sequence) Number 24)

[0292] (Example 2) Differential ligand inhibition by receptor fusion protein variants in cell-based assays TGFβ1, TGFβ2, and TGFβ3, hTβRII(G4S)2-hFc, h TβRII(G4S)3-hFc, hTβRII(G4S)4-hFc, hTβRII- Biac The results were evaluated in vitro using the ore(trademark) instrument. The results are shown in Figures 4A and 4B. In short, each of the fusion proteins binds to TGFβ1 and TGFβ3 with high affinity. It was possible, but surprisingly, the linker length was longer than or equal to (G4S)4. A structure having (G4S)4 has a higher affinity than a structure having a shorter linker length than (G4S)4. It was able to bind to both GFβ1 and TGFβ3. It was also able to bind to either TGFβ2 or the construct. The bond between them was weak or transient. Deglycosylation of the structure was The combination did not change.

[0293] Using a reporter gene assay in A549 cells, TGFβ1, TGFβ2 Furthermore, the ability of the hTβRII-hFc mutant to inhibit TGFβ3 activity was determined. The assay was performed using the pGL3(CAGA)12 reporter plasmid (Dennler et al., 1998). , EMBO Vol. 17: pp. 3091-3100) transfected, as well as transfected The sea oyster reporter plasmid (pRLCMV), which serves as a control for injection efficiency, was used in the treatment. Based on an infected human lung cancer cell line. The CAGA motif is a TGFβ-responsive gene. Because it is present within the promoter of (for example, PAI-1), this vector is SMAD2 It is commonly used for factors that signal via SMAD3.

[0294] On the first day of the assay, A549 cells (ATCC®: CCL-185®) The mixture was distributed into 48-well plates. On the second day, pGL3(CAGA)12 and pRLCM were added. V and X-tremeGENE 9 (Roche Applied Science) A solution containing OptiMEM (Invitrogen) is pre-incubated. Next, Eagle's Minimum Essential Medium (EMEM, ATCC) supplemented with 0.1% BSA (Registered Trademark) Add this to the standard and sow for incubation overnight at 37°C and 5% CO2. The culture medium was applied to the seeded cells. On the third day, the culture medium was removed and the ligand prepared as described below was used. The cells were incubated overnight at 37°C and 5% CO2 with a mixture of the inhibitor.

[0295] The series dilution of the test substance in assay buffer (EMEM + 0.1% BSA) was performed in 48 wells. The procedure was performed at a rate. An equal volume of assay buffer containing the test ligand was added and determined beforehand. Final ligand concentrations equal to the EC50 were obtained for human TGFβ1, human TGFβ2 and Human TGFβ3 was obtained from PeproTech. The test solution was incubated at 37°C for 30 minutes. The mixture was incubated, and then a portion of the mixture was added to all the wells. The test solution and the mixture were incubated overnight. After incubation, the cells are rinsed with phosphate-buffered saline, and then with passive lysis buffer (P It was dissolved in romega E1941 and stored overnight at -70°C. On the 4th day and the final day... The plate was gently shaken and warmed to room temperature. The cell lysates were then chemiluminescently plated in double repeats. Transfer to a 96-well tray and use Dual-Luciferase Reporter As Using reagents from the SAY system (Promega E1980), the luminometer was used to measure the fractions. The normalized luciferase activity was determined through analysis.

[0296] As illustrated in Figures 5A-5F, hTβRII(G4S)2-hFc, hTβR II(G4S)3-hFc, hTβRII(G4S)4-hFc, hTβRII(G4S )5-hFc, hTβRII(G4S)6-hFc, hTβRII-hFc, and hT All βRII extension hinge-hFc proteins inhibit both TGFβ1 and TGFβ3. It was possible to cause harm. Interestingly, there was an improvement in TGFβ1 and TGFβ3 inhibition, and hTβ RII(G4S)2-hFc, hTβRII(G4S)3-hFc and hTβRII( There was a correlation between the linker length of the G4S)4-hFc structure and this direction (Figure 5E), The above trend is due to hTβRII(G4S)5-hFc and hTβRII(G4S)6-hF Structure c appeared to have reached a steady state (Figure 5F).

