JP2018020980A - Agent and composition for prevention or treatment of metabolic bone disease - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide an agent and composition for the prevention or treatment of metabolic bone disease, that can effectively prevent or treat metabolic bone disease with bone loss.SOLUTION: The present invention provides an agent and composition for the prevention or treatment of metabolic bone disease with bone loss that contains latent Transforming growth factor (TGF)-β3 or latent TGF-β1 as an active ingredient.SELECTED DRAWING: None

Description

  The present invention relates to a preventive or therapeutic agent for bone metabolic diseases accompanied by a decrease in bone mass, and a composition for preventing or treating bone metabolic diseases accompanied by a decrease in bone mass.

  Osteoporosis is a disease in which the risk of fracture increases because osteoblast activity cannot catch up with excessive activation of osteoclasts, resulting in decreased bone mass. Currently, female hormones, calcitonin, bisphosphonate, ipriflavone, raloxifene, vitamin K2, vitamin D and the like are used as osteoporosis therapeutic agents. However, since these osteoporosis therapeutic agents are not locally effective, side effects may be problematic.

  For example, administration of female hormones may increase the incidence of breast cancer, endometrial cancer and the like. In addition, administration of calcitonin may cause facial flushing, nausea, allergic reactions, and the like. In addition, administration of bisphosphonate may cause jaw osteonecrosis, jaw osteomyelitis and the like. In addition, administration of ipriflavone may cause heartbeat, nausea, abdominal pain and the like. In addition, administration of raloxifene may cause hot flashes, nausea, leg cramps, and the like. Also, it may be dangerous to administer vitamin K2 to patients with myocardial infarction. Moreover, administration of vitamin D may result in excessive blood calcium.

  Meanwhile, transforming growth factor (TGF) -β is a kind of cytokine produced by almost all cells in the living body such as kidney, bone marrow, and platelets. It is known that TGF-β has an inactive latent form and an active form having activity, and the activity is regulated by conversion from the latent form to the active form. Moreover, it is known that TGF-β has five types of subtypes (β1 to β5). TGF-β possessed by mammals is TGF-β1, TGF-β2, and TGF-β3 among five subtypes.

  There are many unclear points about the function of TGF-β in bone metabolism. For example, it has been reported that TGF-β acts in an inhibitory manner on bone resorption and osteoclast formation, and at the same time, reports that it acts in an accelerated manner (see, for example, Non-Patent Document 1).

Yan T, Riggs BL, Boyle WJ, Khosla S. 2001. Regulation of osteoclastogenesis and RANK expression by TGF-beta1.J Cell Biochem 83: 320-5.

  An object of the present invention is to provide a preventive or therapeutic agent that can effectively prevent or treat a bone metabolic disease accompanied by a decrease in bone mass with few side effects.

The present invention includes the following aspects.
(1) A preventive or therapeutic agent for bone metabolic diseases accompanied by a decrease in bone mass, comprising latent TGF-β1 or latent TGF-β3 as an active ingredient.
(2) It contains latent TGF-β1, and when the latent TGF-β1 contains any one of the following amino acid sequences (a) to (c) and is converted to an active form, TGF-β The preventive or therapeutic agent according to (1), which is a protein having an activity of transmitting a signal.
(A) Amino acid sequence described in SEQ ID NO: 1 (b) Amino acid sequence in which one or more amino acids are deleted, substituted or added in the amino acid sequence described in SEQ ID NO: 1 (c) described in SEQ ID NO: 1 An amino acid sequence having a sequence identity of 80% or more with the amino acid sequence of (3) latent TGF-β3, wherein the latent TGF-β3 is any one of the following amino acid sequences (d) to (f): And a prophylactic or therapeutic agent according to (1), which is a protein having an activity of transmitting a TGF-β signal when converted into an active form.
(D) Amino acid sequence represented by SEQ ID NO: 3 (e) Amino acid sequence in which one or more amino acids are deleted, substituted, or added in the amino acid sequence represented by SEQ ID NO: 3 (f) SEQ ID NO: 3 (4) the bone metabolic disease is osteoporosis, metastatic bone tumor, multiple myeloma, hypercalcemia due to malignant tumor or deformability The prophylactic or therapeutic agent according to any one of (1) to (3), which is osteoarthritis.
(5) A composition for preventing or treating a bone metabolic disease accompanied by a decrease in bone mass, comprising the preventive or therapeutic agent according to any one of (1) to (4) and a pharmaceutically acceptable carrier. .

