Treatment of osteogenesis imperfecta

Abstract

The present disclosure provides methods of treating and ameliorating osteogenesis imperfecta (OI) in a subject by administering to the subject a therapeutically effective amount of an agent that binds to and neutralizes transforming growth factor beta (TGF-β).

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority from U.S. Provisional Patent Application No. 63 / 274,503, filed November 1, 2021, and European Patent Application Publication No. 22315238.0, filed October 13, 2022. The disclosures of the two priority applications are incorporated herein by reference in their entireties.

[0002] Sequence Listing This application contains a Sequence Listing that has been submitted electronically in ASCII format and is incorporated herein by reference in its entirety. Said ASCII copy, created on October 26, 2022, is named 022548WO028.XML and is 20,163 bytes in size. [Background technology]

[0003] Osteogenesis imperfecta (OI) is a genetically and phenotypically heterogeneous, Mendelian disorder of the skeletal system with an estimated prevalence of 1 in 10,000-20,000 births. Skeletal manifestations of OI include low bone mass, bone fragility, recurrent fractures, scoliosis, and bone deformities. Extraskeletal manifestations include loss of muscle mass, muscle weakness, dentinogenesis imperfecta, hearing loss, and pulmonary disease (Marini, Nat Rev Dis Primers (2017) 3:17052; Marom et al., Am J Med Genet C Semin Med Genet. (2016) 172(4):367-83; Patel et al., Clin Gen. (2015) 87(2):133-40; Rossi et al., Curr Opin Pediatr. (2019) 31(6):708-15; Tam et al., Clin Gen. (2018) 94(6):502-11; DiMeglio et al. J Bone Miner Res. (2006) 21:132-40; Gatti et al., J Bone Miner Res. (2005) 20(5):758-63); Gatti et al., Calcified Tissue Int. (2013) 93(5):448-52). Management of individuals with OI usually involves a multidisciplinary approach.Mainstream treatments for OI bone fragility include repurposing of drugs used to treat osteoporosis (Adami et al., J Bone Miner Res. (2003) 18(1): 126-30; Bishop et al., Ear Hum Dev. (2010) 86(11): 743-6; Chevrel et al., J Bone Miner Res. (2006) 21(2): 300-6; Glorieux et al., NEJM (1998) 339(14): 947-52; Rauch et al., J Bone Miner Res. (2009) 24(7): 1282-9; Rauch et al., J Bone Miner Res. (2003) 18(4): 610-4; Orwoll et al., J Clin Invest (2014) 124(2): 491-8; Hoyer-Kuhn et al. al., J Musculoskelet Neuronal Interact. (2016)16(1):24-32; Anissipour et al., J Bone Joint Surg Am. (2014)96(3):237-43).

[0004] Despite significant progress, most pharmacological interventions in OI have not provided the disease-modifying effects observed in other bone diseases such as osteoporosis and are less effective in certain types of OI. Due to the complexity of the underlying biochemical processes involved in OI and bone remodeling, translation of pharmacokinetic / pharmacodynamic (PK / PD) relationships to the clinic is challenging. Bisphosphonates (BPNs), a class of antiresorptive drugs that reduce bone resorption, are currently the standard of care, especially in pediatric OI. In children, BPN has been shown to have beneficial effects on areal and volumetric bone mineral density (aBMD and vBMD), progression of scoliosis, quality of life, and in some studies, fracture incidence (Bishop et al., Lancet (2013) 382(9902):1424-32; Rauch et al., 2003, supra; Bains et al., JBMR Plus (2019) 3(5):e10118; Rauch et al., Bone (2007) 40(2):274-80). However, given the heterogeneity of OI and variability in clinical study design, the effects of BPN are inconsistent. In adults, the benefits and outcomes of long-term treatment with bisphosphonates are less certain (Adami et al., supra; Shi et al., Am J Ther. (2016) 23(3):e894-904). Furthermore, in a randomized trial involving adults with OI, treatment with the anabolic agent teriparatide resulted in increases in aBMD and vBMD in individuals with the mild form (OI type I) but not in moderate to severe forms of impairment (OI types III and IV). Furthermore, none of these repurposed therapies address specific pathogenic mechanisms in OI and therefore have no effect on extraskeletal manifestations. Summary of the Invention [Problem to be solved by the invention]

[0005] Thus, there remains a large unmet need for effective treatments that target various forms of OI. [Means for solving the problem]

[0006] The present disclosure provides a method of treating osteogenesis imperfecta (OI) in a human subject in need thereof, comprising administering to the subject a therapeutically effective amount of an anti-TGF-β antibody, wherein the antibody comprises heavy chain complementarity determining regions (CDRs) 1-3 comprising SEQ ID NOs: 4-6, respectively, and light chain CDRs 1-3 comprising SEQ ID NOs: 7-9, respectively, and wherein the antibody comprises a human IgG4 constant region having a proline at position 228 (Eu numbering), wherein the therapeutically effective amount is 1-8 mg / kg, optionally 2, 2.5 or 5 mg / kg, administered twice a year (e.g., every 6 months or Q6M) or 0.1-1 mg / kg, optionally 0.35, 0.4 or 0.5 mg / kg, administered every 3 months (Q3M).

[0007] In some embodiments, the antibody herein comprises a heavy chain variable domain comprising SEQ ID NO: 10 and a light chain variable domain comprising SEQ ID NO: 11. In a further aspect, the antibody comprises a human IgG4 constant region and / or a human kappa light chain constant region. In a particular embodiment, the antibody comprises or consists of a heavy chain comprising SEQ ID NO: 3 and a light chain comprising SEQ ID NO: 2.

[0008] In some embodiments, the antibody comprises a bone-targeting moiety, and optionally the bone-targeting moiety is a polyarginine peptide (e.g., SEQ ID NO: 14). In further embodiments, the antibody comprises one or more polyarginine peptides. In certain embodiments, the antibody is fused to a polyarginine peptide at the N-terminus or C-terminus or both termini of the antibody heavy chain and / or the C-terminus of the antibody light chain.

[0009] In some embodiments, the OI treated herein is moderate to severe OI or type IV OI, hi some embodiments, the OI is type I, type II, or type III.

[0010] In some embodiments, the human subject is an adult patient (age 18 or older) or a pediatric patient (age under 18). In some embodiments, the human subject has a mutation in the COL1A1 or COL1A2 gene, optionally, the mutation is a glycine substitution mutation in the COL1A1 or COL1A2 gene or a valine deletion in the COL1A2 gene.

[0011] In some embodiments, the treatments herein improve bone parameters selected from the group consisting of bone mineral density (BMD), bone volume density (BV / TV), total bone surface (BS), bone surface density (BS / BV), trabecular number (Tb.N), trabecular thickness (Tb.Th), trabecular spacing (Tb.Sp) and total volume (Dens TV).