[0297] Inclusion by reference All published documents and patents referred to herein are individual published documents or patents. It is as if each of them is specifically and individually shown to be incorporated by reference, The entire text is incorporated herein by reference.

[0298] While we have discussed specific embodiments of the subject matter, the above specification is intended to be helpful in the explanation. This is not limited to the above. By reconsidering this specification and the claims below, Many variations will become apparent to those skilled in the art. The entire scope of the present invention is equivalent to the claims of this invention. By referring to the entire scope of the object, and this specification together with such variations, It should be decided by reference. The present invention provides, for example, the following items: (Item 1) A transforming growth factor β receptor II (TβRII) fusion polypeptide, , a) The extracellular domain of the TβRII portion, b) Different parts, c) The linker is a linker moiety that is at least 10 amino acids long and Includes, The TβRII extracellular domain portion is i) Starting at any of positions 23-35 of sequence number 1, and position 153-159 of sequence number 1 An array ending in any of the following, ii) Starting at any position between 23 and 60 of Sequence ID 2, and between 178 and 184 of Sequence ID 2 Arrays ending in any position A polypeptide containing at least 80% identical amino acid sequence. (Item 2) The TβRII extracellular domain portion begins at any of positions 23-35 of Sequence ID No. 1. , and is at least 80% identical to the sequence ending at position 153-159 of sequence number 1. A polypeptide containing the amino acid sequence described in item 1. (Item 3) The TβRII extracellular domain portion begins at any of positions 23-35 of Sequence ID No. 1. , and is at least 90% identical to the sequence ending at position 153-159 of sequence number 1. A polypeptide containing the amino acid sequence described in item 1. (Item 4) The TβRII extracellular domain portion begins at any of positions 23-35 of Sequence ID No. 1. , the sequence ending at any of positions 153-159 of sequence number 1 is at least 95% identical. A polypeptide containing the amino acid sequence described in item 1. (Item 5) The TβRII extracellular domain portion begins at any of positions 23-35 of Sequence ID No. 1. The sequence ending at position 153-159 of sequence number 1 is at least 97% identical. A polypeptide containing the amino acid sequence described in item 1. (Item 6) The TβRII extracellular domain portion begins at any of positions 23-35 of Sequence ID No. 1. , containing an amino acid sequence ending at any of positions 153-159 of SEQ ID NO: 1, as described in item 1. polypeptide. (Item 7) The TβRII extracellular domain portion begins at any of positions 23-60 of Sequence ID No. 2. The sequence ending at position 178-184 of sequence number 2 is at least 80% identical. A polypeptide containing the amino acid sequence described in item 1. (Item 8) The TβRII extracellular domain portion begins at any of positions 23-60 of Sequence ID No. 2. The sequence ending at position 178-184 of sequence number 2 is at least 90% identical. A polypeptide containing the amino acid sequence described in item 1. (Item 9) The TβRII extracellular domain portion begins at any of positions 23-60 of Sequence ID No. 2. The sequence ending at position 178-184 of sequence number 2 is at least 95% identical. A polypeptide containing the amino acid sequence described in item 1. (Item 10) The TβRII extracellular domain portion begins at any of positions 23-60 of Sequence ID No. 2. The sequence ending at position 178-184 of sequence number 2 is at least 97% identical. A polypeptide containing the amino acid sequence described in item 1. (Item 11) The TβRII extracellular domain portion begins at any of positions 23-60 of Sequence ID No. 2. , containing an amino acid sequence ending at any of positions 178-184 of SEQ ID NO: 2, as described in item 1. polypeptide. (Item 12) The TβRII extracellular domain portion is at least 80% identical to that of SEQ ID NO: 18. A polypeptide