MicroRNA 19A / 19B for use in treating pathological conditions associated with bone loss or reduced muscle function

Abstract

Provided is an agent for treating or preventing a disease or pathological condition associated with bone loss or decreased muscle function, or an agent for treating or preventing cancer-related bone destruction. [Solution] A therapeutic or prophylactic agent comprising an inhibitor of microRNA19a and / or microRNA19b, wherein the inhibitor is a nucleic acid or a nucleic acid analog thereof selected from any one of the following (1) to (4): (1) a DNA or RNA oligonucleotide containing or consisting of a sequence capable of hybridizing with microRNA19a and / or microRNA19b to form a duplex that prevents the microRNA from binding to its target mRNA, (2) a microRNA sponge, (3) a lentivector-based anti-microRNA (miRZip), or (4) a tough decoy inhibitor.

Description

[Technical field]

[0001] The present invention relates to inhibitors of microRNA 19a and 19b, and the use of the inhibitors to treat or prevent conditions or diseases associated with bone loss, particularly osteoporosis and osteogenesis imperfecta (OI). The inhibitors are also useful for inducing bone anabolism when administered alone or in combination with parathyroid hormone or a recombinant fragment thereof. The present invention further relates to inhibitors of microRNA 19a and 19b, and the use of the inhibitors to treat or prevent conditions or diseases associated with muscle function loss, particularly muscle degeneration and muscle atrophy. The inhibitors are also useful for stabilizing and / or enhancing muscle function. Furthermore, inhibitors of microRNA 19a and 19b can be used to treat or prevent cancer-related bone destruction or bone metastasis. [Background technology]

[0002] Osteoporosis is the most common disease of the musculoskeletal system in women and men, frequently resulting in fragility fractures. These fractures often represent a major medical and socio-economic burden, as they increase morbidity and mortality, especially in the elderly. Bone is a highly dynamic tissue that is constantly degraded and remodeled throughout life by bone-resorbing osteoclasts and bone-forming osteoblasts, but these processes are tightly coupled and balanced to keep bone mass constant. With age, bone resorption increases while bone formation decreases, leading to reduced bone mass and ultimately osteoporosis.

[0003] Established anti-resorptive drugs such as bisphosphonates and RANKL neutralizing antibodies (Denosumab™) offer a therapeutic approach for osteoporosis by inhibiting osteoclast activity and thereby preventing further loss of bone mass. In contrast, administration of a recombinant fragment of parathyroid hormone (referred to in the literature as PTH 1-34 or teriparatide (Forteo™ / Forsteo™)) is the only bone anabolic therapy currently available. However, PTH therapy requires daily injections of the PTH 1-34 fragment, and safety concerns limit therapy to a full duration of 24 months. Thus, additional therapies that prevent bone loss and / or result in increased bone mass and bone density are needed.

[0004] MicroRNAs (miRNAs) are non-coding RNAs that can block the expression of protein-coding genes. miRNAs have been found to be encoded in the genomes of many organisms, including mammals and non-mammalian animals. miRNAs can bind to the 3' untranslated region (3'-UTR) of target gene mRNAs through their base complementarity. Depending on the binding sequence and the complementarity of the protein involved, miRNAs either inhibit mRNA translation or induce mRNA degradation. Partial base complementarity between miRNA and cognate mRNA usually results in inhibition of mRNA translation, whereas complete base complementarity induces mRNA degradation.

[0005] miRNAs are usually 18–24 nucleotides long and are generated by stepwise processing of precursor molecules. miRNAs are transcribed by polymerase II or III from either the encoding gene or from an intron. The primary transcript resulting from transcription of a gene is called primary microRNA (pri-miRNA). It has a length of 500–3000 nucleotides and carries a 7-methylguanosine cap at the 5' end and a poly-A tail at the 3' end. The pri-miRNA is processed in the cell nucleus by the RNAse III enzyme Drosha and the dsRNA-binding protein DGCR8, leading to precursor microRNAs (pre-miRNAs) of 70–80 nucleotides in length. The pre-miRNAs form characteristic hairpin structures and are transported through nuclear pores to the cytoplasm, where they are processed by the RNAse III enzyme Dicer into ds-miRNAs of 17–24 nucleotides in length. Dicer interacts with proteins that specifically bind to ds-miRNAs to unwind these duplexes. The resulting ss-miRNAs can interfere with the expression of their respective target mRNAs.

[0006] Several miRNAs have been reported to be involved in the regulation of numerous physiological and developmental processes. Recently, miRNAs have attracted great attention as potential therapeutic targets for treating diseases such as hepatitis C [1], chronic lymphocytic leukemia [2], colorectal cancer [3], obesity [4], or schizophrenia [5].

[0007] The present invention is based on the insight that miRNA-19a and miRNA-19b are involved in the regulation of bone formation, and that inhibition of these miRNAs is a highly efficient way to prevent or reduce bone loss. Thus, inhibition of these miRNAs is useful for the treatment of diseases such as osteoporosis that are associated with bone loss. Inhibitors of miRNA-19a and miRNA-19b exert anabolic effects and can be used alone or in combination with each other. In a preferred embodiment, one or more inhibitors of miRNA-19a and / or miRNA-19b are combined with parathyroid hormone, or a fragment thereof, such as PTH1-34, to enhance the bone anabolic effect. The present invention further shows that miRNA-19a and miRNA-19b are involved in the regulation of muscle regeneration, in particular in preventing the loss of muscle mass, strength and / or performance. As shown in the following examples, inhibition of these miRNAs is effective in preventing the loss of muscle function under various conditions. Therefore, inhibition of these miRNAs is also useful for treating diseases such as muscular dystrophy, muscle atrophy, sarcopenia, or cachexia, such as cancer-induced cachexia. The present invention further shows that miRNA-19a and miRNA-19b inhibitors can be used to prevent or treat cancer-induced bone destruction and bone metastasis, particularly breast cancer metastasis to bone. Summary of the Invention

