Use of FGFR3 tyrosine kinase inhibitors for the treatment of FGFR-associated bone repair and bone formation disorders - Patent Application 20070233333

Abstract

The present invention relates to a method for treating FGFR-related bone repair and bone formation and bone quality disorders.The inventors provide data that demonstrates the impaired bone formation and repair process in HCH mandibles, characterized by the presence of pseudoarthrosis in many calluses and bone structure similar to osteoporotic bone, due to the abnormal activation of FGFR3 signal transduction.Interestingly, treatment with (BGJ398) partially restores the defective bone formation and repair.Therefore, the present invention relates to a method for treating FGFR-related bone repair and bone formation disorders in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of at least one FGFR3 tyrosine kinase inhibitor.

Description

[Technical field]

[0001] Field of the invention: The present invention relates to methods and pharmaceutical compositions for the treatment of FGFR-associated bone repair and bone formation and quality disorders. [Background technology]

[0002] Background of the invention: Fibroblast growth factor receptors (FGFRs) are key genes involved in bone formation. Patients with craniosynostosis and chondrodysplasias associated with gain-of-function mutations in FGFRs exhibit craniofacial and mandibular malformations (Kolar JC 2017). FGFR2-related craniostenosis (e.g., Crouzon, Apert and Pfeiffer syndromes) is characterized by the presence of unicoronal or bicoronal craniosynostosis with fusion of one or more cranial sutures, resulting in various cranial deformities with hypertelorism, exophthalmos, midfacial hypoplasia and mandibular prognathism. FGFR3-related craniosynostosis (e.g., Muenke and Crouzon syndromes and acanthosis nigricans) presents with craniofacial congenital anomalies, i.e., unicoronal or bicoronal craniosynostosis, and midfacial hypoplasia. FGFR3-related chondrodysplasias, such as achondroplasia, severe achondroplasia with developmental delay and acanthosis nigricans (SADDAN) and hypochondroplasia nigricans (HCH), are characterized by short limbs, skull base birth defects, macrocephaly, deafness, midface hypoplasia and mandibular prognathism. Maxillofacial and neurosurgery is indicated for these craniosynostosis and chondrodysplasia patients to correct the craniofacial and skull birth defects. The process of bone healing occurs in two different ways depending on the mechanical stability. Stable fracture healing occurs via membranous ossification. In contrast, healing of unstable fractures is under the control of endochondral ossification. In this case, a large cartilage template forms in the fracture gap, which is replaced by bone to bridge both ends of the broken bone.

[0003] Fracture healing is a complex process involving a series of cellular events including initial bleeding and inflammation, recruitment and proliferation of mesenchymal cells, followed by the formation of a cartilaginous callus and its gradual replacement by a bony callus. Diverse growth factors / cytokines regulate skeletal development and homeostasis and can also regulate fracture healing. It is well known that receptor tyrosine kinases (RTKs) play a role in bone repair. Among the RTKs, FGFR3 regulates the formation of the cartilaginous callus and its replacement by bone. A mouse model of chondrodysplasia (Fgfr3 Y367C / + ), gain-of-function mutations in FGFR3 impair bone regeneration in unstabilized tibial fractures and induce a pseudarthrosis phenotype in the callus (Julien et al., 2020).

[0004] Understanding skeletal repair and bone formation is essential for the development of therapies used to improve bone healing after surgical osteotomies or traumatic fractures. The exact function of FGFRs in mandibular formation and repair remains to be understood. In the maxillofacial skeleton, the mandible is the largest and most robust bone of the face, and its abnormal growth and development contributes to imbalance of the upper and lower jaws and subsequent congenital anomalies of dental occlusion and facial developmental variations. Two processes of ossification control mandibular development: 1) endochondral ossification regulates the formation of the condylar process and Meckel's cartilage, and 2) membranous ossification controls the formation and elongation of the mandibular ramus.

[0005] To investigate abnormal FGF signaling during mandibular bone formation and repair, we used a mouse model of HCH (Fgfr3 N534K / +) was studied. The HCH mouse model presents the maxillofacial features of HCH, i.e. macrocephaly and mandibular prognathism. Unstabilized fractures were performed on the mandibles of adult HCH mice, and the formation of the callus repair was analyzed at different critical time points of the repair. Our data demonstrate impaired bone formation and resorption, bone quality, and bone repair process in HCH mandibles, characterized by the presence of pseudoarthrosis in many calluses, due to abnormal activation of FGFR3 signaling. Interestingly, treatment with an FGFR3 tyrosine kinase inhibitor (BGJ398) partially restores the incomplete bone repair of the callus and increases bone volume / total volume without pseudoarthrosis.

[0006] Summary of the invention: The present invention relates to methods and pharmaceutical compositions for the treatment of FGFR-associated bone repair and bone formation and quality disorders. In particular, the present invention is defined by the claims.

[0007] Detailed description of the invention: The present invention relates to a method for treating an FGFR-associated bone repair and bone formation disorder in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of at least one FGFR3 tyrosine kinase inhibitor.