containing an acid sequence, as described in item 1. (Item 13) The TβRII extracellular domain portion is at least 90% identical to that of SEQ ID NO: 18. A polypeptide containing an acid sequence, as described in item 1. (Item 14) The TβRII extracellular domain portion is at least 95% identical to that of SEQ ID NO: 18. A polypeptide containing an acid sequence, as described in item 1. (Item 15) The TβRII extracellular domain portion is at least 97% identical to that of SEQ ID NO: 18. A polypeptide containing an acid sequence, as described in item 1. (Item 16) The TβRII extracellular domain portion includes the amino acid sequence of SEQ ID NO: 18, item 1 The polypeptide described. (Item 17) The TβRII extracellular domain portion begins at any of positions 23-35 of Sequence ID No. 1. , and is at least 80% identical to the sequence ending at position 153-159 of sequence number 1. A polypeptide consisting of the amino acid sequence described in item 1. (Item 18) The TβRII extracellular domain portion begins at any of positions 23-35 of Sequence ID No. 1. , and is at least 90% identical to the sequence ending at position 153-159 of sequence number 1. A polypeptide consisting of the amino acid sequence described in item 1. (Item 19) The TβRII extracellular domain portion begins at any of positions 23-35 of Sequence ID No. 1. , the sequence ending at any of positions 153-159 of sequence number 1 is at least 95% identical. A polypeptide consisting of the amino acid sequence described in item 1. (Item 20) The TβRII extracellular domain portion begins at any of positions 23-35 of Sequence ID No. 1. The sequence ending at position 153-159 of sequence number 1 is at least 97% identical. A polypeptide consisting of the amino acid sequence described in item 1. (Item 21) The TβRII extracellular domain portion begins at any of positions 23-35 of Sequence ID No. 1. , consisting of an amino acid sequence ending at any of positions 153-159 of sequence number 1, as described in item 1. Polypeptide. (Item 22) The TβRII extracellular domain portion begins at any of positions 23-60 of Sequence ID No. 2. The sequence ending at position 178-184 of sequence number 2 is at least 80% identical. A polypeptide consisting of the amino acid sequence described in item 1. (Item 23) The TβRII extracellular domain portion begins at any of positions 23-60 of Sequence ID No. 2. The sequence ending at position 178-184 of sequence number 2 is at least 90% identical. A polypeptide consisting of the amino acid sequence described in item 1. (Item 24) The TβRII extracellular domain portion begins at any of positions 23-60 of Sequence ID No. 2. The sequence ending at position 178-184 of sequence number 2 is at least 95% identical. A polypeptide consisting of the amino acid sequence described in item 1. (Item 25) The TβRII extracellular domain portion begins at any of positions 23-60 of Sequence ID No. 2. The sequence ending at position 178-184 of sequence number 2 is at least 97% identical. A polypeptide consisting of the amino acid sequence described in item 1. (Item 26) The TβRII extracellular domain portion begins at any of positions 23-60 of Sequence ID No. 2. , consisting of an amino acid sequence ending at any of positions 178-184 of sequence number 2, as described in item 1. Polypeptide. (Item 27) The TβRII extracellular domain portion is at least 80% identical to that of SEQ ID NO: 18. A polypeptide consisting of an acid sequence, as described in item 1. (Item 28) The TβRII extracellular domain portion is at least 90% identical to that of SEQ ID NO: 18. A polypeptide consisting of an acid sequence, as described in item 1. (Item 29) The TβRII extracellular domain portion is at least 95% identical to that of SEQ ID NO: 18. A polypeptide consisting of an acid sequence, as described in item 1. (Item 30) The TβRII extracellular domain portion is at least 97% identical to that of SEQ ID NO: 18. A polypeptide consisting of an acid sequence, as described in item 1. (Item 31) The extracellular domain portion of TβRII consists of the amino acid sequence of SEQ ID NO: 18, item 1 Polypeptides as described. (Item 32) A polypeptide containing an N-terminal leader