[0008] In one embodiment, the present invention provides an inhibitor of microRNA19a and / or microRNA19b for use in a method for treating a disease associated with bone loss. A disease associated with a pathological degree of bone loss may be, for example, a disease characterized by abnormal bone resorption and / or bone formation in the presence of high bone metabolism. Hereinafter, it is shown that inhibition of microRNA19a and / or microRNA19b results in a strong bone anabolic effect, which demonstrates the usefulness of microRNA19a and / or microRNA19b inhibitors for treating diseases and conditions in which pathological levels of bone loss occur. Such diseases and conditions include, in particular, osteoporosis and osteogenesis imperfecta (OI). Thus, in a preferred embodiment, the disease and condition treated by microRNA19a and / or microRNA19b inhibitors is osteoporosis. As used herein, the term "osteoporosis" includes all forms of osteoporosis, such as primary osteoporosis, secondary osteoporosis, and postmenopausal osteoporosis. Also included are osteoporosis induced by sex steroid deficiency and osteoporosis induced by glucocorticoids. The inhibitors are also useful for treating conditions requiring bone anabolism. For example, the temporary use of the inhibitors in patients suffering from bone fractures can promote the natural healing process by inducing bone mass. In a preferred embodiment, the inhibitors of microRNA19a and / or microRNA19b are used alone, i.e., without any other active ingredient. In another preferred embodiment, the inhibitors of microRNA19a and / or microRNA19b are used in combination with parathyroid hormone or a fragment thereof, such as PTH1-34. The inhibitors of microRNA19a and / or microRNA19b effectively enhance the bone anabolism imparted by PTH1-34.

[0009] In another embodiment, the present invention provides an inhibitor of microRNA19a and / or microRNA19b for use in a method for treating a disease associated with loss of muscle mass or function. The inventors have found that inhibiting the expression of microRNA19a and / or microRNA19b significantly increases or restores muscle function. Thus, inhibitors of microRNA19a and / or microRNA19b are also suitable for therapeutic use in treating diseases such as muscular dystrophy or muscle atrophy. In a preferred embodiment, microRNA19a and / or microRNA19b inhibitors are used to treat muscular dystrophy, sarcopenia, cachexia such as cancer-induced cachexia, or muscle atrophy. In another preferred embodiment, microRNA19a and / or microRNA19b inhibitors are used to treat loss of muscle function associated with osteoporosis, such as sex steroid deficiency-induced or glucocorticoid-induced osteoporosis. In another preferred embodiment, microRNA19a and / or microRNA19b inhibitor is used to treat the loss of muscle function associated with bone metastasis of breast cancer.In yet another preferred embodiment, microRNA19a and / or microRNA19b inhibitor is used to treat the loss of muscle function associated with osteogenesis imperfecta (OI).In yet another preferred embodiment, microRNA19a and / or microRNA19b inhibitor is used to treat the loss of muscle function associated with muscular dystrophy, particularly hereditary muscular dystrophy due to mutation of dystrophin gene, such as Duchenne type or Becker type.

[0010] In another aspect, the present invention provides an inhibitor of microRNA19a and / or microRNA19b for use in a method for preventing or treating cancer-related bone destruction.Herein, it has been found that the inhibitor of microRNA19a and / or microRNA19b effectively inhibits the proliferation of bone-destroying MDA-MB-231 metastatic breast cancer cells.Therefore, the inhibitor of microRNA19a and / or microRNA19b can be used for therapeutic treatment or prevention of cancer-related bone destruction, treatment or prevention of bone metastasis, such as bone metastasis resulting from breast cancer.

[0011] In certain preferred embodiments, the present invention provides inhibitors of microRNA19a and / or microRNA19b for use in methods of treating osteogenesis imperfecta (OI). Osteogenesis imperfecta, also known as brittle bone disease, is a rare genetic disease affecting bones caused by one or more mutations in the gene encoding type I collagen. Several mutations and subtypes causing OI have been identified. People with OI may break bones with minor or no trauma. OI ranges from very severe to mild. Individuals with the most severe form of OI may die at birth. Those with severe OI who survive may have bowed arms and legs, be very short, and be unable to walk. Those with the mildest form of OI may break bones only occasionally, be of normal height, and have a normal life span. Fractures can occur in any bone, but are most common in the arms and legs. The current standard of care for severe forms of OI includes the use of bisphosphonates and surgery to insert rods into the bone to strengthen it. Antiresorptive treatment with bisphosphonates targets the reduction of osteoclast-dependent bone resorption. By reducing resorption, greater bone mass is obtained. Although bone quality remains poor, greater bone mass significantly reduces the risk of fracture. Furthermore, bone-forming agents have been shown to be effective in further increasing bone mass and are currently being investigated in clinical trials. Herein, inhibitors of microRNA19a and / or microRNA19b have been shown to improve bone formation and bone mass under various conditions, making them suitable for the treatment of OI. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0012] MicroRNAs 19a and / or 19b are known to be expressed from the polycistronic cluster miR-17-92, which includes miRNAs 17, 18, 19a, 20a, 19b-1, and 192. Several of these microRNAs have already been associated with human cancers.

[0013] The present invention proposes to use the inhibitor of microRNA 19a and / or 19b to treat diseases related to bone loss or muscle function loss.Although the present invention is not limited in this respect, the subject suffering from disease or condition is preferably a human subject.Preferably, the subject is at least 30 years old, preferably at least 40 years old, at least 45 years old, at least 50 years old, at least 55 years old, at least 60 years old, or at least 65 years old.

[0014] In principle, any type of molecule that interferes with microRNA19a and / or microRNA19b or their pre-miRNAs can be used to reduce or block the expression or function of these microRNAs or their corresponding pre-miRNAs. The inhibitors used herein are preferably nucleic acid molecules, such as ribonucleic acid (RNA) or deoxyribonucleic acid (DNA) molecules, or their derivatives modified as described below. It is particularly preferred that the inhibitors are DNA or RNA oligonucleotides that contain or consist of sequences that can hybridize with the cognate miRNA, i.e., microRNA19a and / or microRNA19b, to form a duplex that prevents the microRNA from binding to the target mRNA. Other techniques for the inhibition of microRNA19a and / or microRNA19b or their pri-miRNAs are known to those skilled in the art and include microRNA sponges, lentivector-based anti-microRNA (miRZip) and tough decoy inhibitors.

[0015] The term "oligonucleotide" as used herein refers to an oligomer or polymer of nucleotide monomers. According to the present invention, an oligonucleotide can be of any length, but preferably has a length of 8-50 nucleotides, more preferably 10-40 nucleotides, 12-30 nucleotides, 15-25 nucleotides, and most preferably 18-24 nucleotides. For example, an oligonucleotide can have a length of 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48 or 50 nucleotides. In some applications, an oligonucleotide may contain more than 50 nucleotides, for example 60, 70, 80, 90 or 100 nucleotides. An oligonucleotide can be a single-stranded, double-stranded, or partially double-stranded nucleic acid molecule. Methods for generating oligonucleotides are widely known and include chemical synthesis or PCR-based methods.