[0008] The present invention also relates to a method for restoring defective bone repair in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of at least one FGFR3 tyrosine kinase inhibitor.

[0009] As used herein, the term "subject" or "patient" refers to a mammal, such as a rodent, cat, dog, and primate. In particular, the subject of the present invention is a human. In particular, the subject of the present invention is an adult. In particular, the subject of the present invention is a child, a teenager, or an elderly person. In some embodiments, the patient is under 15 years old. In some embodiments, the patient is under 10 years old. In some embodiments, the patient is under 7 years old. In some embodiments, the patient is under 5 years old. In some embodiments, the patient is under 3 years old. In some embodiments, the patient is an adult. In some embodiments, the subject is over 15 years old. In some embodiments, the subject is over 20 years old. In some embodiments, the subject is over 25 years old. In some embodiments, the subject is over 30 years old. In some embodiments, the subject is over 35 years old.

[0010] As used herein, the term "bone" refers to hard tissue that constitutes part of the skeleton of most vertebrates. Bones protect various organs of the body, produce red and white blood cells, store minerals, structure and support the body, and provide mobility. Bones vary in shape and size and have complex internal and external structures. They are light, yet strong and rigid, and perform multiple functions. Bone tissue (bone tissue) is a hard tissue and a type of specialized connective tissue. It has a honeycomb-like matrix inside, which helps give bone rigidity. Bone tissue is composed of different types of bone cells. Osteoblasts and osteocytes are involved in the formation and mineralization of bone; osteoclasts are involved in the resorption of bone tissue. Modified (flattened) osteoblasts become surface cells that form a protective layer on the bone surface. The mineralized matrix of bone tissue has an organic component, mainly consisting of collagen called ossein, and an inorganic component of bone mineral, which is composed of various salts. Bone tissue is a mineralized tissue of two types: cortical bone and cancellous bone (also called trabecular bone). Other types of tissue found in bone include bone marrow, endosteum, periosteum, nerves, blood vessels, growth plates, and articular cartilage.

[0011] As used herein, the terms "osteogenesis", "osteogenesis" or "ossification" refer to the process of bone formation. After precursor cells form osteoblast layers, they proceed to three developmental stages of cell differentiation, called proliferation, matrix maturation and mineralization. Based on their embryological origin, there are two types of ossification, called intramembranous ossification, which occurs in mesenchymal cells that differentiate directly into osteoblasts in ossification centers without preceding cartilage formation, and endochondral ossification, in which bone tissue mineralization is formed first through chondrogenesis. In intramembranous ossification, bone development occurs directly. In this process, mesenchymal cells proliferate into highly vascularized areas of embryonic connective tissue in the formation of cell aggregates or primary ossification centers. The cells synthesize bone matrix in the periphery, and mesenchymal cells continue to differentiate into osteoblasts. The bone is then remodeled and replaced by mature lamellar bone. Endochondral ossification forms the primary ossification center, where cartilage is extended by chondrocyte proliferation and deposition of cartilage matrix. After this formation, chondrocytes in the central region of the cartilage begin to proceed to maturation into hypertrophic chondrocytes. After the primary ossification center is formed, the medullary cavity begins to widen toward the epiphysis. Subsequent stages of endochondral ossification then occur in several areas of the bone.

[0012] In one embodiment, the treatment of the present invention improves bone healing following surgical osteotomy or traumatic fracture.

[0013] As used herein, the term "surgical osteotomy" refers to a procedure in which a surgeon removes or sometimes adds a wedge of bone near an injured joint, which transfers weight from an area with damaged cartilage to an area with more or healthier cartilage.

[0014] As used herein, the term "bone healing" or "fracture healing" or "bone repair" has its common meaning in the art and refers to the proliferative physiological process by which the body promotes the repair of a fracture. The bone healing process has three overlapping stages. 1. Inflammation begins immediately after a fracture occurs and continues for several days. Once a fracture occurs, blood rushes to the area, causing inflammation and blood clotting at the fracture site. This provides the initial structural stability and framework for new bone formation. 2. Bone production begins when the blood clot formed by inflammation is replaced with fibrous tissue and cartilage (known as the soft callus). As healing progresses, the soft callus is replaced with hard bone (known as the hard callus), which is visible on an x-ray several weeks after the fracture. 3. Bone remodeling, the final stage of bone healing, continues for several months. During remodeling, bone continues to form, compress, and return to its original shape. In addition, blood circulation to the area improves. Once adequate bone healing has occurred, weight bearing (e.g., standing or walking) aids in bone remodeling.

[0015] In some embodiments, the subject suffers from a bone fracture.As used herein, the term "bone fracture" refers to a medical condition in which there is partial or complete destruction of the continuity of bone.In more severe cases, bone may be destroyed into several pieces.