  According to the present invention, it is possible to provide a preventive or therapeutic agent that can effectively prevent or treat a bone metabolic disease accompanied by a decrease in bone mass with few side effects.

(A) is the microscope picture which shows the result of the TRAP dyeing | staining which compared the effect | action with respect to osteoclast formation of TGF- (beta) 1-3 in Experimental example 1. FIG. (B) is the graph which quantified the dyeing | staining result of (a). (A) And (b) is a graph which shows the result of Experimental example 3. FIG. (A) And (b) is a micro CT image which shows the result of administering latent type TGF-β1-Fc to a bone destruction model mouse in Experimental Example 4. (C) is a graph which shows the result of having digitized the number of absorption pits confirmed by the images of (a) and (b). In Experimental example 4, it is a fluorescence micrograph which shows the result of having immunostained with the antibody with respect to the cathepsin K which is a marker of an osteoclast, after administering latent type TGF- (beta) 1-Fc to the bone destruction model mouse. (A) And (b) is a micro CT image which shows the result of administering latent type TGF-β3-Fc to a bone destruction model mouse in Experimental Example 5.

[Prophylactic or therapeutic agent]
In one embodiment, the present invention provides a preventive or therapeutic agent for bone metabolic diseases accompanied by a decrease in bone mass, comprising latent TGF-β1 or latent TGF-β3 as an active ingredient.

  It is known that mammalian TGF-β has subtypes of TGF-β1, TGF-β2, and TGF-β3. It is known that TGF-β is biosynthesized as latent TGF-β and undergoes intramolecular cleavage to become active TGF-β.

  Latent TGF-β is a protein that does not have TGF-β activity and exhibits activity to transmit a TGF-β signal when activated and converted to active TGF-β. The activity of transmitting a TGF-β signal can be measured, for example, by detecting the activation of Smad existing downstream of the TGF-β receptor. Latent TGF-β is known not to have the activity of transmitting TGF-β signals.

  As will be described later in Examples, the inventors have shown that bone destruction can be suppressed in vivo by administering latent TGF-β1 or latent TGF-β3 to a bone destruction model mouse. Therefore, latent TGF-β1 or latent TGF-β3 can be used as a prophylactic or therapeutic agent for bone metabolic diseases accompanied by bone loss.

  In the preventive or therapeutic agent of the present embodiment, being effective for the prevention or treatment of bone metabolic diseases accompanied by a decrease in bone mass means, for example, alleviation of symptoms of the disease, prevention of progression, or prevention of disease onset. Or more than one is achieved. Whether it is effective for the prevention or treatment of bone metabolic diseases accompanied by bone loss is, for example, osteoclast formation inhibitory effect, bone resorption inhibitory effect by osteoclasts, bone formation promoting effect, or It can be determined by having one or more of the actions such as the whole body or local bone mass increasing action.

  Specific examples of latent TGF-β1 include a protein consisting of the amino acid sequence set forth in SEQ ID NO: 1 or 2. The protein consisting of the amino acid sequence shown in SEQ ID NO: 1 is human latent TGF-β1. Examples of the nucleic acid encoding the protein consisting of the amino acid sequence shown in SEQ ID NO: 1 include the nucleic acid consisting of the base sequence shown in SEQ ID NO: 5.

  The protein consisting of the amino acid sequence set forth in SEQ ID NO: 2 is mouse latent TGF-β1. Examples of the nucleic acid encoding the protein consisting of the amino acid sequence shown in SEQ ID NO: 2 include a nucleic acid consisting of the base sequence represented by SEQ ID NO: 6.