[0012] In some embodiments, the treatments herein decrease bone turnover and / or bone cell density, optionally, decreased bone turnover is indicated by a decrease in serum CTX or an increase in serum osteocalcin (OCN).

[0013] In some embodiments, the antibody is administered at 2 mg / kg twice a year or 0.4 mg / kg Q3M, optionally, administration results in about a 5% increase in BMD in the subject. In some embodiments, the antibody is administered at 5 mg / kg twice a year or 0.5 mg / kg Q3M, optionally, administration results in about a 5% increase in BV in the subject. In some embodiments, the antibody is administered at 2.5 mg / kg twice a year or 0.35 mg / kg Q3M, optionally, administration results in a decrease in TGF-β levels to homeostatic values ​​in the subject.

[0014] In some embodiments, the antibody is administered by intravenous infusion. In some embodiments, the treatment herein includes another therapeutic agent, such as a bisphosphonate, parathyroid hormone, calcitonin, teriparatide, or an anti-sclerostin agent. In further embodiments, the bisphosphonate is selected from alendronate, pamidronate, zoledronate, and risedronate.

[0015] Also provided herein are anti-TGF-β antibodies for use in the treatment of osteogenesis imperfecta in the subject therapeutic methods; use of anti-TGF-β antibodies in the manufacture of a medicament for the treatment of osteogenesis imperfecta in the subject methods; and articles of manufacture or kits comprising anti-TGF-β antibodies for use in the treatment of osteogenesis imperfecta in the subject methods.

[0016] Also provided herein are anti-TGF-β antibodies, or antigen-binding fragments thereof, for use in the treatment of osteogenesis imperfecta in the therapeutic methods herein, and the use of anti-TGF-β antibodies, or antigen-binding fragments thereof, in the manufacture of a medicament for the treatment of osteogenesis imperfecta in the therapeutic methods herein.

[0017] Also provided are articles of manufacture (eg, kits) that contain anti-TGF-β antibodies, or antigen-binding fragments thereof, for use in treating osteogenesis imperfecta in the therapeutic methods herein.

[0018] Other features, objects and advantages of the present invention will become apparent from the following detailed description. It should be understood, however, that this detailed description, while illustrating embodiments and aspects of the present invention, is given by way of illustration only, not by way of limitation. Various changes and modifications within the scope of the present invention will become apparent to those skilled in the art from this detailed description. [Brief description of the drawings]

[0019] [Figure 1]Graphs showing a multi-model approach to assess the concentration-response relationship of Ab1 (GC2008) to bone mass density (BMD), bone strength and TGF-β kinetics in bones of OI patients. FIG. 1A shows PK / PD modeling based on clinical data with fresolimumab (GC1008), a fully human anti-TGF-β antibody. FIG. 1B shows PK / PD modeling based on preclinical data with 1D11, a murine anti-TGF-β antibody (U.S. Pat. No. 5,571,714; ATCC deposit no. HB9849; available, for example, at Thermo Fisher catalog no. MA5-23795). FIG. 1C shows physiologically based pharmacokinetic modeling (PBPK) based on physicochemical (PC) properties of Ab1, another fully human anti-TGF-β antibody. [Diagram 2] 1 is a pair of graphs showing the use of PK data of fresolimumab (1 mg / kg or 4 mg / kg intravenous ("IV") dose) in serum of patients with focal segmental glomerulosclerosis (FSGS) in informing the PK / BMD response of fresolimumab in patients with OI. [Diagram 3] 1 is a pair of graphs showing the use of Ab1 PK data in predicting PK / BMD response to Ab1 in patients with OI. [Figure 4]FIG. 1 is a panel of graphs showing the use of mouse 1D11 PK data in predicting PK / BV response of Ab1 in OI patients. Graph A shows concentration vs. time (left axis) and bone volume fraction (right axis) for 5 mg / kg dosing three times per week in mice. Graph B shows concentration vs. time (left axis) and bone volume fraction (right axis) for 5 mg / kg dosing weekly in mice. Graph C shows concentration vs. time (left axis) and bone volume fraction (right axis) for 5 mg / kg dosing every 2 weeks in mice. Graph D shows concentration vs. time (left axis) and bone volume fraction (right axis) for 5 mg / kg dosing every 4 weeks in mice. Graph E shows concentration vs. time (left axis) and bone volume fraction (right axis) for 0.5 mg / kg dosing every 3 months in humans. Graph F shows concentration vs. time (left axis) and bone volume fraction (right axis) for 5 mg / kg dosing every 6 months in humans. Symbols are mean bone volume fraction data and error bars indicate their standard deviations. [Diagram 5] FIG. 1 is a panel of graphs showing the use of physicochemical properties of Ab1 in modeling physiologically based PK (PBPK) responses of Ab1 in patients with OI. Graph A shows concentrations of Ab1 for single IV doses of 0.05, 0.25, 1, and 3 mg / kg. Symbols are individual subject data and lines show PBPK model predictions. Graph B shows a comparison of plasma (solid line) and bone (dotted line) PK for a single IV dose of 0.05 mg / kg Ab1. Graph C shows plasma PK predictions for 0.35 mg / kg dosing every 3 months and 2.5 mg / kg dosing every 6 months. Graph D shows TGFβ target kinetics in bone following dosing of Ab1 at 0.35 mg / kg every 3 months or 2.5 mg / kg every 6 months. [Figure 6] FIG. 1 is a panel of graphs showing Ab1 population PK assessment plots. Graph A shows observed Ab1 concentrations vs. individual predictions. Graph B shows observed Ab1 concentrations vs. population predictions. Graph C shows normalized prediction distribution error vs. Ab1 population predictions. Graph D shows normalized prediction distribution error vs. time. The straight line represents the identity (y=x) line and the curve is a spline interpolation. [Figure 7]1D11 PK response following 5 mg / kg IP administration in OI mice. Circles represent OI mouse data and solid lines represent one-compartment model simulation. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0020] The present disclosure provides a method for treating OI in a human patient by administering a monoclonal antibody that binds to and neutralizes all isoforms of human TGF-β. The method is developed based on a multiple model-based approach that relies on preclinical and clinical PK and PD data to inform the concentration-response relationship of anti-TGF-β antibody Ab1 and its impact on bone mineral density (BMD), bone strength and TGF-β expression levels in OI patients.

[0021] I. Osteogenesis imperfecta OI encompasses a group of congenital bone disorders characterized by the deficiency of one or more proteins involved in bone matrix deposition or homeostasis. There are more than 19 types of OI defined by their specific genetic mutations, the resulting protein deficiencies, and the phenotype of affected individuals. Classification includes findings on x-rays and other imaging studies. The main types of OI are as follows (information from the John Hopkins University website):

[0022] Type I is the mildest and most common type. Approximately 50% of all affected children have this type. Fewer fractures and deformities occur.