[0016] In one embodiment, the FGFR3 tyrosine kinase inhibitor of the present invention increases bone mineral density or bone volume / total volume (BV / TV).As used herein, the term "bone mineral density" or "bone volume / total volume" (BV / TV) refers to the amount of bone mineral in bone tissue.The concept is the mass of mineral per bone volume.

[0017] In one embodiment, the FGFR3 tyrosine kinase inhibitor of the present invention can be used to treat osteoporosis.As used herein, the term "osteoporosis" refers to a bone disease characterized by the loss of bone tissue / bone density, and over time bones become very thin and fragile.

[0018] As used herein, the term "craniofacial congenital anomalies" refers to a diverse group of deformities in the growth of the bones of the head and face. Congenital anomalies refers to the medical term meaning "irregular" or "different from normal." These anomalies are present at birth (congenital) and include a number of variants. Some are mild, while others are severe and require surgery.

[0019] As used herein, the term "craniosynostosis" refers to a condition in which one or more of the fibrous sutures of a young infant's skull fuses prematurely by turning into bone (ossification), thereby altering the growth pattern of the skull. This may involve a growth abnormality of the facial skeleton known as facio-cranio-stenosis, or may even be within the scope of a multiple malformation syndrome.

[0020] As used herein, the term "mandible", also known as the lower jaw or mandible bone, is the largest and strongest lowermost bone in the human facial skeleton. It forms the lower jaw and holds the lower teeth in place. The mandible is located below the maxilla. It is the only mobile bone in the skull (excluding the ossicles of the middle ear). It is connected to the temporal bone by the temporomandibular joint.

[0021] In some embodiments, the subject will have or be suffering from an FGFR-associated bone repair and bone formation disorder. In some embodiments, the subject carries a gain-of-function mutation in an FGFR.

[0022] As used herein, the term "FGFR-related bone repair and bone formation and bone quality disorder" or "FGFR-related bone repair and bone formation disorder" refers to the lack of bone repair and bone formation progression.FGFR-related bone repair disorder refers to defective bone repair.

[0023] As used herein, the term "fibroblast growth factor" (FGF) refers to a family of cell signaling proteins; they are involved in a wide variety of processes, especially as critical elements for normal development in animal cells. Any irregularity in their function leads to a wide range of developmental defects. These growth factors typically act as systemic or local circulating molecules of extracellular origin that activate cell surface receptors. A defining property of FGFs is that they bind to the co-receptors heparin and heparan sulfate. Thus, some are sequestered in the extracellular matrix of tissues containing heparan sulfate proteoglycans and are released locally upon injury or tissue remodeling.

[0024] As used herein, the term "fibroblast growth factor receptor" (FGFR) refers to a receptor that binds to a member of the fibroblast growth factor (FGF) family of proteins. Some of these receptors are involved in pathological conditions. Distinct membrane FGFRs have been identified in vertebrates, all of which belong to the tyrosine kinase superfamily: FGFR1 (also see fibroblast growth factor receptor 1) (=CD331), FGFR2 (also see fibroblast growth factor receptor 2) (=CD332), FGFR3 (also see fibroblast growth factor receptor 3) (=CD333), FGFR4 (also see fibroblast growth factor receptor 4) (=CD334), FGFRL1 (also see fibroblast growth factor receptor-like 1) and FGFR6.

[0025] In certain embodiments, the subject carries a gain-of-function mutation in FGFR3.

[0026] As used herein, the terms "FGFR3", "FGFR3 tyrosine kinase receptor" and "FGFR3 receptor" are used interchangeably throughout the present specification and refer to all naturally occurring isoforms of FGFR3. An exemplary human amino acid sequence of FGFR3 is represented by SEQ ID NO:1. [ka]