sequence, as described in any one of items 1 to 31. (Item 33) The N-terminal leader sequence contains one of the amino acid sequences from SEQ ID NOs. 22 to 24. , polypeptides as described in item 32. (Item 34) The aforementioned N-terminal leader sequence includes the amino acid sequence of SEQ ID NO: 23, item 32 or 33 Polypeptides as described. (Item 35) The aforementioned heterogeneous portion is an immunoglobulin Fc domain, according to any one of items 1 to 34. The polypeptide described. (Item 36) The immunoglobulin Fc domain is a human immunoglobulin Fc domain, item 3 Polypeptides as described in 5. (Item 37) The heterogeneous portion includes an amino acid sequence that is at least 80% identical to sequence number 20. Polypeptides as described in item 35. (Item 38) The heterogeneous portion includes an amino acid sequence that is at least 90% identical to sequence number 20. Polypeptides as described in item 35. (Item 39) The aforementioned heterogeneous portion includes an amino acid sequence that is at least 95% identical to sequence number 20. Polypeptides as described in item 35. (Item 40) The aforementioned heterogeneous portion includes an amino acid sequence that is at least 97% identical to sequence number 20. Polypeptides as described in item 35. (Item 41) The heterogeneous portion comprises the amino acid sequence of SEQ ID NO: 20, as described in item 35 of the polypeptide. . (Item 42) The linker is less than 25 amino acids in length, as described in any one of items 1 to 41. polypeptide. (Item 43) The linker is one of items 1 to 41, with a length between 10 and 25 amino acids. Polypeptides as described. (Item 44) The linker is one of items 1 to 41, with a length of 15 to 25 amino acids. Polypeptides as described. (Item 45) The linker is one of items 1 to 41, with a length between 17 and 22 amino acids. Polypeptides as described. (Item 46) The linker is 21 amino acids long, as described in any one of items 1 to 41. Lipeptide. (Item 47) The aforementioned linker, (GGGGS) n Any of items 1 to 41 that include (where n = ≥ 2 in the formula) A polypeptide as described in item 1. (Item 48) The aforementioned linker, (GGGGS) nAny of items 1 to 41 that include (where n = ≥ 3 in the formula) A polypeptide as described in item 1. (Item 49) The aforementioned linker, (GGGGS) n Any of items 1 to 41 that include (where n = ≥ 4 in the formula) A polypeptide as described in item 1. (Item 50) The aforementioned linker, (GGGGS) n Any of items 1 to 44 that include (where n ≠ ≥ 5 in the formula) A polypeptide as described in item 1. (Item 51) The linker includes the amino acid sequence of SEQ ID NO: 21 in any one of items 1 to 50. The polypeptide described. (Item 52) The linker contains one of the amino acid sequences from sequence numbers 4 to 7, items 1 to 45. A polypeptide as described in any one of the following items. (Item 53) The linker includes the amino acid sequence of SEQ ID NO: 6, as described in any one of items 1 to 52. Polypeptide. (Item 54) Items containing an amino acid sequence that is at least 80% identical to the amino acid sequence of Sequence ID No. 11. A polypeptide as described in any one of items 1 to 53. (Item 55) Items containing an amino acid sequence that is at least 90% identical to the amino acid sequence of Sequence ID No. 11. A polypeptide as described in any one of items 1 to 53. (Item 56) Items containing an amino acid sequence that is at least 95% identical to the amino acid sequence of Sequence ID No. 11. A polypeptide as described in any one of items 1 to 53. (Item 57) The polypeptide described in any one of items 1 to 53, including the amino acid sequence of SEQ ID NO: 11 Do. (Item 58) Amino acids that are at least 80% identical to the amino acid sequence of SEQ ID NOs: 13, 53, or 56 A polypeptide containing an array, as described in any one of items 1 through 53. (Item 59) Amino acids that are at least 90% identical to the amino acid sequence of SEQ ID NOs: 13, 53, or 56. A polypeptide containing an array, as described in any one of items 1 through 53. (Item 60) Amino acids that are at least 95% identical to the amino acid sequence of SEQ ID NOs: 13, 53, or 56 A polypeptide