  As will be described later in the Examples, it became clear that latent TGF-β1-Fc, which is a fusion protein of latent TGF-β1 and immunoglobulin constant region (Fc), can suppress bone destruction in vivo. It was. Therefore, any protein or polypeptide may be added to the latent TGF-β1. Here, examples of the protein or polypeptide that may be added include an Fc tag, a histidine tag, a GST tag, a GFP tag, a myc tag, and a FLAG tag.

  The latent TGF-β1 may have a mutation with respect to the amino acid sequence described in SEQ ID NO: 1 as long as it is a protein having an activity of transmitting a TGF-β signal when converted to an active form.

Specific examples of the latent TGF-β1 include a protein having an amino acid sequence of any of the following (a) to (c) and having an activity of transmitting a TGF-β signal when converted to an active form: Can be mentioned.
(A) amino acid sequence described in SEQ ID NO: 1 (b) amino acid sequence described in SEQ ID NO: 1, wherein one or more amino acids are deleted, substituted or added (c) described in SEQ ID NO: 1 Amino acid sequence whose sequence identity is 80% or more

  In this specification, 1 or a plurality may be 1-10 pieces, for example, may be 1-5 pieces, for example, may be 1-4 pieces, for example, 1-3 pieces, for example. For example, it may be 1-2.

  In the present specification, the sequence identity of amino acid sequences may be 80% or more, 85% or more, 90% or more, 95% or more, It may be 98% or more.

Here, the sequence identity of the target amino acid sequence with respect to the reference amino acid sequence can be determined, for example, as follows. First, the reference amino acid sequence and the target amino acid sequence are aligned. Here, a gap may be included in each amino acid sequence so as to maximize the sequence identity. Subsequently, in the reference amino acid sequence and the target amino acid sequence, the number of matched amino acids is calculated, and the sequence identity can be obtained according to the following formula.
Sequence identity (%) = number of matched amino acids / total number of amino acids in the subject amino acid sequence × 100

  Specific examples of latent TGF-β3 include a protein consisting of the amino acid sequence set forth in SEQ ID NO: 3 or 4. The protein consisting of the amino acid sequence set forth in SEQ ID NO: 3 is human latent TGF-β3. Examples of the nucleic acid encoding the protein consisting of the amino acid sequence shown in SEQ ID NO: 3 include a nucleic acid consisting of the base sequence represented by SEQ ID NO: 7.

  The protein consisting of the amino acid sequence set forth in SEQ ID NO: 4 is murine latent TGF-β3. Examples of the nucleic acid encoding the protein consisting of the amino acid sequence shown in SEQ ID NO: 4 include a nucleic acid consisting of the base sequence represented by SEQ ID NO: 8.

  The latent TGF-β3 may have a mutation with respect to the amino acid sequence described in SEQ ID NO: 3 as long as it is a protein having an activity of transmitting a TGF-β signal when converted to an active form.

As a specific example of latent TGF-β3, a protein having an amino acid sequence of any of the following (d) to (f) and having an activity of transmitting a TGF-β signal when converted to an active form is Can be mentioned. Here, the sequence identity of one or more and amino acid sequences is the same as described above.
(D) The amino acid sequence described in SEQ ID NO: 3 (e) The amino acid sequence described in SEQ ID NO: 3 wherein one or more amino acids are deleted, substituted, or added (f) described in SEQ ID NO: 3 Amino acid sequence whose sequence identity is 80% or more

  Examples of bone metabolic diseases accompanied by bone loss include diseases such as osteoporosis, metastatic bone tumors, multiple myeloma, hypercalcemia due to malignant tumors, and osteoarthritis. Bone loss may be systemic, such as osteoporosis, or local, such as metastatic bone tumors.

  According to the preventive or therapeutic agent of the present embodiment, it is possible to effectively prevent or treat a bone metabolic disease accompanied by a decrease in bone mass.