[0023] Type II is the most severe type. Infants have very short arms and legs, a small chest, and a soft skull. Infants may be born with fractured bones and may have low birth weight and underdeveloped lungs. Infants with Type II OI usually die within the first few weeks of life.

[0024] Type III is the most severe type in infants who do not die as newborns. At birth, infants may have slightly shorter than normal arms and legs and fractures of the arms, legs and ribs. Infants may also have a larger than normal head, a triangular shaped face, a deformed chest and spine, and breathing and swallowing problems.

[0025] Type IV is a type of OI where symptoms range from mild to severe. Babies with Type IV may be diagnosed at birth. Babies may not break bones until they begin to crawl or walk. The bones in the arms and legs may not be straight. Babies may not grow normally.

[0026] Type V is similar to Type IV. Symptoms can be moderate to severe. Enlarging areas of thickening (hypertrophic callus) are common in areas where large bones are fractured.

[0027] Type VI is very rare. Symptoms are moderate and similar to type IV.

[0028] Type VII may be similar to Type IV or Type II. It is typically shorter than normal height. It is also typically shorter than normal upper arms and femurs.

[0029] Type VIII is similar to types II and III. Patients have very soft bones and severe growth retardation.

[0030] The phenotype varies depending on the type of OI, but common symptoms include incomplete ossification of bones and teeth, bone loss, brittle bones, and pathological fractures. Specific symptoms include easy fractures, bone deformities (e.g., bowing of the legs), discoloration of the whites of the eyes (sclera), barrel chest, curved spine, triangular face, loose joints, muscle weakness, easily bruised skin, hearing loss in early adulthood, and / or soft, discoloured teeth. Complications of OI include respiratory infections (e.g., pneumonia), heart problems (e.g., poor heart valve function), kidney stones, joint problems, hearing loss and abnormal eye conditions (including vision loss). OI can be diagnosed or monitored by x-rays, laboratory tests (e.g., blood and genetic tests), dual energy x-ray absorptiometry scans (DXA or DEXA scans), and bone biopsies.

[0031] Although multiple pathogenic gene mutations can cause the various subtypes of OI, more than 90% are caused by pathogenic variants in the COL1A1 gene (encoding the α1 chain of type I collagen) or the COL2A1 gene (encoding the α1 chain of type II collagen) or in genes encoding proteins that post-translationally modify type I collagen (CRTAP, PPIB, and LEPRE1) (Patel et al., supra; Lim et al., Bone (2017) 102:40-49).

[0032] In some embodiments, the patient's OI is caused by a mutation (eg, a glycine substitution) in COL1A1 or COL1A2 or a biallelic pathogenic variant in CRTAP, PPIB, or LEPRE1.

[0033] II. Anti-TGF-β antibody TGF-β is a multifunctional cytokine involved in cell proliferation and differentiation, embryonic development, extracellular matrix formation, bone development, wound healing, hematopoiesis, and immune and inflammatory responses. Secreted TGF-β protein is cleaved into latency-associated peptide (LAP) and mature TGF-β peptide, which is found in latent and active forms. Mature TGF-β peptide forms both homodimers and heterodimers with other TGF-β family members.

[0034] There are three human (h)TGF-β isoforms: TGF-β1, TGF-β2, and TGF-β3 (UniProt accession numbers P01137, P08112, and P10600, respectively). TGF-β1 differs from TGF-β2 by 27 and from TGF-β3 by 22, mostly conservative amino acids. Human TGF-β is highly similar to mouse TGF-β. Human TGF-β1 has only one amino acid difference from mouse TGF-β1; human TGF-β2 has only three amino acid differences from mouse TGF-β2; and human TGF-β3 is identical to mouse TGF-β3.

[0035] Binding of TGF-β proteins to homodimeric or heterodimeric TGF-β transmembrane receptor complexes activates the canonical TGF-β signaling pathway mediated by intracellular SMAD proteins. Deregulation of TGF-β leads to pathological processes that are involved in many conditions in humans, such as birth defects, cancer, chronic inflammatory diseases, autoimmune diseases, and fibrotic diseases (see, e.g., Border et al., Curr Opin Nephrol Hypertens. (1994) 3(4): 446-52; Border et al., Kidney Int Suppl. (1995) 49: S59-61).

[0036] In the present invention, the anti-TGF-β antibody can be a pan-specific antibody, i.e., an antibody that binds with high affinity to and neutralizes all three isoforms of TGF-β. In some embodiments, the antibody is fresolimumab. Fresolimumab is a recombinant human antibody. Its heavy chain is shown below. [ka] In the above sequence, positions 1 to 120 correspond to the heavy chain variable domain (V H ), and the heavy chain CDRs ("HCDRs"; Kabat definition) are framed. The heavy chain comprises a human IgG4 constant region.

[0037] The light chain of fresolimumab is shown below. [ka] In the above sequence, positions 1 to 108 correspond to the light chain variable domain (V L ), with the light chain CDRs ("LCDRs"; Kabat definition) underlined. The light chain comprises a human Cκ constant region.

[0038] In some embodiments, the anti-TGF-β antibody herein is Ab1, a variant of fresolimumab. The heavy chain of Ab1 differs from that of fresolimumab only at a residue in the IgG4 hinge region. The residue is S228 (Eu numbering), and Ab1 has a proline at that position, i.e., an S228P substitution relative to fresolimumab. Ab1 and fresolimumab have the same light chain. The heavy chain of Ab1 is shown below. [ka] In the above sequence, the HCDRs are boxed and the S228P substitution is in bold.

[0039] In some embodiments, the anti-TGF-β antibody comprises one or more (e.g., all six) of the HCDRs 1-3 and LCDRs 1-3 of fresolimumab. In other words, the antibody comprises one or more (e.g., all six) of the following HCDRs and LCDRs: HCDR1 SNVIS (SEQ ID NO: 4) HCDR2 GVIPIVDIANYAQRFKG (SEQ ID NO: 5) HCDR3 TLGLVLDAMDY (SEQ ID NO: 6) LCDR1 RASQSLGSSYLA (SEQ ID NO: 7) LCDR2 GASSRAP (SEQ ID NO: 8) LCDR3 QQYADSPIT (SEQ ID NO: 9)

[0040] In some embodiments, the anti-TGF-β antibody is fresolimumab or the V Hand / or V L In other words, the antibody comprises the following sequence: V H : [ka] V L : [ka] Includes one or both of the following:

[0041] In some embodiments, the anti-TGF-β antibody is of the human IgG isotype, such as the human IgG4 isotype. In certain embodiments, the human IgG4 constant region has the following amino acid sequence: [ka] In further embodiments, the human IgG4 constant region has a mutation at position 228 (Eu numbering). In some embodiments (e.g., Ab1), the mutation is a serine to proline mutation (S228P). In the above sequence, S228 serine is framed.