[0016] Oligonucleotides are preferably composed of nucleotide monomers linked by phosphodiester bridges. However, oligomers or polymers in which the sugar phosphate backbone is replaced by a functionally similar structure, such as a peptide backbone, or a phosphoramide, thiophosphate, methylphosphonate or dithiophosphate backbone, can also be used. In some cases, these modifications increase the resistance of the oligonucleotide to enzymatic degradation by nucleases after administration to a therapeutic subject.

[0017] In addition, oligonucleotides with one or more modified sugar residues are also included in the present invention. For example, one or more hydroxyl groups can be replaced with halogen or aliphatic groups, or functionalized with ether, amine, or similar structures. In a preferred embodiment, the oligonucleotide comprises one or more sugars whose 2' hydroxyl has been modified by adding a methyl (OMe) or methoxyethyl (MOE) group. These sugar modifications provide greater stability and resistance to enzymatic degradation.

[0018] The oligonucleotides of the invention may also contain modified bases in addition to or in place of the naturally occurring bases adenine, guanine, thymine, cytosine and uracil. Such modified bases include, for example, 5-methylcytosine, 5-hydroxymethylcytosine, xanthine, hypoxanthine, 2-aminoadenine, 6-methyladenine, 6-methylguanine, 2-thiouracil, 2-thiothymine, 2-thiocytosine, 5-halouracil, 5-halocytosine, 5-propynyl-uracil, 5-propynylcytosine, 6-azouracil, 6-azocytosine, 6-azothymine, 5-uracil, 4-thiouracil, 5-hydroxymethylcytosine ... These include 8-thioalkyladenine, 8-hydroxyladenine, 8-thioladenine, thioalkylguanine, 8-hydroxylguanine, 8-thiolguanine, 7-methylguanine, 7-methyladenine, 8-azaguanine and 8-azaadenine, 7-deazaguanine, 7-deazaadenine, 3-deazaguanine and / or 3-deazaadenine.

[0019] Apart from the above, the oligonucleotides of the invention may also contain one or more modifications at their 3' and / or 5' ends, for example, one or both ends may be bound to a protecting group to render the oligonucleotide less susceptible to enzymatic degradation by nucleases.

[0020] Preferably, the inhibitors of microRNA 19a and / or microRNA 19b are substantially complementary to microRNA 19a and / or microRNA 19b. The sequences of the inhibitors anti-microRNA 19a (5'-TGCATAGATTTGCAC-3') and anti-microRNA 19b (5'-TGCATGGATTTGCAC-3') are set forth in SEQ ID NO: 1 and SEQ ID NO: 2, respectively. The sequences of microRNA 19a and microRNA 19b are set forth in SEQ ID NO: 3 and SEQ ID NO: 4, respectively. An oligonucleotide is substantially complementary to any of these sequences if it comprises at least 70% sequence identity for a length of at least 10, 12, 14, 16, 18 or 20 nucleotides. Preferably, the oligonucleotide has at least 70% sequence identity for a length of 20 nucleotides. Preferably, the sequence identity is at least 80%, at least 90%, or at least 95%. It is particularly preferred that the oligonucleotide is fully complementary for a length of at least 10, 12, 14, 16, 18 or 20 nucleotides. In one embodiment, the regions of complementarity have fewer than 5 mismatches, for example 4, 3, 2, or 1 mismatch.

[0021] In a particularly preferred embodiment, the inhibitor of microRNA 19a and / or microRNA is (a) the sequence of SEQ ID NO: 1 or SEQ ID NO: 2, or (b) the complement of the sequence of SEQ ID NO: 1 or SEQ ID NO: 2, or (c) a variant of any of the sequences of (a) or (b) having at least 70% sequence identity thereto and capable of hybridizing to microRNA 19a and / or microRNA 19b. It is a DNA containing.

[0022] The inhibitor disclosed above is preferably administered as a pharmaceutical composition.Accordingly, in another aspect, the present invention provides a pharmaceutical composition comprising (a) an inhibitor of microRNA19a and / or microRNA19b, such as a nucleic acid or a nucleic acid analog, and (b) a pharmaceutically acceptable carrier.As mentioned above, the inhibitor is preferably a DNA or RNA molecule.In one aspect, the pharmaceutical composition is preferably for use in a method for treating a disease associated with bone loss.In another aspect, the pharmaceutical composition is preferably for use in a method for treating a disease associated with loss of muscle function.

[0023] The preparation of pharmaceutical compositions containing nucleic acid-based inhibitors is well known to those working in the field of pharmacology. Typically, such compositions are prepared for injection as liquid solutions or suspensions. When used in human patients, pharmaceutical active inhibitors can be mixed with excipients that are compatible with the inhibitors. The pharmaceutical compositions of the present invention usually contain a physiologically acceptable carrier with the microRNA inhibitor molecules dissolved or dispersed as active ingredients.

[0024] Pharmaceutically acceptable carriers include, for example, water, physiological saline, Ringer's solution, or dextrose solution. Further suitable carriers for compositions containing microRNA inhibitors are described in standard textbooks, for example, "Remington's Pharmaceutical Sciences", Mack Pub.Co., New Jersey (1991). In addition to carriers, the pharmaceutical compositions of the present invention may contain any concentration of wetting agent, buffer, stabilizer, dye, preservative, etc., provided that they do not interfere with the inhibitory activity of the microRNA inhibitors of the present invention.

[0025] Various routes of administration are feasible to provide the microRNA inhibitor to the site in need of treatment. The pharmaceutical composition may be formulated, for example, for parenteral administration. Parenteral administration includes intravenous, intradermal, intraarterial, subcutaneous, topical, transmucosal, transdermal or rectal administration. According to a particularly preferred embodiment, the pharmaceutical composition is formulated for injection or infusion.

[0026] Pharmaceutical compositions suitable for injection usually include sterile aqueous solutions or dispersions, and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. Moreover, the compositions should be stable under the usual conditions of manufacture and storage. To maintain sterility, pharmaceutical compositions usually contain preservatives, such as parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like, to inhibit the growth of microorganisms in the product. For intravenous or intraarterial administration, suitable carriers may include physiological saline, bacteriostatic water, Cremophor EL™ (BASF) or phosphate buffered saline (PBS). The carrier may also be a solvent or dispersion medium, including, for example, water, ethanol, polyol (such as glycerol, propylene glycol, and liquid polyethylene glycol), and suitable mixtures thereof. Prolonged absorption of the injectable compositions can be achieved by including in the composition an agent that delays absorption, for example, aluminum monostearate or gelatin. Sterile injection solutions can be prepared by mixing the required amount of microRNA inhibitors in a suitable solvent with one or more of the above ingredients, followed by sterile filtration. Similarly, dispersions are prepared by incorporating the microRNA inhibitors into a sterile vehicle that contains a dispersion medium and other ingredients as outlined above as necessary. Sterile solutions can also be obtained by providing the microRNA inhibitors in the form of a sterile powder by methods known in the art, such as vacuum drying or freeze-drying, and reconstituting the powder with a sterile liquid to obtain a final solution for injection. Alternatively, the pharmaceutical composition according to the present invention can also be administered by continuous infusion.