[0027] As used herein, the expressions "gain-of-function mutation of FGFR3", "constitutively active FGFR3 receptor variant", "constitutively active mutant of FGFR3" or "mutant FGFR3 exhibiting constitutive activity" are used interchangeably and refer to mutants of said receptors that exhibit biological activity (i.e., induce downstream signaling) and / or exhibit greater biological activity than the corresponding wild-type receptor in the presence of an FGF ligand. Constitutively active FGFR3 variants according to the invention are in particular selected from the group consisting of (residues numbered according to their position in the precursor of fibroblast growth factor receptor 3 isoform 1 - 806 amino acids long -): a variant in which the serine residue at position 84 is replaced by a lysine (hereinafter designated S84L); a variant in which the arginine residue at position 200 is replaced by a cysteine ​​(hereinafter designated R200C); a variant in which the arginine residue at position 248 is replaced by a cysteine ​​(hereinafter designated R248C); a variant in which the serine residue at position 249 is replaced by a cysteine ​​(hereinafter designated S249C); a variant in which the proline residue at position 250 is replaced by an arginine (hereinafter designated P250R); a variant in which the asparagine residue at position 262 is replaced by a histidine (hereinafter designated N262H); a mutant in which the glycine residue at position 268 is replaced by a cysteine ​​(hereinafter designated G268C); a mutant in which the tyrosine residue at position 278 is replaced by a cysteine ​​(hereinafter designated Y278C); a mutant in which the serine residue at position 279 is replaced by a cysteine ​​(hereinafter designated S279C); a mutant in which the glycine residue at position 370 is replaced by a cysteine ​​(hereinafter designated G370C); a mutant in which the serine residue at position 371 is replaced by a cysteine ​​(hereinafter designated S371C); a mutant in which the tyrosine residue at position 373 is replaced by a cysteine ​​(hereinafter designated Y373C); a mutant in which the glycine residue at position 380 is replaced by an arginine (hereinafter designated G380R); a mutant in which the valine residue at position 381 is replaced by a glutamic acid (hereinafter designated V381E);Mutants in which the alanine residue at position 391 is replaced by glutamic acid (hereinafter designated A391E); mutants in which the asparagine residue at position 540 is replaced by lysine (hereinafter designated N540K); mutants in which the stop codon has been eliminated by a base substitution, in particular mutants in which the stop codon has been mutated at an arginine, cysteine, glycine, serine or tryptophan codon (hereinafter designated X807R, X807C, X807G, X807S and X807W, respectively); mutants in which the lysine residue at position 650 is replaced by another residue, in particular methionine, glutamic acid, asparagine, etc. a mutant in which the methionine residue at position 528 is replaced by isoleucine (hereinafter referred to as M528I); a mutant in which the isoleucine residue at position 538 is replaced by valine (hereinafter referred to as I538V); a mutant in which the asparagine residue at position 540 is replaced by serine (hereinafter referred to as N540S); a mutant in which the asparagine residue at position 540 is replaced by threonine (hereinafter referred to as N540T). Typically, the constitutively active FGFR3 variant according to the present invention is an N540K, K650N, K650Q, M528I, I538V, N540S, N540T or A391E variant;

[0028] In certain embodiments, the subject has an FGFR3-associated skeletal disease.

[0029] As used herein, the term "FGFR3-associated skeletal disease" is intended to mean a skeletal disease caused by abnormally increased activation of FGFR3, in particular by expression of a constitutively active mutant of the FGFR3 receptor, in particular a constitutively active mutant of the FGFR3 receptor as described above.

[0030] In certain embodiments, the FGFR3-related skeletal disease is preferably FGFR3-related chondrodysplasia and FGFR3-related craniosynostosis.

[0031] As used herein, "FGFR3-associated chondrodysplasia" includes, but is not limited to, dwarfism, such as hypochondroplasia (HCH), thanatophoric dysplasia (TD) type I, thanatophoric dysplasia type II, achondroplasia (ACH) and SADDAN (severe achondroplasia with developmental delay and acanthosis nigricans).

[0032] In particular, the FGFR3-associated skeletal disorder is dwarfism.

[0033] As used herein, the term "dwarfism" has its ordinary meaning in the art and refers to short stature resulting from genetic or medical conditions. Dwarfism is generally defined as an adult height of 147 centimeters or less.

[0034] In particular, an FGFR3-associated skeletal disease is hypochondroplasia (HCH).

[0035] As used herein, the term "hypoplasia achondroplasia" (HCH) has its common meaning in the art and relates to disproportionately short stature, shortened limbs and a head that appears large in comparison to underdeveloped parts of the body.

[0036] In certain embodiments, the FGFR3-associated chondrodysplasia is hypochondroplasia caused by expression of the constitutively active N540K, K650N, K650Q, M528I, I538V, N540S or N540T mutants of the FGFR3 receptor.

[0037] In particular, an FGFR3-related skeletal disease is achondroplasia (ACH).

[0038] As used herein, the term "achondroplasia" (ACH) has its common meaning in the art and relates to a genetic defect in which the arms and legs are short while the trunk is usually of normal length, with an enlarged head and prominent forehead.

[0039] In particular, the FGFR3-related skeletal disease is thanatophoric dysplasia (TD).

[0040] As used herein, the term "thanatophoric dysplasia" (TD) has its common meaning in the art and relates to severe skeletal defects characterized by a disproportionately small rib cage, extremely short limbs and excess skin folds on the arms and legs.

[0041] In certain embodiments, the FGFR3-associated skeletal disease is FGFR3-associated craniosynostosis. In some embodiments, the FGFR3-associated craniosynostosis represents an inherited or sporadic disease.

[0042] In particular, FGFR3-associated craniosynostosis is Muenke syndrome, which is caused by expression of the constitutively active P250R mutant of the FGFR3 receptor.

[0043] In particular, FGFR3-associated craniosynostosis is Crouzon syndrome with acanthosis nigricans (CAN), which is caused by expression of the constitutively active A391E mutant of the FGFR3 receptor.

[0044] As used herein, the term "craniosynostosis" has its common meaning in the art and relates to a condition in which one or more of the fibrous sutures of a subject's skull fuse earlier than normal by turning into bone (ossification), thereby altering the growth pattern of the skull. "Crouzon syndrome with acanthosis nigricans (CAN)" is an extremely rare form of craniosynostosis.