  Latent TGF-β1 and latent TGF-β3 are converted to active forms by the action of osteoclasts. The conversion from latent to active forms of TGF-β1 and TGF-β3 is probably caused by the acidic conditions around the osteoclasts.

  Conventionally, the effect of TGF-β on osteoclasts has been reported to both promote osteoclast formation and suppress osteoclast formation. For this reason, it was unclear whether TGF-β has a preventive or therapeutic effect on bone metabolic diseases accompanied by bone loss. In addition, as described later in Examples, in osteoclast primary culture in a medium containing latent TGF-β, the significance of the osteoclast inhibitory effect was not recognized.

  In contrast, as described later in the Examples, the inventors have clarified that bone destruction can be suppressed in vivo by administering latent TGF-β1 or latent TGF-β3 to a bone destruction model mouse. . Such a result is difficult for those skilled in the art to predict.

  Bone is kept homeostatic due to the balance between bone resorption by osteoclasts and remodeling in which bone formation by osteoblasts occurs in the bone resorbed by osteoclasts. Conventionally, there is no preventive or therapeutic agent for bone metabolic diseases using this remodeling mechanism.

  According to the prophylactic or therapeutic agent of the present embodiment, it exhibits an action in accordance with the above-described remodeling mechanism, and can exhibit activity only in a local area where bone is resorbed by osteoclasts, thereby suppressing bone resorption. There are few side effects on other tissues and cells. Moreover, according to the preventive or therapeutic agent of the present embodiment, not only the suppression of bone resorption but also the effect of promoting bone formation by active TGF-β can be expected.

[Prophylactic or therapeutic composition]
In one embodiment, the present invention provides an agent for preventing or treating a bone metabolic disease with bone loss, comprising the above-described preventive or therapeutic agent for a bone metabolic disease with bone loss, and a pharmaceutically acceptable carrier. Prophylactic or therapeutic compositions are provided.

  The preventive or therapeutic composition of the present embodiment is preferably formulated into a dosage form used parenterally. Examples of dosage forms used parenterally include injections, ointments, patches and the like.

  As the pharmaceutically acceptable carrier, those usually used for preparations can be used without particular limitation. More specifically, for example, binders such as gelatin, corn starch, tragacanth gum and gum arabic; excipients such as starch and crystalline cellulose; swelling agents such as alginic acid; solvents for injection such as water, ethanol and glycerin; Examples thereof include adhesives such as rubber adhesives and silicone adhesives.

  The therapeutic composition may contain additives. Additives include lubricants such as magnesium stearate; sweeteners such as sucrose, lactose and saccharin; flavoring agents such as peppermint and red mono oil; stabilizers for benzyl alcohol and phenol; buffers such as phosphate and sodium acetate Agents; solubilizing agents such as benzyl benzoate and benzyl alcohol; antioxidants; preservatives and the like.

  The therapeutic composition can be formulated by combining the above pharmaceutically acceptable carriers and additives as appropriate and admixing them in a generally accepted unit dosage form.

  The dosage of the therapeutic composition varies depending on the dosage form of the therapeutic composition, the patient's symptoms, body weight, age, sex, etc., and cannot be generally determined. For example, in the case of injections, the dosage per dosage unit form 0.01 to 50 mg of active ingredient (latent TGF-β1 or latent TGF-β3) may be administered by subcutaneous injection, intravenous injection, infusion or the like.

  In addition, the daily dose of the therapeutic composition varies depending on the patient's symptoms, body weight, age, sex, etc., and cannot be determined unconditionally. For example, an adult daily dose of 0.005 to 100 mg / kg body weight The active ingredient may be divided and administered about once a day to 6 months.

[Other Embodiments]
In one embodiment, the present invention provides for the prevention of bone metabolic disorders with bone loss comprising administering an effective amount of latent TGF-β1 or latent TGF-β3 to a patient in need of treatment. Alternatively, a method of treatment is provided. Examples of the latent TGF-β1 or the latent TGF-β3 include those described above.