[0027] The administration of the pharmaceutical composition may be achieved by transmucosal or transdermal delivery. For transmucosal or transdermal administration, the pharmaceutical composition containing the microRNA inhibitor of the present invention may contain a penetrant suitable for passing through the skin or mucosal barrier. Such penetrants are known in the art, and include, for example, for transmucosal administration, surfactants, bile salts, and fusidic acid derivatives. Transmucosal administration can be achieved by using nasal sprays or suppositories. For transdermal administration, the active compound is formulated into ointments, salves, gels, or creams as is generally known in the art. Transdermal compositions can be administered, for example, by patch or microneedle. Preferably, the compound is prepared in the form of a suppository, using a conventional suppository base, such as cocoa butter and other glycerides for rectal delivery.

[0028] In one embodiment, the microRNA inhibitor is prepared with a carrier that protects the inhibitor from elimination from the body, such as a controlled release formulation, including implants and microencapsulated delivery systems. Biodegradable biocompatible polymers can be used, such as ethylene vinyl acetate, polyanhydrides, polyglycolic acid, collagen, polyorthoesters, and polylactic acid. Methods for preparing controlled release formulations are well known in the art. In addition, sustained release compositions may be prepared. Suitable examples of sustained release preparations include semipermeable matrices of solid hydrophobic polymers, which matrices are in the form of shaped articles, such as films or microcapsules. Examples of sustained release matrices include polyesters, hydrogels, polylactides, copolymers of L-glutamic acid and ethyl-L-glutamic acid, non-degradable ethylene-vinyl acetate, degradable lactic acid-glycolic acid copolymers, and the like.

[0029] In a further aspect, the present invention provides a method for treating a disease or condition associated with pathological levels of bone loss, such as osteoporosis or OI, comprising administering a therapeutically effective amount of an inhibitor of microRNA19a and / or microRNA19b, or a pharmaceutical composition as defined above comprising such an inhibitor. In yet another aspect, the present invention provides a method for treating a disease or condition associated with loss of muscle function, such as muscular dystrophy, sarcopenia, cachexia, such as cancer-induced cachexia, or muscle atrophy, comprising administering a therapeutically effective amount of an inhibitor of microRNA19a and / or microRNA19b, or a pharmaceutical composition as defined above comprising such an inhibitor.

[0030] A therapeutically effective amount of a microRNA inhibitor is typically an amount sufficient to effectively block or reduce the expression of microRNA 19a and / or microRNA 19b, or their respective pri- or pre-miRNAs, upon administration. In principle, any type of molecule that interferes with the expression of microRNA 19a and / or microRNA 19b can be used for the purposes of the present invention.

[0031] It is understood by those skilled in the art that the amount of inhibitor to be administered to a patient depends on several factors, such as the age and weight of the patient, and the nature and severity of the condition to be treated.The amount of inhibitor that exerts the desired therapeutic effect can be determined in each patient case by using routine experimentation.Typically, the dosage of microRNA inhibitor per body weight varies from about 0.1 mg per kg of patient body weight to about 50 mg per kg of patient body weight, more preferably from about 0.5 mg per kg of patient body weight to about 20 mg per kg of patient body weight, and even more preferably from 1 mg per kg of patient body weight to about 10 mg per kg of patient body weight.The administration schedule can include one or more daily administrations of inhibitor.

[0032] For example, the treatment may last for at least 2 weeks, at least 3 weeks, at least 4 weeks, at least 5 weeks, at least 6 weeks, at least 7 weeks, at least 8 weeks, at least 9 weeks, at least 10 weeks, at least 11 weeks, or at least 12 weeks. In some indications, the treatment may last for at least 4 months, at least 5 months, at least 6 months, at least 7 months, at least 8 months, at least 9 months, at least 10 months, at least 11 months, at least 12 months, or at least 18 months. The duration of the treatment is determined by the physician, taking into account several factors, such as the nature and severity of the condition being treated, the route of administration, etc. [Brief description of the drawings]