[0045] As used herein, the term "acanthosis nigricans" refers to a brown to black, poorly defined, velvety hyperpigmentation of the skin.

[0046] As used herein, the term "treatment" or "treating" refers to both preventative or prophylactic treatments, including treatments of patients at risk of or suspected of having a disease, as well as patients who have been diagnosed as ill or suffering from a disease or medical condition, and curative, patient-improving, or disease-modifying treatments, including suppression of clinical recurrence. Treatments may be administered to subjects with a medical disorder or at risk of eventually acquiring a disorder, to prevent, cure, delay the onset of, reduce the severity of, or improve one or more symptoms of the disorder or recurrent disorder, or to prolong the survival of the subject beyond that expected in the absence of such treatment. "Therapeutic regimen" refers to a pattern of treatment of a disease, e.g., a pattern of dosing used during treatment. The therapeutic regimen may include induction regimens and maintenance regimens. The phrase "induction regimen" or "induction period" refers to a therapeutic regimen (or a portion of a therapeutic regimen) used for the initial treatment of a disease. The general goal of an induction regimen is to provide high levels of drug to the patient during the initial period of the treatment regimen. The induction regimen may employ (partially or entirely) a "loading regimen", which may include administering a higher dose of drug than the physician would employ during a maintenance regimen, administering a drug more frequently than the physician would administer during a maintenance regimen, or both. The phrase "maintenance regimen" or "maintenance period" refers to a treatment regimen (or a portion of a treatment regimen) used to maintain a patient during disease treatment, for example, to keep the patient in remission for an extended period of time (months or years). A maintenance regimen may employ continuous therapy (e.g., administering a drug at regular intervals, e.g., daily, weekly, monthly, yearly, etc.) or intermittent therapy (e.g., discontinued treatment, intermittent treatment, treatment upon relapse, or treatment upon reaching a certain predetermined criterion (e.g., symptoms of disease, etc.).

[0047] As used herein, the term "preventing" is intended to characterize a prophylactic method or process aimed at delaying or preventing the onset of the disorder or condition to which such term applies.

[0048] The term "expression", when used in relation to the expression of a gene or nucleic acid, refers to the conversion of the information contained in the gene into a gene product.A gene product can be the direct transcription product of a gene (e.g., mRNA, tRNA, rRNA, antisense RNA, ribozyme, structural RNA or any other type of RNA) or a protein (i.e., FGFR3) produced by translation of an mRNA.

[0049] As used herein, the term "inhibitor" includes not only a drug for inhibiting the activity of a target molecule, but also a drug for inhibiting the expression of a target molecule.

[0050] In one embodiment, the inhibitor of the present invention is a tyrosine kinase inhibitor.As used herein, the term "FGFR3 tyrosine kinase inhibitor" (TKI) refers to a compound (natural or synthetic) that is effective in inhibiting the tyrosine kinase activity of FGFR3.In addition, inhibitors that have specific activity against tyrosine kinase can be preferred.

[0051] Examples of FGFR3 tyrosine kinase inhibitors include PD173074 (CAS No. 219580-11-7), AZD4547 (CAS No. 1035270-39-3), BGJ398 (CAS No. 872511-34-7), AP24534 (CAS No. 943319-70-8), BIBF1120 (CAS No. 656247-17-5), JNJ-42756493 (CAS No. 1346242-81-6), TKI-258 (CAS No. 405169-16-6), PHA-739358 (CAS No. 827318-97-8), BMS-540215 (CAS No. 649735-46-6), TKI-258 dilactic acid (CAS No. 852433-84-2), MK-2461 (CAS No. 917879-39-1), BMS-582664 (CAS No. 649735-63-7), SSR128129E (CAS No. 848318-25-2), PRN1371 (CAS No. 1802929-43-6), PD166866 (CAS No. 192705-79-6), BLU554 (CAS No. 1707289-21-1), S49076 (CAS No. 1265965-22-7), SU5402 (CAS No. 215543-92-3), BLU9931 (CAS No. 1538604-68-0), FIN-2 (CAS No. 1633044-56-0), TKI-258 lactate (CAS No. 915769-50-5), CH5183284 (CAS No. 1265229-25-1), LY2874455 (CAS No. 1254473-64-7) or ASP5878 (CAS No. 1453208-66-6). As is well recognized, the CAS (chemical abstracts service) number assigned to each molecule is a unique identifier for each compound. In one embodiment, the FGFR3 tyrosine kinase inhibitor is TYRA-300 from Tyra Biosciences Inc.

[0052] In certain embodiments, the FGFR3 tyrosine kinase inhibitor is BGJ398, which is a potent inhibitor of FGFR3 family.As used herein, the term "BGJ398" has its general meaning in the art and refers to 3-(2,6-dichloro-3,5-dimethoxyphenyl)-1-[6-[4-(4-ethylpiperazin-1-yl)anilino]pyrimidin-4-yl]-1-methylurea.This term is also known as infigratinib, NVP-BGJ398, or BGJ-398.