containing an array, as described in any one of items 1 through 53. (Item 61) The amino acid sequence of sequence number 13, 53, or 56 is included in any one of items 1-53. The polypeptide described. (Item 62) Items containing an amino acid sequence that is at least 80% identical to the amino acid sequence of Sequence ID No. 15. A polypeptide as described in any one of items 1 to 53. (Item 63) Items containing an amino acid sequence that is at least 90% identical to the amino acid sequence of Sequence ID No. 15. A polypeptide as described in any one of items 1 to 53. (Item 64) Items containing an amino acid sequence that is at least 95% identical to the amino acid sequence of Sequence ID No. 15. A polypeptide as described in any one of items 1 to 53. (Item 65) The polypeptide described in any one of items 1 to 53, including the amino acid sequence of SEQ ID NO: 15 Do. (Item 66) The TβRII polypeptide does not contain amino acids 185-592 of SEQ ID NO: 2. A polypeptide as described in any one of items 1 to 65. (Item 67) The TβRII polypeptide mentioned above does not contain amino acids 1-22 of SEQ ID NO: 2, item 1- A polypeptide as described in any one of paragraphs 66. (Item 68) a) The amino acid sequence of SEQ ID NO: 18 and at least 85%, 90%, 95%, 97% or Contains amino acid sequences that are 99% identical, and are 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1 The TβRII polypeptide moiety, which contains one or fewer additional amino acids, b) The amino acid sequence of SEQ ID NO: 6 and at least 85%, 90%, 95%, 97%, or 9% It contains 9% identical amino acid sequences and 5, 4, 3, 2, or 1 or fewer additional amino acids. Includes the linker section, c) The amino acid sequence of SEQ ID NO: 20 and at least 85%, 90%, 95%, 97% or Contains amino acid sequences that are 99% identical, and are 25, 20, 15, 10, 5, 4, 3, 2 or A heterogeneous portion containing one or fewer additional amino acids, d) Leader array (e.g., array 23) as needed Polyp Petit de. (Item 69) a) Containing the amino acid sequence of SEQ ID NO: 18, 10, 9, 8, 7, 6, 5, 4, 3, 2 The TβRII polypeptide moiety contains one or fewer additional amino acids, b) Containing the amino acid sequence of SEQ ID NO: 6, with 5, 4, 3, 2 or 1 or fewer additional amino acids The linker portion contains acid, c) Containing the amino acid sequence of SEQ ID NO: 20, 25, 20, 15, 10, 5, 4, 3, 2 or a heterogeneous portion containing one or fewer additional amino acids, d) Leader array (e.g., array 23) as needed Polyp Petit de. (Item 70) a) The extracellular domain of the TβRII portion, which has at least 85 sequences from sequence number 18. The extracellular domain containing amino acid sequences that are identical by %, 90%, 95%, 97%, or 99% In and b) At least 85%, 90%, 95%, 97%, or 99% of the sequence of sequence number 20 is identical to that sequence. A heterologous region containing a single amino acid sequence, c) The linker is at least 85%, 90%, 95%, 9% of the amino acid sequence of SEQ ID NO: 6 The extracellular domain and the heterogeneous portion include amino acid sequences that are 7% or 99% identical. The linker part to connect and A polypeptide, including any one of items 1 through 69. (Item 71) a) The extracellular domain of the TβRII portion, which includes the amino acid sequence of SEQ ID NO: 18 The extracellular domain and b) A heterologous portion containing the amino acid sequence of Sequence ID No. 20, c) The linker contains the amino acid sequence of SEQ ID NO: 6, the extracellular domain and the heterogene Linker section that connects parts and A polypeptide, including any one of items 1 through 70. (Item 72) The amino acid sequence of SEQ ID NO: 48 and at least 85%, 90%, 95%, 97%, or 99% A polypeptide described in any one of items 1 to 71, containing an amino acid sequence that is % identical. (Item 73) The polypeptide described in any one of items 1 to 72, including the amino acid sequence of SEQ ID NO: 48 Do. (Item 74) The Po in item 73, which does not include the leader sequence, or has the leader sequence removed. Lipeptide. (Item 75) Glycosylated amino acids, pegylated amino acids, farnesylated amino acids, acetylated amino acids Acids, biotinylated