[0033] [Figure 1]Figure 1 shows the in vivo increase in bone mass by inhibition of miR-19a / b. (a): Venn diagram of miRNAs decreased by treating mice with PTH or antibodies against sclerostin, respectively. (b): Expression of miR-19a and miR-19b in bones of mice treated with PTH or Scl-Ab. (c): Expression of miR-19a and miR-19b in calvarial osteoblasts stimulated in vitro with PTH or Wnt3a. (d): Differentiation of calvarial osteoblasts transfected with a scrambled control miRNA inhibitor (scr), inhibitors of miR-19a or miR-19b (anti-miR), or a combination of anti-miR-19a and anti-miR-19b (19a / b), assessed by staining for alkaline phosphatase activity. (e): Von Kossa staining of proximal tibia and fluorescent double labeling of bone formation (insets). (f): Histomorphometric analysis of the proximal tibia. (g): Von Kossa staining of the 4th lumbar vertebrae of the same animals as in (e)-(f). (h): BV / TV of the 4th lumbar vertebrae of the same animals as in (e)-(f). Mean ± SEM. *p<0.05, **p<0.01, ***p<0.001 vs. vehicle, #p<0.05, ###p<0.001 vs. scramble. (i)-(k): Micro-computed tomography (μCT) scans of the distal femur (i,j) and mid-shaft femoral cross-section (k) of the same animals as in (e) after the end of treatment. (l): Quantification of the μCT analysis of the distal femur of the same animals as in (e). Mean ± SEM. *p<0.05 vs. scramble. Abbreviations: BV / TV, bone volume / tissue volume; MS / BS, bone mineralization surface / bone surface; BFR / BS, bone formation rate / bone surface; MAR, bone mineralization rate; OS / BS, osteoid surface / bone surface; Ob.S / BS, osteoblast surface / bone surface; N.Ob / BS, osteoblast number / bone surface; ES / BS, erosion surface / bone surface; Oc.S / BS, osteoclast surface / bone surface; N.Oc / BS, osteoclast number / bone surface; Tb.Th, trabecular width; Tb.Sp, trabecular spacing; Tb.N, trabecular number; SMI, structural modeling index. [Diagram 2]Figure 2 shows that inhibition of miR-19a and miR-19b enhances the bone anabolic effects of iPTH treatment and restores bone mass in a mouse model of osteoporosis. (a): Von Kossa staining and (b): histomorphometric analysis of the proximal tibia of 12-week-old male mice of genotype Dmp1-Cre-;Tgif1fl / fl after treatment with intermittent PTH and / or weekly injections of anti-miR-19a / b for 4 weeks. For abbreviations, see Fig. 1 legend. Scale bar indicates 1 mM. (c): Time course of relative BV / TV in the tibia of female mice in which osteoporosis was induced by ovariectomy (OVX) 21 days before the start of weekly treatment with anti-miR-19a / b. (d)-(f), μCT scans of distal femur (d, e) and midshaft femoral cross-section (f) of the same animals as in c, after the end of treatment. (g)-(i), μCT analysis of trabecular BV / TV of femur (g), tibia (h), and 4th lumbar vertebra (i) 70 days after ovariectomy. (j) Model of miR-19a / b action. Mean ± SEM. *p<0.05, ***p<0.001 vs. scr, vehicle; ##p<0.01 vs. miR-19a / b, vehicle; §§§p<0.001 vs. scr, iPTH. *p<0.05, vs. sham; scr#p<0.05, ##p<0.01 vs. OVX; scr. [Diagram 3]FIG. 3 shows the effect of anti-miR-19a / b on ovariectomy-induced loss of muscle function in mice. (a): Histological analysis of tibialis anterior muscle from C56Bl / 6 wild-type male mice injected with FAM-labeled scrambled or anti-miR-19a / b oligonucleotides. Arrows indicate nuclear centers. (b): Ex vivo contractility of extensor digitorum longus in ovariectomized or sham-operated female C57Bl / 6J mice after administration of scrambled (Scr) or anti-miR-19a / b oligonucleotides. (c): Ex vivo contractility of extensor digitorum longus in ovariectomized or sham-operated female C57Bl / 6J mice after administration of Scr or anti-miR-19a / b oligonucleotides. (d): Ex vivo fatigue of extensor digitorum longus in ovariectomized or sham-operated female C57Bl / 6J mice after administration of Scr or anti-miR-19a / b oligonucleotides. (e): Ex vivo fatigue of the soleus muscle in ovariectomized or sham-operated female C57Bl / 6J mice following administration of Scr or anti-miR-19a / b oligonucleotides. *p<0.05, vs. OVX;scr. [Figure 4] Figure 4 shows the effect of anti-miR-19a / b on the in vitro proliferation of bone metastatic MDA-MB-231 breast cancer cells. (a): Proliferation of MDA-MB-231 cells transfected with Scr or anti-miR-19a / b oligonucleotides. (b): Proliferation of MDA-MB-231 cells transfected with Scr or miR-19a / b oligonucleotides. **p<0.01 vs. scr. [Diagram 5]Figure 5 shows that inhibition of miR-19a and miR-19b reduces breast cancer bone metastasis and restores breast cancer-induced loss of bone mass and muscle function. (a) Quantification of bioluminescence signal in tumor-bearing mice treated for 4 weeks with scrambled (Scr, squares) or anti-miR-19a / b oligonucleotides (triangles). (b) μCT analysis of the proximal tibia in non-tumor-bearing mice (control) and mice with bone metastases after treatment with Scr or anti-miR-19a / b oligonucleotides. (c) Ex vivo contractility of soleus muscle in non-tumor-bearing mice (control) and tumor-bearing mice after treatment with Scr or anti-miR-19a / b oligonucleotides. (d) Ex vivo fatigue of soleus muscle in non-tumor-bearing mice (control) and tumor-bearing mice treated with Scr or anti-miR-19a / b oligonucleotides. Mean ± SEM. *p<0.05 vs. non-tumor-bearing mice. [Figure 6]Figure 6 shows the effect of anti-miR-19a / b on glucocorticoid-induced loss of bone mass and muscle function in mice. (a) Fat mass of mice treated with placebo or glucocorticoid (prednisolone) and delivery of scrambled (Scr) or anti-miR-19a / b oligonucleotides measured by dual energy X-ray absorptiometry (DXA). (b) Body weight measured by DXA. (c)-(e) DXA analysis of bone density of mice treated with placebo or prednisolone and delivery of Scr or anti-miR-19a / b oligonucleotides in the whole body (c), femur (d) and spine (e). (f)-(h) Wet muscle weights of gastrocnemius (f), quadriceps (g) and tibialis anterior (h) of mice treated with placebo or prednisolone and Scr or anti-miR-19a / b oligonucleotides. (i) In vivo muscle strength determined using the vertical forelimb grip strength test after 2 weeks of treatment with placebo or prednisolone and Scr or anti-miR-19a / b oligonucleotides. (j) In vivo muscle function tests (plantarflexion torque) in mice treated with placebo or prednisolone and Scr or anti-miR-19a / b oligonucleotides. Mean ± SEM. *p<0.05 vs. corresponding placebo-treated mice, #P<0.05 vs. prednisolone and scr-treated mice. EXAMPLES

[0034] Example 1 In vivo analysis of miRNA expression To identify miRNAs regulated in response to bone anabolic stimuli, 8-week-old mice were injected with a recombinant fragment of human parathyroid hormone (PTH1-34) or an antibody against sclerostin (Scl-Ab). Specifically, wild-type male mice were injected with vehicle, 100 μg / kg PTH1-34 (Bachem), or 100 μg / kg Scl-Ab. After 4 h, total RNA, including small RNAs, was isolated from mouse bones using Trizol (Invitrogen) according to the instructions provided by the manufacturer. For expression analysis, cDNA was synthesized from 1 μg of total RNA using the ProtoScript First Strand cDNA Synthesis Kit (NEBioLabs). Quantitative real-time PCR (qRT-PCR) was performed on a CFX96 detection system (BioRad) using SYBR Green Master Mix (BioRad). After normalization of TATA-binding protein (Tbp) mRNA, the relative expression level and fold induction of each target gene were calculated using the comparative CT (ΔΔCT) method. Small RNAs were isolated from various mouse tissues using Trizol. PolyA tails were added to small RNAs for cDNA synthesis using the QuantimiR-kit (SBI System Biosciences) according to the manufacturer's guidelines. Relative miRNA expression was measured by SYBR Green detection (BioRad) using a universal reverse primer and a specific forward primer designed for each miRNA of interest. U6 expression level was used as an internal control, and relative miRNA expression levels were calculated using the ΔΔCT method.