  In one embodiment, the present invention provides latent TGF-β or latent TGF-β3 for the prevention or treatment of bone metabolic diseases associated with bone loss. Examples of the latent TGF-β1 or the latent TGF-β3 include those described above.

  In one embodiment, the present invention provides the use of latent TGF-β1 or latent TGF-β3 for the manufacture of a prophylactic or therapeutic agent for bone metabolic diseases associated with bone loss. Examples of the latent TGF-β1 or the latent TGF-β3 include those described above.

  Next, although an experiment example is shown and this invention is demonstrated in detail, this invention is not limited to the following experiment examples.

[Experimental Example 1]
(Comparison of action of activated mouse TGF-β1 to 3 on osteoclast formation)
Osteoclast progenitor cells obtained from C57 / BL6 background wild-type mice are sometimes referred to as medium (hereinafter referred to as “M”) containing macrophage colony stimulating factor (M-CSF, R & D) at a final concentration of 50 ng / mL. ), A medium containing M-CSF with a final concentration of 50 ng / mL and a receptor activator of nuclear factor kappa B ligand (RANKL, Pepro Tech) with a final concentration of 25 ng / mL (hereinafter sometimes referred to as “M + R”). ), Medium containing a final concentration of 50 ng / mL M-CSF, a final concentration of 25 ng / mL RANKL and a final concentration of 200 ng / mL recombinant BMP2 (Pepro Tech) (hereinafter referred to as “M + R + BMP2”) A medium containing M-CSF having a final concentration of 50 ng / mL, RANKL having a final concentration of 25 ng / mL, and active TGF-β1 (R & D) having a final concentration of 10 ng / mL (hereinafter referred to as “M + R + TGFβ1”). ), Medium containing M-CSF with a final concentration of 50 ng / mL, RANKL with a final concentration of 25 ng / mL, and activated TGF-β2 (R & D) with a final concentration of 10 ng / mL (hereinafter referred to as “M + R + TGFβ2”) Or a medium containing M-CSF with a final concentration of 50 ng / mL, RANKL with a final concentration of 25 ng / mL and active TGF-β3 (R & D) with a final concentration of 10 ng / mL (hereinafter referred to as “M + R + TGFβ3”) Incubated for 5 days. Thereafter, tartrate-resistant acid phosphatase (TRAP), which is known as an osteoclast marker, was stained with a commercially available kit (Sigma) to evaluate the degree of osteoclast formation.

  FIG. 1 (a) is a photomicrograph showing the results of TRAP staining of each cell after 5 days of culture. FIG. 1B is a graph showing the results of measuring the number of TRAP-positive multinucleated cells based on FIG. The presence or absence of a significant difference was examined by Student t test. “*” Indicates that there is a significant difference at P <0.05, and “***” indicates that there is a significant difference at P <0.001. As a result, it was revealed that the formation of osteoclasts was significantly suppressed by adding active TGF-β1 or active TGF-β3 to the medium. On the other hand, it was revealed that osteoclast formation was significantly promoted by addition of active TGF-β2 to the medium.

[Experiment 2]
(Production of latent TGF-β-Fc)
A DNA fragment having a base sequence excluding the base corresponding to the stop codon of the base sequence (SEQ ID NO: 6) encoding the latent mouse TGF-β1 was ligated to an Fc fusion protein expression vector, and the latent TGF-β-Fc An expression vector was prepared. Subsequently, the latent TGF-β-Fc expression vector is transfected into COS7 cells to express the latent TGF-β1-Fc and purified using a protein G column to obtain the latent TGF-β1-Fc. It was.

  The latent TGF-β3-Fc was similar to the above except that the nucleotide sequence (SEQ ID NO: 8) encoding the latent mouse TGF-β3 was used instead of the DNA fragment encoding the latent mouse TGF-β1. Got.