amino acids, amino acids conjugated to lipid substructures, and organic inducers One or more modified amino acids selected from amino acids conjugated to a conductive agent. A polypeptide containing an acid residue, as described in any one of items 1 to 74. (Item 76) A glycosylated polypeptide as described in item 75. (Item 77) Having a glycosylation pattern specific to the expression of the polypeptide in CHO cells, A polypeptide listed in any one of items 1 through 75. (Item 78) Equilibrium dissociation constant (K) less than 100 pM D ) which binds to human TGFβ1, items 1-77 A polypeptide as described in any one of the items. (Item 79) Equilibrium dissociation constant (K) less than 75 pM D ) Binds to human TGFβ1, items 1-78 A polypeptide as described in item one. (Item 80) Equilibrium dissociation constant (K) less than 60 pM D ) Binds to human TGFβ3, items 1-79 A polypeptide as described in item one. (Item 81) Equilibrium dissociation constant (K) less than 50 pM D ) Binds to human TGFβ3, items 1-80 A polypeptide as described in item one. (Item 82) Determined using a reporter gene assay, IC2 is less than 1.0 nM. 50 TGFβ A polypeptide that inhibits item 1, as described in any one of items 1 to 81. (Item 83) Determined using a reporter gene assay, IC2 is less than 0.25 nM. 50 TGF A polypeptide that inhibits β1, as described in any one of items 1 to 81. (Item 84) Determined using a reporter gene assay, IC2 is less than 0.1 nM. 50 TGFβ A polypeptide that inhibits item 1, as described in any one of items 1 to 81. (Item 85) Determined using a reporter gene assay, IC2 is less than 0.05 nM. 50 TGF A polypeptide that inhibits β1, as described in any one of items 1 to 81. (Item 86) Determined using a reporter gene assay, IC2 is less than 0.3 nM. 50 TGFβ A polypeptide described in any one of items 1 to 81 that inhibits item 3. (Item 87) Determined using a reporter gene assay, IC2 is less than 0.1 nM. 50 TGFβ A polypeptide described in any one of items 1 to 81 that inhibits item 3. (Item 88) Determined using a reporter gene assay, IC2 is less than 0.05 nM. 50 TGF A polypeptide that inhibits β3, as described in any one of items 1 to 81. (Item 89) Determined using a reporter gene assay, IC2 is less than 0.04 nM. 50 TGF A polypeptide that inhibits β3, as described in any one of items 1 to 81. (Item 90) The reporter gene assay is the CAGA reporter assay, items 82-8 A polypeptide as described in any one of item 8. (Item 91) A homodimer containing two polypeptides listed in any one of items 1 through 90. (Item 92) An isolated polypeptide containing the coding sequence of any one of items 1 to 90. Renucleotide. (Item 93) A set comprising a promoter sequence operably linked to a polynucleotide as described in item 92. Substitute polynucleotide. (Item 94) At least 80%, 85%, or 90% of any one of sequence numbers 10, 12, or 14, Items 92 or 93 contain nucleotide sequences that are 95%, 97%, or 100% identical. Polynucleotides as described above. (Item 95) Cells transformed with any one of the polynucleotides described in items 92-94. (Item 96) A mammalian cell, as described in item 95. (Item 97) Cells as described in item 95, which are CHO cells or human cells. (Item 98) A polypeptide described in any of items 1 to 90 or a homodimer described in item 91, A pharmaceutical product containing pharmaceutically acceptable excipients. (Item 99) A polypeptide or its homodimer described in any one of items 72 to 74, and a pharmaceutical A pharmaceutical product containing an excipient that is permissible in the field. (Item 100) A method for modulating the cellular response to TGFβ superfamily members. This refers to a polypeptide listed in any of items 1-90 or a homodimer listed in item 91. A method comprising exposing the cells to the method. (Item 101) In patients who require it, diseases related to TGFβ superfamily members or a method for treating a condition, wherein the polypeptide described in any one of items 1 to 90 A method comprising administering an effective amount of the homodimer described in item 91 to the patient. (Item 102) The item is that the TGFβ superfamily member is TGFβ1 or TGFβ3. The method described in item 101. (Item 