[0035] Results: Treatment of mice with PTH or Scl-Ab reduced the expression of 35 or 129 miRNAs in bone, respectively. 22 miRNAs were found to be downregulated by both treatments (Fig. 1a). Among the commonly downregulated miRNAs, miR-19a and miR-19b were identified. Direct quantification of miR-19a / b expression in bone confirmed their negative regulation by PTH and Scl-Ab (Fig. 1b).

[0036] Example 2 In vitro analysis of miRNA expression To verify the effect of stimulation of calvarial osteoblasts with PTH1-34 and Wnt3a, an in vitro assay was performed. Calvarial osteoblasts were isolated from 2-3 day old mice and grown in α-MEM containing 10% FBS and P / S. Osteoblasts were stimulated with vehicle, PTH1-34 (100 μM) or Wnt3a (100 ng / ml) for 4 h, and small RNA was extracted from the cells using the miRNEasy kit (Qiagen). After cDNA synthesis, qRT-PCR was used to analyze the expression levels of miR-19a and miR-19b. U6 expression was used as an internal control, and the relative miRNA expression levels were calculated using the ΔΔCT method.

[0037] result : Both PTH1-34 and Wnt3a stimulation induced a decrease in miR-19a / b expression in calvarial osteoblasts (Figure 1c).

[0038] Example 3 : In vitro analysis of osteoblast differentiation To verify whether calvarial osteoblast differentiation could be affected by interference with miR-19a / b expression, cells were transfected with oligonucleotides designed to bind and inhibit endogenous miR-19a (anti-miR-19a: 6-FAM / TGCATAGATTTGCAC) and miR-19b (anti-miR-19b: 6-FAM / TGCATGGATTTGCAC). These synthetic oligonucleotides contain phosphorothioate backbone linkages for optimized pharmacokinetic and pharmacodynamic properties and minimal toxicity, in addition to optimal use in functional studies. Furthermore, the 5' fluorescein FAM label allows for monitoring of transfection efficiency in vitro and tissue delivery in vivo. Cells were transfected with anti-miR-19a / b (50 nM) using the Neon electroporation system (Invitrogen). Osteoblast differentiation was induced by supplementing α-MEM with 0.2 mM L-ascorbic acid and 10 mM β-glycerophosphate (both from Millipore). Osteoblast differentiation was determined by alkaline phosphatase (ALP) staining after fixing the cells with 4% neutral buffered formaldehyde solution. For ALP staining, cells were incubated with Naphthol AS-MX / Fast Blue (both from Sigma-Aldrich) in Tris-HCl solution for 15 min at room temperature.

[0039] result It was found that differentiation of calvarial osteoblasts was synergistically enhanced by transfection with inhibitors against miR-19a and miR-19b (FIG. 1d).

[0040] Example 4 : Bone analysis of mice treated with anti-miR-19a / b To verify whether treatment with a combination of inhibitors against miR-19a and miR-19b (anti-miR-19a / b) could exert a bone anabolic effect in vivo, 8-week-old mice were treated for 28 days with weekly intravenous (i.v.) injections of anti-miR-19a / b followed by measurement of bone mass. Mice were injected with calcein (40 mg / kg) and democycline (20 mg / kg, Sigma-Aldrich) 7 and 2 days before sacrifice, respectively. Tibiae and fourth lumbar (L4) vertebral bodies were collected and fixed in 3.7% PBS-buffered formaldehyde. For histomorphometric analysis, tibiae and L4 were embedded in methyl methacrylate and toluidine blue. Von Kossa staining and tartrate-resistant acid phosphatase (TRAP) staining were performed using 4 μm sagittal sections. Quantitative measurements of bone histomorphometry were performed using the OsteoMeasure system (OsteoMetrics) according to a standardized protocol (ASBMR standard) [6]. Micro-computed tomography (μCT) was used for the three-dimensional analysis of bone properties of long bones and vertebral bodies. The fourth lumbar (L4) vertebral bodies of mice were analyzed using high-resolution micro-computed tomography with a fixed isotropic voxel size of 10 μm (70 peak kV with an integration time of XμA400 ms, Viva80 micro-CT, Scanco Medical AG). This threshold was verified by manually evaluating 10 single tomographic slices from four samples per group, isolating mineralized tissues and preserving their morphology while excluding non-mineralized tissues. All analyses were performed on digitally extracted bone tissues using 3D distance techniques (Scanco Medical AG).

[0041] resultAnti-miR-19a / b treatment increased trabecular bone mass in the proximal tibia. Histomorphometric analysis revealed that anti-miR-19a / b treatment activated bone remodeling by enhancing osteoblast and osteoclast parameters, promoted bone formation, and led to a net increase in bone mass (Figure 1e and Figure 1f). The bone anabolic effect of anti-miR-19a / b treatment was confirmed using micro-computed tomography (μCT) analysis in the L4 vertebral body (Figure 1g and Figure 1h), and in the distal femur (Figure 1i-l).

[0042] Example 5 Simultaneous treatment with miR-19a / b inhibitors and intermittent PTH(1-34) Because both PTH and anti-miR-19a / b increase bone mass, co-treatment was evaluated. A recombinant fragment of human parathyroid hormone (PTH1-34; 100 μg / kg body weight, Biochem) or vehicle was administered intraperitoneally to 8-week-old male mice five times per week for three weeks. Anti-miR-19a / b or scr control were administered once per week as co-treatment.

[0043] result: Anti-miR-19a / b enhanced PTH-mediated bone mass increase in the tibia of male mice (FIGS. 2a and 2b).

[0044] Example 6 : miR-19a / b inhibitors in osteoporosis models Because bone loss is a hallmark of osteoporosis, we tested whether increasing bone mass by anti-miR-19a / b administration could also be useful for treating osteoporosis in females. To this end, 8-week-old female mice were ovariectomized to deprive them of sex steroids, leading to a marked decrease in trabecular bone mass in the tibia. To mimic the clinical situation, anti-miR-19a / b treatment with weekly injections of anti-miR-19a / b or scrambled control was initiated after osteoporosis was fully established 21 days after ovariectomy.

[0045] resultThis treatment attenuated bone loss in this osteoporosis model, preventing further bone loss 14 days after the first treatment. Quantification of trabecular bone mass in the distal femur, proximal tibia, and L4 vertebrae after the end of treatment confirmed that anti-miR-19a / b treatment had a protective effect against osteoporotic bone loss.