[Experiment 3]
(Investigation of the activity of latent TGF-β1-Fc in vitro)
Bone marrow cells taken from the femur and tibia of C57 / BL6 background wild type mice were added to α-MEM medium containing 10% (v / v) serum and M-CSF to a final concentration of 50 ng / mL. The cells were cultured for 72 hours in the added medium.

Thereafter, the adherent cells were collected, and as primary culture, a medium containing M-CSF having a final concentration of 50 ng / mL (hereinafter sometimes referred to as “M”), and M- with a final concentration of 50 ng / mL. Medium containing CSF and RANKL having a final concentration of 25 ng / mL (hereinafter sometimes referred to as “MR”), M-CSF having a final concentration of 50 ng / mL, RANKL having a final concentration of 25 ng / mL, and a final concentration of 10 ng / mL Medium containing latent TGF-β1 (hereinafter sometimes referred to as “latent TGFβ1”), final concentration of 50 ng / mL M-CSF, final concentration of 25 ng / mL RANKL, and final concentration of 10 ng / mL activity medium containing type TGF? 1 (hereinafter, may be referred to as "active TGF? 1".) Now, 1 × 10 ⁵ cells dense cells / well In were cultured for 5 days were seeded in 96-well plates.

  After primary culture, RNA was collected from each cell, and expression of osteoclast markers cathepsin K (Cathepsin K, Ctsk) and nuclear factor of activated T cells 1 (NFATc1) was analyzed by real-time PCR.

  FIG. 2 (a) is a graph showing the expression levels of Ctsk and NFATc1 after primary culture. In FIG. 2 (a), the data standardize the average expression level of Ctsk or NFATc1 with the average expression level of β-actin and indicate the average value ± SD. The presence or absence of a significant difference was examined by Student t test. “**” indicates that there is a significant difference at P <0.01, “***” indicates that there is a significant difference at P <0.001, and “NS” indicates that there is no significant difference. Indicates.

  Subsequently, secondary culture was performed. Culture supernatant of MR medium in primary culture (hereinafter sometimes referred to as “MR SUP”) and culture supernatant of latent TGFβ1 medium (hereinafter sometimes referred to as “latent TGFβ1 SUP”). Adherent cells similar to those described above were cultured for 5 days using the medium.

  After secondary culture, RNA was collected from each cell, and expression of osteoclast markers, cathepsin K (Cathepsin K, Ctsk) and nuclear factor of activated T cells 1 (NFATc1) was analyzed by real-time PCR.

  Fig. 2 (b) is a graph showing the expression levels of Ctsk and NFATc1 after secondary culture. In FIG. 2 (b), the data standardize the average expression level of Ctsk or NFATc1 with the average expression level of β-actin, and show the average value ± SD. The presence or absence of a significant difference was examined by Student t test. “*” Indicates that there is a significant difference at P <0.05, and “***” indicates that there is a significant difference at P <0.001.

  As a result, in primary culture, the effect of latent TGF-β1 on osteoclast formation was not observed. However, significant suppression of osteoclast formation was observed in the secondary culture compared to the TGF-β1 non-added group. This result indicates that latent TGF-β1 was converted to active TGF-β1 by osteoclasts activated in primary culture.

[Experimental Example 4]
(Administration of latent TGF-β1-Fc to bone destruction model mouse)
Lipopolysaccharide (LPS) was administered intradermally on the calvaria at 5 μg / five female mice at 500 μg / mouse to prepare bone destruction model mice. As a control, phosphate buffer (PBS) was administered. Upon administration of LPS or PBS, 20 μg / mouse latent TGF-β1-Fc (hereinafter sometimes referred to as “TGFβ1-Fc”) or 20 μg / mouse CD4-Fc (control) were simultaneously administered. .

  On day 5 after administration, mice were sacrificed and bone destruction was evaluated by micro CT. Moreover, the calvaria was embedded in paraffin to prepare a tissue section, and cathepsin K (Cathepsin K, Ctsk), which is a marker of osteoclast, was detected by immunostaining.