103) The method according to item 100 or 101, wherein the disease or condition is cancer. (Item 104) The aforementioned cancers include stomach cancer, intestinal cancer, skin cancer, breast cancer, melanoma, bone cancer, and thyroid cancer. The method described in item 102, selected from the options provided. (Item 105) The disease or condition is a fibrous or sclerotic disease or condition, item 100 or The method described in 101. (Item 106) The aforementioned fibrous or sclerotic disease or condition is scleroderma, lupus erythematosus, pulmonary fibrosis, etc. Arteriosclerosis, hepatic fibrosis, diffuse systemic sclerosis, glomerulonephritis, nerve scarring, dermis Selected from scarring, radiation-induced fibrosis, hepatic fibrosis, idiopathic pulmonary fibrosis, and myelofibrosis. The method described in item 105. (Item 107) The method according to item 106, wherein the disease or condition is myelofibrosis. (Item 108) The aforementioned disease or condition is primary myelofibrosis, myelofibrosis after polycythemia eucarcosis, and The method described in item 107, selected from the group consisting of myelofibrosis following essential thrombocythemia. (Item 109) The aforementioned disease or condition, according to the International Prognostic Scoring System (IPSS), Alternatively, according to Dynamic IPSS (DIPSS), low risk, intermediate-1 risk, intermediate-2 risk. The method described in item 107, selected from the group consisting of, or high-risk myelofibrosis. (Item 110) The method according to item 106, wherein the disease or condition is idiopathic pulmonary fibrosis. (Item 111) The method according to item 100 or 101, wherein the disease or condition is a heart disease. (Item 112) The aforementioned disease or condition is hereditary hemorrhagic telangiectasia (HHT), Marfan syndrome, Loeys-Dietz syndrome, familial thoracic aortic aneurysm syndrome, tortuosomatic artery syndrome, pre-eclampsia, ate Items selected from Rohm's arteriosclerosis, restenosis, and hypertrophic cardiomyopathy / congestive heart failure. The method described in 100 or 101. (Item 113) The method according to item 100 or 101, wherein the disease or condition is pulmonary hypertension. (Item 114) The aforementioned pulmonary hypertension is classified as Class I, Class II, or Class I by the World Health Organization. The method described in item 113 for pulmonary hypertension of class II or class IV. (Item 115) If the aforementioned disease or condition is a kidney-related disease or condition, see item 100 or 101. Method of loading. (Item 116) The aforementioned kidney-related disease or condition is chronic kidney disease (or failure), acute kidney disease (or (Kidney failure), primary kidney disease, non-diabetic kidney disease, glomerulonephritis, interstitial nephritis, diabetic kidney Diseases, diabetic nephropathy, glomerulosclerosis, rapidly progressive glomerulonephritis, renal fibrosis, Alport syndrome Group, IDDM nephritis, mesangial proliferative glomerulonephritis, membranoproliferative glomerulonephritis, crescent formation Glomerulonephritis, interstitial renal fibrosis, focal segmental glomerulosclerosis, membranous nephropathy, minimal change nephropathy -se syndrome, microimmune rapidly progressive glomerulonephritis, IgA nephropathy, polycystic kidney disease, Dent's disease, Renal cystinosis, Hymann's nephritis, autosomal dominant (adult) polycystic kidney disease, autosomal recessive (childhood) ) Polycystic kidney disease, acute kidney injury, nephrotic syndrome, renal ischemia, glomerular epithelial cell disease or Disorders, proteinuria, glomerular disorders, membranous glomerulonephritis, focal segmental glomerulonephritis, pre-eclampsia, eclampsia, Kidney disorders, collagen disease vascular disorders, benign orthostatic (postural) proteinuria, IgM nephropathy, membranous nephropathy, Lucoidosis, diabetes mellitus, drug-induced kidney damage, Fabry disease, aminoaciduria, f Fanconi syndrome, hypertensive nephrosclerosis, interstitial nephritis, sickle cell anemia, hemoglobinuria, Myoglobinuria, Wegener's granulomatosis, glycogen storage disease type 1, chronic kidney disease, chronic renal failure, hypotrichosis