[0046] Example 7 Muscle histological analysis The effect of anti-miR-19a / b on ovariectomy-induced loss of muscle function was analyzed. FAM-labeled scrambled or anti-miR-19a / b oligonucleotides were injected into 8-week-old C56Bl / 6 wild-type male mice once a week (10 mg / kg, iv) for 4 weeks and histological analysis was performed. One week after the last treatment, mice were sacrificed and muscles (tibialis anterior) were harvested. Muscles were embedded in optimal cutting temperature compound (OCT) before frozen sectioning in a cryostat. Histological sections were stained with periodic acid-Schiff (PAS) and hematoxylin solutions.

[0047] result At steady state, nuclei are located at the edges of myofibers. Interestingly, in myofibers treated with anti-miR-19a / b, a shift of nuclei from a peripheral to a central location was observed (Figure 3A), suggesting active and spontaneous muscle tissue remodeling in response to miR-19a / b inhibition.

[0048] Example 8 : Muscle function analysis To investigate the functional implications of the findings reached in Example 7, ex vivo functional studies were performed using freshly harvested muscles (extensor digitorum longus and soleus). Eight-week-old female C57Bl / 6J mice were ovariectomized or sham-operated and three weeks later were treated with scrambled (Scr) or anti-miR-19a / b (10 mg / kg, iv) once a week for seven weeks. One week after the last injection, the mice were sacrificed and the ex vivo contractility of the extensor digitorum longus (EDL) and soleus muscles was analyzed using a special device (Aurora Scientific) designed to measure the properties of mouse muscles in situ, ex vivo and in vivo. For this purpose, the EDL and soleus muscles were dissected from the hind limbs, steel hooks were tied to the tendons of the muscles and the muscles were mounted between force transducers. The muscles were stimulated to contract using a supramaximal stimulation between two electrodes. In the fatigue test, the extensor digitorum longus and soleus muscles were stimulated repeatedly 50 times at 70 Hz to detect the maximum tetanic force. Data were collected and analyzed using the Dynamic Muscle Control / Data acquisition (DMC) and Dynamic Muscle Control Data Analysis (DMA) programs (Aurora Scientific).

[0049] result Functional testing revealed that ovariectomy significantly reduced the strength (Figure 3b and 3c) and elasticity (Figure 3d and 3e) of the extensor digitorum longus and soleus muscles. Anti-miR-19a / b (10 mg / kg, iv, once a week for 7 weeks) restored both strength and elasticity to sham control levels (Figure 3b-e), strongly suggesting that antagonizing miR-19a / b prevents ovariectomy-induced loss of muscle function. These findings indicate that miR-19a / b is a novel target for the treatment of muscle dysfunction.

[0050] Example 9 : Proliferation of metastatic breast cancer cells in bone The above findings support the notion that antagonizing miR-19a / b is bone-protective. Therefore, we tested whether anti-miR-19a / b treatment could reduce its activity in osteodestructive MDA-MB-231 breast cancer cells. To test this hypothesis, MDA-MB-231 breast cancer cells were transfected with anti-miR-19a / b, miR-19a / b nucleotides, or the respective scr controls. Cells were grown in vitro for 4 days, and cell proliferation was measured by MTS assay every day until the end of the experiment.

[0051] result Treatment of bone-destructive MDA-MB-231 breast cancer cells with anti-miR-19a / b nucleotides significantly reduced cancer cell proliferation by days 3 and 4, whereas delivery of miR-19a / b nucleotides increased cancer cell proliferation compared to controls at the same time points. These data strongly suggest that inhibiting endogenous miR-19a / b in MDA-MB-231 breast cancer cells may prevent them from exerting their bone-destructive potential in the context of metastatic breast cancer disease.

[0052] Example 10 Treatment of metastatic bone disease We tested whether anti-miR-19a / b treatment could attenuate the activity of osteodestructive MDA-MB-231 breast cancer cells and prevent breast cancer-induced osteolytic disease in vivo. MDA-MB-231 breast cancer cells stably expressing the luciferase gene were injected into the left ventricle of 8-week-old immunodeficient SCID mice. Micrometastases were detected 2 weeks after breast cancer cell injection by bioluminescence imaging (BLI), and mice were randomly divided into two treatment groups. One group received scrambled oligonucleotide (Scr; 10 mg / kg) and the other group received anti-miR-19a / b (10 mg / kg) intravenously (iv) once a week for 4 weeks. Tumor burden was measured weekly by BLI. One week after the last injection, mice were sacrificed and ex vivo contractility of the soleus muscle was analyzed using a special device (Aurora Scientific) designed to measure mouse muscle properties in situ, ex vivo, and in vivo. For this purpose, the soleus muscle was dissected from the hind limb, a hook was tied to the muscle tendon, and the muscle was mounted between a force transducer. The muscle was stimulated to contract using supramaximal stimulation between two electrodes. For fatigue testing, the soleus muscle was stimulated 50 times repeatedly at 70 Hz to detect the maximum tetanic force. Data were collected and analyzed using the Dynamic Muscle Control / Data acquisition (DMC) and Dynamic Muscle Control Data Analysis (DMA) programs (Aurora Scientific). Micro-computed tomography (μCT) was used for the three-dimensional analysis of the long bones. Mouse long bones were analyzed using high-resolution micro-computed tomography with a fixed isotropic voxel size of 10 μm (70 peak kV with an integration time of XμA 400 ms; Viva80 micro-CT; Scanco Medical AG). This threshold was verified by manually evaluating 10 single tomographic slices from four samples per group, isolating the mineralized tissue and preserving its morphology while excluding the non-mineralized tissue. All analyses were performed on digitally extracted bone tissue using 3D distance technology (Scanco Medical AG).

[0053] result BLI imaging revealed that tumor growth in bones was significantly reduced in mice treated with anti-miR-19a / b compared to Scr-treated controls (Figure 5a). Breast cancer induced significant bone loss in Scr-treated mice, which was prevented by weekly anti-miR-19a / b treatment (Figure 5b). Muscle function tests revealed that anti-miR-19a / b (10 mg / kg, iv, weekly for 4 weeks) restored the cancer-induced loss of both strength and endurance in the soleus muscle (Figure 5c and d). This strongly suggests that antagonizing miR-19a / b reduces bone metastasis burden and prevents breast cancer-induced loss of bone mass and muscle function.