[0053] As used herein, the term "administering" or "administration" refers to the act of injecting or otherwise physically delivering a substance (e.g., FGFR3 tyrosine kinase inhibitor) when present outside the body to a subject, such as by mucosal, intradermal, intravenous, subcutaneous, intramuscular delivery and / or any other physical delivery method described herein or known in the art.When a disease or its symptoms are treated, administration of the substance is typically performed after the onset of the disease or its symptoms.When a disease or its symptoms are prevented, administration of the substance is typically performed before the onset of the disease or its symptoms.

[0054] "Therapeutically effective amount" refers to an amount effective at the dosage and duration required to achieve the desired therapeutic result. The therapeutically effective amount of a drug may vary depending on factors such as the disease state, age, sex, and weight of the individual, and the ability of the drug to elicit a desired response in the individual. A therapeutically effective amount is also an amount in which any toxic or adverse effects of the antibody or antibody portion are outweighed by the therapeutically beneficial effects. The effective dosage and dosing regimen of the drug depends on the disease or condition to be treated and can be determined by one of ordinary skill in the art. A physician of ordinary skill in the art can easily determine and prescribe the effective amount of the pharmaceutical composition required. For example, a physician can start the dose of the drug used in the pharmaceutical composition at a level lower than required to achieve the desired therapeutic effect, and gradually increase the dosage until the desired effect is achieved. In general, a suitable dose of the composition of the present invention will be the amount of the compound that is the lowest dose effective to produce a therapeutic effect according to a particular dosing regimen. Such an effective dose will generally depend on the factors described above. For example, a therapeutically effective amount in therapeutic applications can be measured by its ability to stabilize the progression of a disease. One of skill in the art would be able to determine such amounts based on factors such as the size of the subject, the severity of the subject's symptoms, and the particular composition or route of administration selected. An exemplary, non-limiting range for a therapeutically effective amount of a drug is about 0.1-100 mg / kg, such as about 0.1-50 mg / kg, such as about 0.1-20 mg / kg, such as about 0.1-10 mg / kg, for example, about 0.5, such as about 0.3, about 1, about 3 mg / kg, about 5 mg / kg, or about 8 mg / kg. Administration may be, for example, intravenous, intramuscular, intraperitoneal, or subcutaneous, but may be administered proximal to the target site, for example. Dosage regimens in the above methods and uses of treatment are adjusted to produce the optimum desired response (e.g., therapeutic response). For example, a single bolus may be administered, several divided doses may be administered over time, or the dose may be proportionally reduced or increased as indicated by the exigencies of the therapeutic situation. In some embodiments, the effectiveness of treatment is monitored during therapy, eg, at predetermined time points.As a non-limiting example, treatment according to the present invention may be administered at a daily dose of about 0.1 to 100 mg / kg per day, e.g., 0.2, 0.5, 0.9, 1.0, 1.1, 1.5, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 45, 50, 60, 70, 80, 90 or 100 mg / kg of the agent of the present invention, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 45, 50, 60, 70, 80, 90 or 100 mg / kg, or at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 45, 50, 60, 70, 80, 9 on at least one of days 6, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39 or 40, or alternatively on at least one of weeks 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20, or any combination thereof, using single or divided doses every 24, 12, 8, 6, 4 or 2 hours, or any combination thereof.

[0055] In some embodiments, the patient is administered a pharmaceutical composition comprising a therapeutically effective amount of an FGFR3 tyrosine kinase inhibitor as an active ingredient and at least one pharma- ceutically acceptable excipient.

[0056] As used herein, the terms "active principle" or "active ingredient" are used interchangeably. As used herein, the term "pharmaceutical composition" refers to a composition described herein or a pharma- ceutically acceptable salt thereof with other agents, such as carriers and / or excipients. Pharmaceutical compositions as provided herein typically include a pharma- ceutically acceptable carrier.

[0057] As used herein, the term "pharmaceutical acceptable carrier" includes any and all solvents, diluents, or other liquid media, dispersion or suspension aids, surfactants, isotonicity agents, thickening or emulsifying agents, preservatives, solid binders, lubricants, and the like, as appropriate for the particular dosage form desired. Remington's Pharmaceutical-Sciences, Sixteenth Edition, EW Martin (Mack Publishing Co., Easton, Pa., 1980) discloses various carriers used in formulating pharmaceutical compositions and known techniques for their preparation.