[0042] In some embodiments, the anti-TGF-β antibody (e.g., Ab1 and fresolimumab) comprises a human kappa light chain constant region (Cκ). In certain embodiments, the human Cκ has the amino acid sequence: [ka] Includes.

[0043] In some embodiments, antigen-binding fragments of complete anti-TGF-β antibodies may also be used. The term "antigen-binding fragment" or similar terms refers to a portion of an antibody that contains amino acid residues that interact with an antigen and confer to the binder its specificity and affinity for the antigen. Non-limiting examples of antigen-binding fragments include: Fab fragments, F(ab')2 fragments, Fd fragments, Fv fragments, single chain Fvs (scFvs), dAb fragments, and minimal recognition units consisting of amino acid residues that mimic the hypervariable domains of an antibody.

[0044] In some embodiments, the antibodies or antigen-binding fragments herein are linked to a bone-targeting moiety. In further embodiments, the bone-targeting moiety is a polyarginine (poly D) peptide. As used herein, the term "poly D peptide" refers to a peptide sequence having a plurality of aspartic acid or aspartic acid or "D" amino acids, such as about 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30 or more aspartic acid amino acids (residues). For example, a poly D peptide can contain about 2 to about 30, or about 3 to about 15, or about 4 to about 12, or about 5 to about 10, or about 6 to about 8, or about 7 to about 9, or about 8 to about 10, or about 9 to about 11, or about 12 to about 14 aspartic acid residues. A poly D peptide can contain only aspartic acid residues or can contain one or more other amino acids or similar compounds. As used herein, the term "D10" refers to a contiguous sequence of 10 aspartic acid amino acids as found in SEQ ID NO: 14. In some embodiments, an antibody or antibody fragment of the invention may comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12 or more than 12 poly-D peptides.

[0045] The poly-D peptide may be linked to the anti-TGF-β antibody or antigen-binding fragment by recombinantly fused such that the poly-D is linked to the antibody or fragment via a peptidyl bond (i.e., the antibody or fragment is a fusion protein). For example, the poly-D peptide may be fused to the N-terminus or C-terminus or both of the heavy chain and / or the N-terminus or C-terminus or both of the light chain. The poly-D peptide may also be linked to the anti-TGF-β antibody or antigen-binding fragment by chemical conjugation, e.g., chemical reaction with cysteine ​​or lysine residues on the antibody or antibody-binding fragment with or without a linker moiety (e.g., maleimide functional groups and polyethylene glycol (PEG)). See, e.g., WO 2018 / 136698.

[0046] In certain embodiments, the antibody is fresolimumab fused to the D10 peptide at the N-terminus, C-terminus, or both termini of the heavy chain. In some aspects, the antibody is fresolimumab fused to the D10 peptide at the C-terminus of the light chain. In certain embodiments, the antibody is fresolimumab fused to the D10 peptide at both termini of the heavy chain and the C-terminus of the light chain.

[0047] In certain embodiments, the antibody is an Ab1 fused to the D10 peptide at the N-terminus, C-terminus, or both termini of the heavy chain. In some embodiments, the antibody is an Ab1 fused to the D10 peptide at the C-terminus of the light chain. In certain embodiments, the antibody is an Ab1 fused to the D10 peptide at both termini of the heavy chain and the C-terminus of the light chain.

[0048] The anti-TGF-β antibodies or antigen-binding fragments thereof of the present disclosure can be produced by methods well established in the art. The DNA sequences encoding the heavy and light chains of the antibody can be inserted into an expression vector such that the genes are operably linked to the necessary expression control sequences, such as transcriptional and translational control sequences. Expression vectors include plasmids, retroviruses, adenoviruses, adeno-associated viruses (AAV), plant viruses, such as cauliflower mosaic virus, tobacco mosaic virus, cosmids, YACs, EBV-derived episomes, and the like. The antibody light chain coding sequence and the antibody heavy chain coding sequence can be inserted into separate vectors and operably linked to the same or different expression control sequences (e.g., promoters). The expression vectors encoding the antibodies of the present disclosure are introduced into host cells for expression. The host cells are cultured under conditions suitable for expression of the antibody, and the antibody is then harvested and isolated. The host cells include mammalian, plant, bacterial, or yeast host cells. Mammalian cell lines available as hosts for expression are well known in the art and include many immortalized cell lines available from the American Type Culture Collection (ATCC). These include, among others, Chinese hamster ovary (CHO) cells, NS0 cells, SP2 cells, HEK-293T cells, 293Freestyle cells (Invitrogen), NIH-3T3 cells, HeLa cells, baby hamster kidney (BHK) cells, African green monkey kidney cells (COS), human hepatocellular carcinoma cells (e.g., Hep G2), A549 cells, and several other cell lines. Cell lines may be selected based on their expression levels. Other cell lines that may be used are insect cell lines such as Sf9 or Sf21 cells. Tissue culture media for host cells may or may not contain animal-derived components (ADCs), such as bovine serum albumin. In some embodiments, culture media without ADCs is preferred for human safety. Tissue culture can be carried out using fed-batch, continuous perfusion, or any other method appropriate for the host cells and desired yield.

[0049] III. Pharmaceutical Compositions and Uses The methods described herein include administering a therapeutically effective amount of an anti-TGF-β antibody or antigen-binding fragment thereof to an OI patient. As used herein, the phrase "therapeutically effective amount" refers to a dose of an antibody that binds to TGF-β that results in a detectable improvement in one or more symptoms associated with OI (e.g., Type I, Type II, Type III, or Type IV OI; or mild, moderate, moderate-severe, or severe OI type), or that causes a biological effect (e.g., a decrease in the level of a particular biomarker) that correlates with an underlying pathological mechanism causing the OI condition or symptom.

[0050] Improved OI may be manifested by reduced bone turnover, reduced rate of bone remodeling and / or reduced bone cell density. In some embodiments, improved OI is indicated by improved bone parameters selected from the group consisting of bone mineral density (BMD), bone volume density (BV / TV), total bone surface (BS), bone surface density (BS / BV), trabecular number (Tb.N), trabecular thickness (Tb.Th), trabecular spacing (Tb.Sp) and total volume (Dens TV).

[0051] In certain embodiments, the improved bone parameter is lumbar spine areal BMD (LS aBMD) as determined by dual energy X-ray absorptiometry. Compared to pre-treatment baseline levels, LS aBMD values ​​may increase by at least 1%, such as at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20% or more.