  FIG. 3A is an image of micro CT acquired in Experimental Example 4, and FIG. 3B is an enlarged image of a part thereof. FIG. 3 (c) is a graph in which the number of resorption pits, which are depressions caused by bone resorption, is quantified. The number of absorbed pits is shown as a relative value where the number of absorbed pits in the LPS and CD4-Fc administration group is 1. The presence or absence of a significant difference was examined by Student t test. “**” indicates that there is a significant difference at P <0.01, and “***” indicates that there is a significant difference at P <0.001.

  As a result, it was revealed that bone destruction was significantly suppressed in vivo by administration of latent TGF-β1-Fc.

  FIG. 4 is a fluorescence micrograph showing the results of detecting cathepsin K (Cathepsin K, Ctsk), which is a marker of osteoclasts, by immunostaining a tissue section of the calvaria. As a control, the results of staining cell nuclei with 4 ', 6-diamidino-2-phenylindole (DAPI) are also shown. As a result, it was revealed that osteoclast formation was remarkably suppressed by latent TGF-β1-Fc administration.

[Experimental Example 5]
(Administration of latent TGF-β3-Fc to bone destruction model mice)
In the above Experimental Example 4, a latent TGF-β3-Fc (hereinafter referred to as “TGFβ3”) was prepared in the same manner as in Experimental Example 4 except that the latent TGF-β3-Fc was used instead of the latent TGF-β1-Fc. -Fc "), or CD4-Fc (control) was administered to bone destruction model mice, and bone destruction was evaluated by micro CT.

  FIG. 5A is an image of a micro CT acquired in Experimental Example 5, and FIG. 5B is an enlarged image of a part thereof.

  As a result, it was revealed that bone destruction was significantly suppressed in the bone destruction model mice administered with latent TGF-β3-Fc compared to the bone destruction model mice administered with CD4-Fc.

  From the above results, it is clear that by administering latent TGF-β1 in vivo, it is converted into active TGF-β1 only in the region where osteoclasts are activated and remarkably suppresses osteoclast formation. It became. It was also revealed that latent TGF-β3 has the same effect as latent TGF-β1.

  According to the present invention, it is possible to provide a preventive or therapeutic agent that can effectively prevent or treat a bone metabolic disease accompanied by a decrease in bone mass.

Claims (5)

  An agent for preventing or treating a bone metabolic disease accompanied by a decrease in bone mass, comprising latent transforming growth factor (TGF) -β1 or latent TGF-β3 as an active ingredient. A latent TGF-β1, which contains any one of the following amino acid sequences (a) to (c) and transmits a TGF-β signal when converted into an active form: The preventive or therapeutic agent according to claim 1, which is a protein having an activity of
(A) amino acid sequence described in SEQ ID NO: 1 (b) amino acid sequence described in SEQ ID NO: 1, wherein one or more amino acids are deleted, substituted or added (c) described in SEQ ID NO: 1 Amino acid sequence whose sequence identity is 80% or more Contains latent TGF-β3, which contains any one of the following amino acid sequences (d) to (f) and transmits a TGF-β signal when converted to an active form: The preventive or therapeutic agent according to claim 1, which is a protein having an activity of
(D) the amino acid sequence represented by SEQ ID NO: 3 (e) the amino acid sequence represented by SEQ ID NO: 3 in which one or more amino acids are deleted, substituted, or added (f) SEQ ID NO: 3 An amino acid sequence having a sequence identity of 80% or more with the amino acid sequence represented by   The prevention or treatment according to any one of claims 1 to 3, wherein the bone metabolic disease is osteoporosis, metastatic bone tumor, multiple myeloma, hypercalcemia due to malignant tumor or osteoarthritis. Agent.   A composition for preventing or treating a bone metabolic disease accompanied by a decrease in bone mass, comprising the preventive or therapeutic agent according to any one of claims 1 to 4 and a pharmaceutically acceptable carrier.