[0058] Pharmaceutically acceptable carriers or excipients refer to any kind of non-toxic solid, semi-solid or liquid fillers, diluents, encapsulating materials or formulation aids. Typically, pharmaceutical compositions contain a pharma- ceutically acceptable medium for injectable preparations. These may be, in particular, isotonic sterile saline solutions (monosodium or disodium phosphate, sodium chloride, potassium chloride, calcium chloride or magnesium chloride, etc., or mixtures of such salts), or dry, especially lyophilized compositions (optionally allowing the constitution of an injectable solution by adding sterile water or saline). Pharmaceutical dosage forms suitable for injection use include sterile aqueous solutions or dispersions; formulations containing sesame oil, peanut oil or aqueous propylene glycol; and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. In any case, the dosage form must be sterile and must be fluid to the extent that it can be easily injected. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms, such as bacteria and fungi. Sterile injectable solutions are prepared by adding the required amount of the agent of the present invention to a suitable solvent with various other ingredients as listed above, as necessary, followed by filtration sterilization.Generally, dispersions are prepared by adding various sterilized active ingredients to a sterile medium that contains the basic dispersion medium and other necessary ingredients from those listed above.In the case of sterile powders for preparing sterile injectable solutions, the preferred preparation method is vacuum drying and freeze-drying technology, which produces powder from a previously sterile-filtered solution of active ingredient and any additional desired ingredient.

[0059] The present invention will be further illustrated by the following figures and examples, which should not, however, be construed as limiting the scope of the present invention in any way. [Brief description of the drawings]

[0060] [Figure 1A](A) MicroCT scan analysis of mandibles 14 days after fracture. Callus volume is unchanged in treated Fgfr3N534K / + compared to untreated Fgfr3N534K / +. BV / TV is increased in treated Fgfr3N534K / + mandibles. [Figure 1B] (B) MicroCT scan analysis of mandibles 28 days after fracture. Callus volume is decreased in treated Fgfr3N534K / + compared to untreated Fgfr3N534K / + mutants. BV / TV is increased in treated Fgfr3N534K / + mandibles. [Figure 1C] (C) Grade repair at day 28. *p<0.05, **p<0.01, ***p<0.001 EXAMPLES

[0061] Definition: As used herein, the term "FGFR3 N534K / + " concerns a mouse model of HCH. Mutant mice exhibit the clinical picture of HCH with growth defects, growth plate congenital anomalies, partial loss of synchondroses, and lordosis. The bones of adult HCH animals have reduced bone mineral density and their bone structure has some characteristics of osteoporotic bone, placing them at high risk of fracture in old age.

[0062] As used herein, the term "FGFR3 Y367C / + " concerns a mouse model that recapitulates the human ACH phenotype and clinical findings of ACH, such as dwarfism with reduced foramen magnum, mandibular hypoplasia, hearing loss, congenital anomalies of the intervertebral disc, chondrocyte proliferation and differentiation defects, and impaired ciliary formation (Pannier et al. 2009, 2010; Mugniery et al. 2012; Di Rocco et al. 2014; Biosse Duplan et al. 2016; Komla Ebri et al. 2016; Martin et al. 2018).

[0063] As used herein, the term "FGFR3 A385E / +" relates to a CAN mouse model in which memory defects were observed.

[0064] result I-Mouse model of hypochondroplasia (Fgfr3 N534K / + A study of mandibular bone repair in HCH mouse model (Fgfr3 N534K / + ) to study mandibular bone repair. The experiments were performed in 6-week-old adult animals and mice were euthanized at four key time points: during bone repair (10, 14, and 21 days after fracture) and at the end point of the normal bone healing process, i.e., 28 days after fracture.

[0065] Fgfr3 N534K / + and Fgfr3 + / + Mice underwent vertical mandibular fractures in the ascending ramus region, a non-stabilized mandibular fracture protocol that allows for the analysis of endochondral bone repair processes.

[0066] Fgfr3 is involved in multiple key steps of repair N534K / + and Fgfr3 + / + Four batches (n=100) of mandibular fractures from mice (days 10, 14, 21, and 28) were studied. Using collagen type II (proliferative cartilage), type X (hypertrophic cartilage), and type I (bone) immunolabeling, it was observed that endochondral processes were disrupted, and Fgfr3 + / + Compared with Fgfr3 N534K / + In the 28-day post-fracture study, we noticed delayed cartilage resorption, a decrease in the size of the hypertrophic chondrocyte area, and defective bone formation within the repair callus. To complete these data, we performed a histomorphometric analysis of the callus using Alcian blue / picrosirius staining. N534K / + We observed the presence of pseudoarthrosis (fibrosis in the callus) in mice, but Fgfr3 + / + 100% bone was observed in the callus. Considering the quality of the newly formed bone on day 28, four repair grades were defined and mice were classified according to the repair grade (1-4). Incomplete repair was observed when Fgfr3 + / + Compared with Fgfr3 N534K / + It was significant in (Figure 1C).

[0067] To analyze the bone microstructure, microCT scans were performed. + / + Compared with Fgfr3 N534K / + We observed microstructural changes in callus formation from days 10 to 28. Bone volume / total volume (BV / TV) was significantly decreased in mutants compared to controls at days 10 (-23%, p<0.05), 14 (-14%, p<0.01), 21 (-14.9%, p<0.005) and 28 (-5.8%, p<0.05) (Figures 1A and 1B).