[0052] In some embodiments, BMD, bone mass and / or bone strength is increased by about 5% to about 200% following treatment with a therapeutically effective amount of an anti-TGF-β antibody or anti-TGF-β fragment. In certain embodiments, the BMD, bone mass and / or bone strength is reduced or improved by about 2%, about 2.1%, about 2.2%, about 2.3%, about 2.4%, about 2.5%, about 2.6%, about 2.7%, about 2.8%, about 2.9%, about 3%, about 4%, about 5% to about 10%, 10% to about 15%, 15% to about 20%, 20% to about 25%, 25% to about 30%, 30% to about 35%, 35% to about 40%, 40% to about 45%, 45% to about 50%, 50% to about 55%, 55% to about 60%, 60% to about 65%, 65% to about 70%, 70% to about 75%, 75% to about 80%, 80% to about 85%, 85% to about 90%, or 95% to about 100% after treatment. increase by 0%, 90% to about 95%, 95% to about 100%, 100% to about 105%, 105% to about 110%, 110% to about 115%, 115% to about 120%, 120% to about 125%, 125% to about 130%, 130% to about 135%, 135% to about 140%, 140% to about 145%, 145% to about 150%, 150% to about 155%, 155% to about 160%, 160% to about 165%, 165% to about 170%, 170% to about 175%, 175% to about 180%, 180% to about 185%, 185% to about 190%, 190% to about 195%, or 195% to about 200%.

[0053] In some embodiments, a therapeutically effective amount may result in decreased bone turnover, as indicated by a decrease in serum or urinary biomarkers such as, for example, urinary hydroxyproline, urinary total pyridinoline (PYD), urinary free deoxypyridinoline (DPD), urinary type I collagen cross-linked N-telopeptide (NTX), urinary or serum type I collagen cross-linked C-terminal telopeptide (CTX), bone sialoprotein (BSP), osteopontin (OPN), and tartrate-resistant acid phosphatase 5b (TRAP). In certain embodiments, the decrease compared to baseline levels (e.g., before treatment) is about 5% to about 200% after treatment with an antibody that binds TGF-β. For example, the reduction may be from about 5% to about 10%, 10% to about 15%, 15% to about 20%, 20% to about 25%, 25% to about 30%, 30% to about 35%, 35% to about 40%, 40% to about 45%, 45% to about 50%, 50% to about 55%, 55% to about 60%, 60% to about 65%, 65% to about 70%, 70% to about 75%, 75% to about 80%, 80% to about 85%, 85% to about 90%, 90% to about 95%, 95% to about 100%, 100% to about 105%, 105% to about 110% after treatment. , 110% to about 115%, 115% to about 120%, 120% to about 125%, 125% to about 130%, 130% to about 135%, 135% to about 140%, 140% to about 145%, 145% to about 150%, 150% to about 155%, 155% to about 160%, 160% to about 165%, 165% to about 170%, 170% to about 175%, 175% to about 180%, 180% to about 185%, 185% to about 190%, 190% to about 195%, or 195% to about 200%.

[0054] In some embodiments, a therapeutically effective amount may result in an increase in the levels of serum or urinary biomarkers of bone deposition, such as total alkaline phosphatase, bone specific alkaline phosphatase, osteocalcin (OCN), and type I procollagen (C-terminal / N-terminal). In certain embodiments, the increase compared to baseline levels (e.g., before treatment) is about 5% to about 200% after treatment. For example, the decrease may be about 5% to about 10%, 10% to about 15%, 15% to about 20%, 20% to about 25%, 25% to about 30%, 30% to about 35%, 35% to about 40%, 40% to about 45%, 45% to about 50%, 50% to about 55%, 55% to about 60%, 60% to about 65%, 65% to about 70%, 70% to about 75%, 75% to about 80%, 80% to about 85%, 85% to about 90%, 90% to about 95%, 95% to about 100%, 100% to about 105%, 105% to about 110% after treatment. , 110% to about 115%, 115% to about 120%, 120% to about 125%, 125% to about 130%, 130% to about 135%, 135% to about 140%, 140% to about 145%, 145% to about 150%, 150% to about 155%, 155% to about 160%, 160% to about 165%, 165% to about 170%, 170% to about 175%, 175% to about 180%, 180% to about 185%, 185% to about 190%, 190% to about 195%, or 195% to about 200%.

[0055] In some embodiments, the therapeutically effective amount promotes bone deposition, hi some embodiments, the therapeutically effective amount improves the function of non-skeletal organs affected by OI, such as hearing, vision, pulmonary function, and renal function.

[0056] In some embodiments, treatment with anti-TGF-β antibodies may be repeated monthly, every 2 months, every 3 months, every 4 months, every 5 months, every 6 months, every 9 months, every 12 months, or every 18 months. In some embodiments, the therapeutically effective amount of Ab1 may be 1-10 mg / kg, for example 1, 2, 2.5, 3, 4, 5, 6, 7, 8, 9, or 10 mg / kg, optionally administered twice a year (every other year, optionally every 6 months or Q6M). In other embodiments, the therapeutically effective amount of Ab1 may be 0.1-1 mg / kg, for example 0.35, 0.4, or 0.5 mg / kg, optionally administered Q3M. In some embodiments, OI patients are treated with this amount of Ab1 by intravenous injection. Treatment may be repeated at intervals that the physician considers appropriate for the patient.

[0057] The patient can be an adult (e.g., a patient aged 18 years or older). The patient can be a pediatric patient (a patient under 18 years of age, e.g., a patient between newborn and 6 years of age, a patient between 6 and 12 years of age, or a patient between 12 and 18 years of age).

[0058] IV. Combination Therapy In some embodiments, the anti-TGF-β antibody therapy may be combined with other OI treatments. Examples of additional therapeutic agents include, but are not limited to, bisphosphonates, calcitonin, teriparatide, and any other compounds known to treat, prevent, or ameliorate OI. Additional therapeutic agents may be administered simultaneously or sequentially with the antibody that binds to TGF-β. Examples of bisphosphonates are etidronate, clodronate, tiludronate, pamidronate, neridronate, olpadronate, alendronate, ibandronate, zoledronate, and risedronate. In some embodiments, additional therapeutic agents are drugs that stimulate bone formation, such as parathyroid hormone analogs and calcitonin.

[0059] Unless otherwise defined herein, scientific and technical terms used in connection with this disclosure shall have the meanings commonly understood by those of ordinary skill in the art. Exemplary methods and materials are described below, however, methods and materials similar or equivalent to those described herein may also be used in the practice or testing of this disclosure. In the event of a conflict, the present specification, including definitions, will control. In general, the nomenclature used in connection with and techniques of cell and tissue culture, molecular biology, immunology, and protein and nucleic acid chemistry and hybridization described herein are those well known and commonly used in the art. Enzymatic reactions and purification techniques are performed according to manufacturer's specifications as commonly accomplished in the art or as described herein. Furthermore, unless otherwise required by context, singular terms shall include the plural and plural terms shall include the singular. Throughout the specification and embodiments, the terms "have" and "comprise" or variations such as "has," "having," "comprises," or "comprises" are understood to mean the inclusion of a recited integer or group of integers, but not the exclusion of any other integer or group of integers. All publications and other references mentioned herein are incorporated by reference in their entirety. Although a number of documents are cited herein, this citation does not constitute an admission that any of these documents form part of the common general knowledge in the art.