Globular filtration rate (GFR), renal vascular sclerosis, lupus nephritis, ANCA positive microimmune crescent shape Adult glomerulonephritis, chronic allograft nephropathy, nephrotoxicity, kidney necrosis, kidney injury, glomerulos Renal tubular injury, renal dysfunction, nephritis syndrome, acute renal failure, chronic renal failure, proximal fallopian tube dysfunction Acute kidney transplant rejection, chronic kidney transplant rejection, non-IgA mesangial proliferative glomerulonephropathy Inflammation, post-infectious glomerulonephritis, vasculitis associated with all types of kidney disorders, all hereditary kidney diseases, All kinds of interstitial nephritis, kidney transplant failure, kidney cancer, and other conditions (e.g., hypertension, diabetes and autoimmune diseases) Autoimmune diseases) related kidney diseases, Dent's disease, nephrogenic cystinosis, Hymann's nephritis, primary kidney disease Diseases, collapsed glomerulosis, dense deposit disease, cryoglobulinemia-associated glomerulonephritis, Ch. Schönlein disease, post-infectious glomerulonephritis, bacterial endocarditis, microscopic polyangiitis, Jarg-Strauss syndrome, anti-GBM antibody-mediated glomerulonephritis, amyloidosis, monoclonal claw Streptococcal immunoglobulinemia, fibrillary glomerulonephritis, immunotactoid glomerulopathy, ischemic urine Tubulointerstitial nephritis, drug-induced tubulointerstitial nephritis, toxic tubulointerstitial nephritis, infectious tubulointerstitial kidney Inflammation, bacterial pyelonephritis, polyomavirus infection, or HIV infection resulting in the virus Russian infectious tubulointerstitial nephritis, metabolic disease-induced tubulointerstitial disease, mixed connective tissue disease, casts Crystalline nephropathy, which can result from urate or oxalate or drug-induced crystal deposition. Acute cellular tubulointerstitial allograft rejection, lymphoma, or post-transplant lymphoproliferative disorder. As a result, tumor-infiltrating diseases, obstructive kidney disease, vascular diseases, thrombotic microangiopathy, kidney Vascular sclerosis, atherothrombotic disease, mixed connective tissue disease, polyarteritis nodosa, calcium stenosis Phosphate inhibitor-induced vascular disease, acute cellular vascular allograft rejection, acute humoral allograft rejection Rejection reaction, early renal failure (ERFD), end-stage renal disease (ESRD), renal vein thrombosis, acute Tubular necrosis, acute interstitial nephritis, pre-existing chronic kidney disease, renal artery stenosis, ischemic nephropathy, uremia, Drug and toxin-induced chronic tubulointerstitial nephritis, reflux nephropathy, kidney stones, Goodpasture disease Syndrome, normocytic normochromic anemia, renal anemia, diabetic chronic kidney disease, IgG4-related disease, phosphatiditis Nhippel-Lindau syndrome, tuberous sclerosis, nephronophthisis, renal medullary cystopathy, renal cell carcinoma, glandular Cancer, nephroblastoma, lymphoma, leukemia, hyposialylation disorder, chronic cyclosporine nephropathy, renal reperfusion Injury, renal dysplasia, hyperuremia, bilateral arterial occlusion, acute uric nephropathy, hypovolemia, acute bilateral occlusion Obstructive urinary tract disease, hypercalcemia nephropathy, hemolytic uremic syndrome, acute urinary retention, malignant nephrosclerosis Postpartum glomerulosclerosis, scleroderma, non-Goodpasture anti-GBM disease, microscopic nodular polyderma Arteritis, allergic granulomatosis, acute radiation nephritis, post-streptococcal glomerulonephritis, Walden Ström's macroglobulinemia, analgesic nephropathy, arteriovenous fistula, arteriovenous graft, dialysis, dysphagia Topical kidney, cavernous kidney, renal osteodystrophy, mononephropathy, hydronephrosis, microalbuminuria, uremia, Hematuria, hyperlipidemia, hypoalbuminemia, lipiduria, acidosis, hyperkalemia, and The method described in item 115, selected from the group consisting of edema. (Item 117) The method according to item 115, wherein the aforementioned kidney-related disease or condition is chronic kidney disease.

Claims

[Claim 1] The invention described in the specification.

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