[0054] Example 11 Treatment of glucocorticoid-induced osteoporosis To investigate whether anti-miR-19a / b is involved in glucocorticoid-induced osteoporosis, placebo or prednisolone (2.1 mg / kg / d) pellets were subcutaneously implanted into 4-month-old female C57Bl / 6J mice. Mice were treated with scrambled (Scr) or anti-miR-19a / b (10 mg / kg, iv) 1 day before pellet implantation and then once a week. Bone mineral density (BMD) and fat mass were measured before and after 4 weeks of treatment using dual energy X-ray absorptiometry (DEXA, Piximus). In vivo muscle strength was measured after 2 weeks of treatment using a vertical forelimb grip strength test. In vivo muscle function tests (plantar flexion torque) were performed using a special device (Aurora Scientific) designed to measure mouse muscle properties in vivo. Maximal muscle contraction force was normalized to body weight. After 4 weeks of treatment, the mice were sacrificed and the wet weights of the gastrocnemius, quadriceps and tibialis anterior (TA) muscles were measured.

[0055] resultDual energy X-ray absorptiometry revealed an increase in fat mass upon glucocorticoid (prednisolone) treatment, which was prevented by weekly delivery of anti-miR-19a / b (Figure 6a). No change in total body weight was observed (Figure 6b). Furthermore, BMD was significantly reduced in the femur and spine of prednisolone-treated mice (Figure 6c-e). Mice treated with anti-miR-19a / b were prevented from glucocorticoid-induced loss of BMD (Figure 6c-e). Although glucocorticoids reduced muscle mass in both Scr and anti-miR-19a / b treated mice, gastrocnemius, quadriceps and tibialis anterior muscle mass were significantly higher in anti-miR-19a / b treated mice compared to Scr-treated controls (Figure 6f-h). Importantly, anti-miR-19a / b treatment prevented glucocorticoid-induced loss of muscle function as determined by grip strength and plantar flexion torque tests (FIG. 6i, FIG. 6j).

[0056] literature 1. Janssen, HLA et al., Treatment of HCV Infection by Targeting MicroRN A, N. Engl. J. Med. 368, 1685-1694 (2013). 2. Musilova K. & Mraz M., MicroRNAs in B cell lymphomas: How a complex biology gets more complex", Leukemia, 2015 29 (5), 1004-17 (2015). 3. Nielsen BS, et al., High levels of microRNA-21 in the stroma of colorec tal cancers predict short disease-free survival in stage II colon cancer patients, Clin Exp Metastasis 28 (1): 27-38 (2010). 4. Romao JM et al., MicroRNA regulation in mammalian adipogenesis, Exp. Biol. Med. 236 (9), 997-1004 (2011). 5. Hommers LG et al., Heterogeneity and Individuality: microRNAs in Mental Disorders, J Neural Transm. 122 (1): 79-97 (2015). 6. Parfitt, AM et al., Bone histomorphometry: standardization of nomenclat ure, symbols, and units. Report of the ASBMR Histomorphometry Nomenclature C ommittee. J. Bone Miner. Res. 2, 595-610 (1987).

[0057] array SEQ ID NO:1 - 5'-TGCATAGATTTGCAC-3' (anti-miR-19a) SEQ ID NO:2 - 5'-TGCATGGATTTGCAC-3' (anti-miR-19b) SEQ ID NO: 3 - 5'-UGUGCAAAUCUAUGCAAAACUGA-3' (miR-19a) SEQ ID NO: 4 - 5'-UGUGCAAAUCCAUGCAAAACUGA-3' (miR-19b)

Claims

1. A therapeutic or preventive agent for a disease or pathological condition associated with bone loss or muscle function decline, or a therapeutic or preventive agent for cancer-related bone destruction, comprising an inhibitor of microRNA 19a and / or microRNA 19b; said inhibitor is a DNA or RNA oligonucleotide comprising or consisting of a sequence capable of hybridizing with microRNA 19a and / or microRNA 19b to form a duplex that prevents the binding of the microRNA to the target mRNA; The disease or pathological condition associated with bone loss is osteoporosis or osteogenesis imperfecta (OI); The disease or pathological condition associated with reduced muscle function is muscle atrophy, sarcopenia, or cachexia; The cancer-related bone destruction is caused by bone metastasis.

2. The therapeutic or prophylactic agent according to claim 1 , wherein the therapeutic or prophylactic agent is used in combination with parathyroid hormone or a fragment thereof.

3. The therapeutic or prophylactic agent according to claim 1 or 2, which is administered intravenously, subcutaneously, transdermally, or transmucosally.

4. The therapeutic or prophylactic agent according to claim 3 , which is administered by infusion or injection.

5. The inhibitor is (a) the sequence of SEQ ID NO:1 or SEQ ID NO:2, or (b) the complement of the sequence of SEQ ID NO: 1 or SEQ ID NO: 2, or (c) a variant of any of the sequences of (a) or (b) having at least 90% sequence identity thereto and capable of hybridizing to microRNA 19a and / or microRNA 19b. The therapeutic or prophylactic agent according to any one of claims 1 to 4, which is a DNA comprising the following:

6. A pharmaceutical composition for treating or preventing a disease or pathological condition associated with bone loss, for treating or preventing a disease or pathological condition associated with reduced muscle function, or for treating or preventing cancer-related bone destruction, comprising (a) an inhibitor of microRNA 19a and / or microRNA 19b, and (b) a pharma- ceutically acceptable carrier, said inhibitor is a DNA or RNA oligonucleotide comprising or consisting of a sequence capable of hybridizing with microRNA 19a and / or microRNA 19b to form a duplex that prevents the binding of the microRNA to the target mRNA; The disease or pathological condition associated with bone loss is osteoporosis or osteogenesis imperfecta (OI); The disease or pathological condition associated with reduced muscle function is muscle atrophy, sarcopenia, or cachexia; The pharmaceutical composition, wherein the cancer-related bone destruction is caused by bone metastasis.

7. The pharmaceutical composition of claim 6 , wherein the inhibitor is DNA or RNA.

8. 8. The pharmaceutical composition of claim 6 or 7, which is administered intravenously, subcutaneously, transdermally, or transmucosally.

9. 9. The pharmaceutical composition of claim 8, which is administered by infusion or injection.

10. The inhibitor is (a) the sequence of SEQ ID NO:1 or SEQ ID NO:2, or (b) the complement of the sequence of SEQ ID NO: 1 or SEQ ID NO: 2, or (c) a variant of any of the sequences of (a) or (b) having at least 90% sequence identity thereto and capable of hybridizing to microRNA 19a and / or microRNA 19b. The pharmaceutical composition according to any one of claims 6 to 9, which is a DNA comprising:

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