[0060] In order that the present invention may be better understood, the following examples are set forth, which are for illustrative purposes only and should not be construed as limiting the scope of the invention in any way. EXAMPLES

[0061] Example 1: A multi-model approach to evaluate anti-TGF-β antibodies for the treatment of osteogenesis imperfecta This example describes a study that characterized the concentration-response relationship of anti-TGF-β antibody Ab1 and its effect on bone mineral density (BMD) and bone strength in patients with OI. The study utilized a model-based approach informed by preclinical and clinical pharmacokinetic (PK) and pharmacodynamic (PD) data. Specifically, nonclinical PK / PD modeling was performed with 1D11, and clinical PK / PD modeling was performed using data obtained from cancer and OI patients treated with fresolimumab (GC1008) or Ab1 during first-in-human studies. 1D11 was used as a rodent surrogate and is a pan-neutralizing TGF-β mouse monoclonal antibody that binds with high affinity and neutralizes the biological activity of all three isoforms of TGF-β. Fresolimumab (GC1008) is a human anti-TGF-β monoclonal antibody that neutralizes all isoforms of TGF-β. Ab1 (GC2008) is a second generation human anti-TGF-β with high sequence similarity to fresolimumab (GC1008), differing only in a single amino acid in the heavy chain (S228P; Eu numbering). 1D11, fresolimumab (GC1008) and Ab1 represent molecules with identical modes of action that differ only in their PK properties.

[0062] method To understand the dose-response relationship, we developed three modeling approaches: 1) a PK / PD approach based on PK and BMD clinical data from OI patients (Figure 1A); 2) a PK / PD approach based on OI mouse pharmacology studies (Figure 1B); 3) a physiologically based pharmacokinetic (PBPK) model approach to predict doses that will reduce OI TGF-β levels in bone to homeostatic levels (Figure 1C).

[0063] In the first modeling approach, PK / PD (BMD) relationships were established from GC1008; then, PK data from Ab1 (GC2008) was used along with GC1008 PD-related parameters to provide dose predictions. In the second modeling approach, PK / PD (BV / TV, maxF) relationships were established from mouse 1D11 data. After scaling the PK, PD parameters were informed based on human bone turnover rate, and the model was used to provide dose predictions. In the third modeling approach, a PBPK model was informed based on the drug's physicochemical (PC) properties and human physiology. After validating the predictions of the PBPK model by comparison with Ab1 PK data, the PBPK model was used to assess bone PK and relevant target (TGF-β) profiles. The multi-model approach is described in more detail below.

[0064] Exposure and PK assessment of fresolimumab (GC1008) in humans The PK of a single-dose infusion of fresolimumab was evaluated during an open-label, dose-ranging, first-in-human study conducted in patients with biopsy-confirmed, treatment-resistant primary focal segmental glomerulosclerosis (FSGS). Sixteen patients received one of four single dose levels of fresolimumab (0.3, 1, 2, 4 mg / kg) and were followed for 112 days with extensive sampling PK. The mean age of the patients was 37 ± 12 years, the mean FSGS duration was 3.0 ± 2.1 years, half were male, 13 were white, and three were black (Trachtman et al., Kidney Intern. (2011) 79 (11): 1236-43). The serum PK of fresolimumab was best described by a two-compartment model with linear clearance (Trachtman et al., ibid). Patient weight was the only significant covariate identified as predictive of pharmacokinetic variability. The half-life was estimated to be 14 days, the mean dose normalized Cmax, and the exposure (AUC) did not change with dose. The PK model parameters are shown in Table 1.

[0065] [Table 1]

[0066] Phase 1 study of fresolimumab in patients with OI A phase 1 study of a single dose of fresolimumab was conducted in eight adults with OI. The study included a single infusion of fresolimumab (GC1008) (1 mg / kg body weight and 4 mg / kg body weight; n=4 in each dose cohort). The primary outcome of the study was the safety of a single dose of fresolimumab (GC1008), while the effect of fresolimumab on bone remodeling biomarkers and lumbar spine areal bone mineral density (LS aBMD) was analyzed as secondary outcomes over a 6-month time frame (Song et al., J Clin Invest. (2022) Feb 3:e152571. doi:10.1172 / JCI152571.

[0067] Phase I study and PK evaluation of Ab1 The PK of Ab1 was evaluated during an open-label, dose-escalation and expansion first-in-human study (NCT03192345) in cancer patients treated with Ab1 alone (Part A) or in combination with cemiplimab (Part B). A total of 52 patients received a 30-minute IV infusion of Ab1 at 0.05 to 15 mg / kg every 2 weeks (Q2W) or 22.5 mg / kg every 3 weeks (Q3W). Blood samples for Ab1 measurement in serum were collected in all treated patients at the start and end of drug infusion, 2.5, 4.5, 8.5 hours on day 1, days 2, 3, 4, 5, 8, and 15 (for Q2W) or 22 (for Q3W) of cycle 1 (i.e., 1 cycle = 1 dose), days 1 and 8 of cycle 2, and day 1 of subsequent cycles (Williamson et al., Developmental Therapeutics-Immunotherapy (2021) 39(15_suppl):2510). The PK of Ab1 was similar to fresolimumab and was described by a two-compartment model with linear clearance (Figure 6). The PK model parameters are shown in Table 2.

[0068] [Table 2]

[0069] 1D11 PK study in OI mouse model A single dose of 5 mg / kg 1D11 was administered intraperitoneally to G610C OI mice (female / 6 and male / 6, 8 weeks old) and blood was collected at 4, 48, 168, 360, 528 and 1032 hours post-dose. All samples were processed for serum, placed on dry ice and transferred to below -60°C prior to analysis.

[0070] Circulating drug levels in serum were determined using an enzyme-linked immunosorbent assay (ELISA)-based bioanalytical method. Briefly, G610C mouse serum samples containing 1D11 were diluted in buffer (PBS, 0.05% Tween-20, 0.05% Triton X-100, 0.01% BSA) at 10,000-fold dilution for all samples except the sample from the last time point (1032 hours), which was diluted 1,000-fold. 96-well plates were coated with TGF-β2 and incubated with mouse serum samples, after which 1D11 was captured using a goat anti-mouse horseradish peroxidase (HRP) conjugated (Sigma, A0168 / 095M4759V) detection antibody, and the optical density was read at 450 nm and 570 nm on a Spectramax® plus (Molecular Devices). The absorbance measured at 570 nm (background) was subtracted from the absorbance measured at 450 nm. A standard curve was constructed to obtain serum 1D11 concentrations. The lower limit of detection of the assay was 1.0 μg / ml. The PK response of 1D11 is shown in Figure 7. The PK parameters are shown in Table 3.