[0068] Here, we conclude that Fgfr3 gain-of-function impaired bone repair in a model of non-stabilized mandibular fractures in HCH mice: 1) bone formation was altered and bone architecture had some characteristics of osteoporotic bone, and 2) formation of pseudoarthrosis in the callus was observed, demonstrating that endochondral ossification was severely disturbed in the mandible.

[0069] II-Improvement of mandibular formation and repair through therapeutic approaches Current therapeutic approaches to address aberrant FGF signaling are numerous and varied, including FGFR3 tyrosine kinase inhibitors (BGJ398), which target signaling pathways downstream of FGFR3.

[0070] Tyrosine kinase inhibitors (TKIs) inhibit the activity of receptor tyrosine kinases. The efficacy of infigratinib was demonstrated in a mouse model of ACH (Fgfr3 Y367C / + Treated mice showed improvements in many of the skeletal elements affected by ACH, including long bones, vertebrae, intervertebral discs, craniofacial geometry, and the foramen magnum (Komla Ebri et al., 2016; Biosse Duplan et al., 2016).

[0071] From day 0 (fracture of the mandible) to day 14 or 28, HCH mice (Fgfr3 N534K / +) were treated with infigratinib (4 mg / kg subcutaneously, 3 times a week). Bone repair analysis was evaluated by microCT scanning on days 14 and 28 after fracture (Figures 1A and 1B). BV / TV (bone volume / total volume) and callus volume (TV (total volume) of callus / TV of the contralateral ascending ramus region) were quantified.

[0072] Callus volume is unchanged at 14 days after fracture, but at 28 days after fracture a significant decrease in callus volume is observed in treated HCH mice (n=10) compared to untreated HCH mice (n=14) (-38.6%, p<0.0001). Improvements in BV / TV are observed in treated HCH mice (n=7) compared to untreated HCH mice (n=9) at 14 days after fracture (+20.8%, p<0.05) and at 28 days after fracture in treated HCH mice (n=10) compared to untreated HCH mice (n=14) (+17.8%, p<0.01) (Figures 1A and 1B).

[0073] Gain-of-function mutations in FGFR3 disrupted bone formation, quality and repair in a mandibular unstabilized fracture model. Callus is abnormal in HCH mice as revealed by quantitative bone analysis by microCT scanning. BV / TV is significantly decreased in HCH mice compared to wild type.

[0074] Four batches of mice (Fgfr3 + / + .Fgfr3 N534K / + and Fgfr3 N534K / + Comparative analysis of mandibular bone repair (BGJ398 + BGJ398) highlighted the incomplete bone formation and repair in HCH mice and the beneficial effects of both treatments in mandibular bone repair. Complete bone healing (grade 1) was observed in 7 / 10 (BGJ398). Restoration of bone continuity and the presence of mild bone defects (grade 2) was observed in 3 / 10 (BGJ398) (Figure 1C). These results demonstrate the beneficial effects of BGJ398 treatment.

[0075] conclusion Taken together, these data provide proof of concept that treatment with an FGFR3 tyrosine kinase inhibitor (BGJ398) improves mandibular bone formation and repair in the context of FGFR3-associated chondrodysplasia.

[0076] References: Throughout this application, various references describe the state of the art to which this invention pertains. The disclosures of these references are incorporated by reference into this disclosure.

Claims

1. A pharmaceutical composition for the treatment of an FGFR-associated bone repair and bone formation disorder in a subject in need thereof, the pharmaceutical composition comprising an FGFR3 tyrosine kinase inhibitor.

2. The pharmaceutical composition of claim 1 , wherein the subject is a child or an adult.

3. The pharmaceutical composition of claim 1, wherein the FGFR3 tyrosine kinase inhibitor is BGJ398.

4. The pharmaceutical composition of claim 1 , wherein the subject has a gain-of-function mutation of FGFR.

5. The pharmaceutical composition according to claim 4, wherein the gain-of-function mutation of FGFR is an FGFR3-associated skeletal disease.

6. 6. The pharmaceutical composition of claim 5, wherein the FGFR3-related skeletal disease is hypochondroplasia (HCH), achondroplasia (ACH), thanatophoric dysplasia (TD), severe achondroplasia with developmental delay and acanthosis nigricans (SADDAN), Muenke syndrome, Crouzon syndrome with acanthosis nigricans, dwarfism, or craniosynostosis.

7. The pharmaceutical composition according to claim 6, wherein the FGFR3-associated skeletal disease is hypochondroplasia (HCH).

8. The pharmaceutical composition according to claim 6, wherein the FGFR3-associated skeletal disease is achondroplasia (ACH).

9. The pharmaceutical composition according to claim 6, wherein the FGFR3-related skeletal disease is craniosynostosis.

10. The pharmaceutical composition according to claim 9, wherein the craniosynostosis is Crouzon syndrome with acanthosis nigricans (CAN).

11. The pharmaceutical composition according to claim 6, wherein the FGFR3-associated skeletal disease is Muenke syndrome.

12. The pharmaceutical composition of claim 1, further comprising a pharmaceutically acceptable excipient.

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