[0071] [Table 3]

[0072] In vivo pharmacological study with 1D11 in OI mice Animals were given food (Auto KF 5 K 52; Lab Diet) and water ad libitum, housed in a pathogen-free, climate-controlled facility with a 12-hour light / dark cycle. G610C OI (Stock No. 007248; Jackson Labs (hereafter referred to as OI mice)) mice carry a mutation in the Col1a2 gene (Col1a2tm1.1Mcbr), which results in low bone mass and a brittle bone phenotype, and therefore represent a good preclinical model for autosomal dominant OI. A dose-response study was performed as follows: 1D11 was administered to male and female G610C OI mice at 0.3, 1 or 5 mg / kg IP for 8 weeks at a dosing frequency of TIW (n=4-8 males, 5-8 females / group). The pharmacodynamic effects of 1D11 across a range of doses were evaluated. A dose-frequency study was evaluated at a dose of 5 mg / kg 1D11 for a total of 12 weeks, administered IP three times per week, once per week, once every two weeks, or once every four weeks (n=5-8 for both uCT and biomechanics) (Greene, B., et al., JBMR Plus, (2021) 5(9):e10530).

[0073] Development of PK / PD models for fresolimumab and 1D11 The stepwise approach followed in this work is shown in Figure 1A-1C. First, a PK / PD model was developed to evaluate the PK / BMD relationship of fresolimumab (GC1008) using a population PK model of fresolimumab performed in a previous study (Trachtman et al., supra) (Figure 1A). BMD kinetics were described by a type III indirect response model simulating the PK-related increase in BMD by induction of the input rate of the effect compartment (Dayneka et al., J Pharmacokinet Biopharm. (1993) 21(4):457-78). After informing the PD-related parameters with available data (Song et al., supra), fresolimumab PK was replaced by the two-compartment PK model of GC2008 (Williamson et al., supra) and this model was used to provide a prediction of Ab1 dose / response (BMD) relationship based on the PK / PD relationship informed by fresolimumab. Population PK analyses of fresolimumab and Ab1 were performed using NONMEM and Monolix, respectively (Bauer et al., CPT: Pharmacometrics & Systems Pharmacology (2019) 8(8):525-537). PK / PD modeling was performed in Matlab R2019a using the ode45 solver for ordinary differential equations.

[0074] In the second modeling approach, PK / PD relationships were established for 1D11 in mice (Figure 1B). The measured PD endpoints were bone volume fraction (bone volume / total volume-BV / TV) and maximum fracture force (maxF), both of which represented improvements in bone physiology. 1D11 PK was described by a one-compartment model (Figure 7, Table 3), and the kinetics of BV / TV and maxF were described by a type III indirect response model. To translate the PK / PD relationships to humans, the Ab1 pop-PK model was used, substituting mouse bone turnover rate (~3 weeks) with human bone turnover rate (~3 months) while keeping PD-related parameters constant. The model was then used to predict Ab1 dose / response (BV / TV) relationships. PK / PD modeling of 1D11 and its forward translation to humans was performed in Matlab R2019a using the ode45 solver for ordinary differential equations.

[0075] PBPK model of Ab1 Finally, a PBPK modeling approach was used to evaluate Ab1 PK in bone and the corresponding TGF-β response in humans. Based on the physicochemical properties of Ab1, TGF-β levels in plasma and bone, and human physiology, a PBPK model was developed using the PK-Sim® software platform (Willmann et al., BIOSILICO (2003) 1(4):121-1240). To validate the PBPK predictions, Ab1 clinical PK data were compared with PBPK simulations for the following scenarios. After validation, the PBPK model was used to evaluate the PBPK / TGF-β response in human plasma and bone tissue. The PK parameters are shown in Table 4.

[0076] [Table 4]

[0077] result First modeling approach The PK of fresolimumab in serum of focal segmental glomerulosclerosis (FSGS) was evaluated in a previous study (Trachtman et al., supra). The PK / BMD response of fresolimumab in OI patients was investigated by using a type III indirect response model for BMD (Figure 2). In the first modeling approach, the PK / PD model was first informed by fresolimumab (GC1008) PK / BMD data. Figure 2 shows the PK / BMD kinetics along with the respective BMD data for single doses of fresolimumab (GC1008) at 1 and 4 mg / kg in OI patients (Song et al., supra). The parameters of the PD model were fitted to the BMD data following administration of fresolimumab (GC1008) at 1 mg / kg (Figure 2, graph A) and 4 mg / kg (Figure 2, graph B). For BMD, administration of 1 mg / kg only minimally affected the kinetics as shown in Figure 2, graph A. The simulations suggest a more pronounced increase in the early period after IV administration of 4 mg / kg. The number of patients in both dose groups was small, and their BMD values ​​maintained significant variability. The model was able to explain the available data without any problems.

[0078] In the second step of this initial modeling approach, the PK portion of the PK / PD model was further informed by Ab1 PK data while holding the PD parameters constant. The PK of Ab1 was further incorporated into the model based on the previous pop-PK analysis. BMD-related parameters were held constant relative to the parameters of fresolimumab. Figure 3 shows the PK / BMD simulated response of Ab1 when administered IV as 2 mg / kg every 6 months (Figure 3, graph A) and 0.4 mg / kg every 3 months (Figure 3, graph B). The doses shown in Figure 3 are those that result in a 5% increase in BMD. Thus, the PK / BMD model of Ab1 predicts twice-yearly administration of 2 mg / kg every 3 months (Figure 3, graph A) or administration of 0.4 mg / kg every 3 months to increase BMD by 5% (Figure 3, graph B).

[0079] The PK behavior of fresolimumab in FSGS was similar in two other disease populations, namely idiopathic pulmonary fibrosis and advanced malignancies (Morris et al., PLoS One (2014) 9 (3): e90353). The underlying hypothesis in this study is that fresolimumab is expected to have a similar pharmacokinetic profile in OI patients, and therefore the PK parameters obtained from the pop-PK analysis of fresolimumab in cancer patients can be used to describe the PK of fresolimumab (GC1008) in OI patients. As shown in the examples herein, a type III PD model was used to model the PD kinetics after administration of fresolimumab (GC1008). This model represents the drug response resulting from the stimulation of factors that control the production of the response variable (Dayneka et al., supra). Although the BMD data can also be modeled using a type II model, which represents a drug response resulting from inhibition of resolution of the response, in some embodiments, a type III model is preferred, based on the underlying physiology of blocking the mechanisms by which anti-TGF-β treatment ultimately induces BMD (Bonewald et al., Clin Orthop Relat Res. (1990)(250):261-76). Due to the small number of BMD data (four subjects), their sparseness and their high variability (Figure 2, graphs A and B), the E of the PD model max / EC 50The parameters were optimized according to the mean BMD values ​​at each time point, while kin / kout was set based on bone turnover and BMD baseline in humans. The model predicts a minimal effect on BMD after a single dose of 1 mg / kg fresolimumab (GC1008), while a dose of 4 mg / kg induces a stronger effect with a more pronounced increase in BMD in the first 100 days. As shown in the examples herein, the BMD response after Ab1 administration was assumed to follow the same kinetics (same PD model and related parameters) as informed by the PK / BMD fit of fresolimumab. Thus, in some embodiments, using the population-PK derived parameters of Ab1 and the BMD-related parameters identified in the first step of this modeling approach, our model provides predictions regarding the PK / PD response of Ab1 (Figure 3, graphs A and B).

[0080] To date, there have been several studies investigating the effect of various treatments on the BMD of patients with OI. In a clinical trial involving 23 men and 23 premenopausal women with OI, Adami et al. tested the effect of neridronate, an amino-bisphosphonate, when administered every 3 months (Adami et al., supra). Within the first 12 months of treatment, bone mineral density at the spine and hip increased by 3% and 4.3%, respectively, with further increases of 3.91% and 1.49% observed during the second year of follow-up. The magnitude of these changes was considered clinically important based on the relationship between BMD change and reduced fracture risk (Hochberg et al., J Clin Endocrinol Metab. (2002) 87(4):1586-92). In the clinical trial by Orwoll et al., 79 adults with OI were randomized in a 1:1 ratio to receive 20 μg / day teriparatide or placebo subcutaneously. Compared to the placebo group, the treatment group demonstrated an increase in lumbar spine (LS) areal BMD (aBMD) of 6.1% versus 2.8% and total hip aBMD of 2.6% versus -2.4%. Additionally, vertebral BMD (vBMD) and strength improved with treatment but declined with placebo. Overall, the results showed that adults with OI showed increases in hip and spine aBMD, vBMD, and estimated strength. In a retrospective analysis (Kuhn et al., J Musculoskelet Neuronal Interact. (2014) 14 (4): 445-53)), the effect of a new physical therapy approach involving lateral alternating whole-body vibration on motor function was analyzed in 53 children with OI. After 12 months, children showed significant increases in motor function and walking distance, which was accompanied by an increase in aBMD from 0.4357 to 0.48 (approximately 10%) and an increase in headless total body BMD from 0.5382 to 0.5529 (approximately 3%). Finally, in a later study, Kuhn et al. showed that denosumab, a RANK ligand antibody that inhibits osteoclast maturation, resulted in a 19% increase in lumbar spine aBMD in 10 children with OI (Kuhn et al., supra). In conclusion, the current data indicate that a significant improvement in clinical outcome is expected for a 5% increase in BMD.Based on model-based predictions of Ab1 PK / PD, a 5% BMD gain is achieved when 0.4 mg / kg Ab1 is administered once every 3 months (Figure 3, graph A) or 2 mg / kg every 6 months (Figure 3, graph B).

[0081] Second Modeling Approach In the second modeling approach, the PK of 1D11 was evaluated in mice (Figure 7). Mouse 1D11 PK was modeled using 1CM. The PK / PD model was further developed based on bone volume changes. Bone volume fraction changes were further evaluated by a type III IDR model (Benjamin et al., JMBR Plus 5.9(2021):e10530). To use the preclinical model for human prediction, bone turnover rate and bone volume fraction baseline were adjusted with human values ​​(Figure 4). PD parameters were fitted to mouse PD. Figure 4 shows the PK / PD response after intravenous administration of 5 mg / kg 1D11 after various regimens. Comparing the simulations (solid lines) with the experimental observations (symbols), the model was able to adequately describe the observed data. The model further predicts that more frequent administration of the same dose results in a faster time to reach steady state in the PD response. To predict PD response in humans, the PK of Ab1 was used to modify the baseline along with the turnover rate parameters of the PD model to represent human values ​​of bone volume fraction and bone turnover accordingly. Figure 4, graphs E and F, show the model-based PK / PD predictions for 0.5 mg / kg IV dosing once every 3 months and 2.5 mg / kg IV dosing every 6 months in humans, respectively. These doses result in a 5% increase in bone volume fraction. The PK / BV model of Ab1 predicts 2.5 mg / kg twice a year (Figure 4, graph F) or 0.5 mg / kg every 3 months to increase BV by 5% (Figure 4, graph E).

[0082] In the second modeling approach (Figure 1B), the available preclinical data of 1D11 are taken into account. The PK of 1D11 was described using a one-compartment model with linear clearance. Although a two-compartment model explained the PK data equally well, the confidence intervals for the parameters of the second compartment were lower, therefore a one-compartment model was chosen. Following the first modeling approach described previously, a type III indirect response model was used to describe the bone volume fraction changes in mice (Figure 4, graphs A-D). According to the bone turnover and bone volume fraction in mice, k in / k out Set E based on available bone volume fraction data max / EC 50 was optimized. As seen in Figure 4, the 1D11 mouse PK / PD model was able to adequately describe the available data. Of note, bone volume fraction measurements were available for one time point limiting the predictive ability of the model, especially for intermediate time points. To translate the mouse 1D11 PK / PD model to the GC2008 PK / PD in humans, three steps were taken. First, the 1D11 PK model was replaced with the previously evaluated GC2008 PK model. In addition, the mouse bone volume fraction baseline was replaced with literature-based values ​​of bone volume fraction in OI patients (Glorieux et al., J Bone Miner Res. (2000) 15(9): 1650-8; and Glorieux et al., J Bone Miner Res. (2002) 17(1): 30-8), and the mouse bone turnover of approximately 3 weeks was replaced with a value of human bone turnover of approximately 3 months (Jilka, ibid.). Based on these changes, a model was used to evaluate the PK / PD response of GC2008 as shown in (Figure 4, graphs E and F). To achieve a 5% increase in bone volume fraction, the PK / PD model predicts dosing of 0.5 mg / kg every 3 months or 2.5 mg / kg twice a year.

[0083] The third (and final) modeling approach In the final modeling approach, a PBPK model of Ab1 was developed and used to predict the dose required to reduce TGF-β in bone to its physiological levels. The developed PBPK model incorporates the physicochemical properties of Ab1 along with information on TGF-β expression in plasma and bone in healthy and OI patients. Predictions based on the PBPK model of Ab1 PK for multiple doses were in close agreement with available data. Figure 5 shows validation of the PBPK model and its forward predictions. Figure 5, graph A, shows the response of the PBPK model to different doses of Ab1. The solid lines show the model-based predictions and the open circles show the individual clinical PK data for the different doses. Comparison of the simulations with the PK data shows that the PBPK model predicts drug exposure in humans well for scenarios that were not used to train the model. Graph B of Figure 5 shows the distribution of Ab1 in plasma (solid line) and bone (dotted line) for a 0.05 mg / kg IV dose of Ab1. The PBPK model predicts that the concentration in bone is nearly 5% of that in plasma. To simulate the OI scenario, TGF-β expression was increased to represent a 3-fold higher concentration of TGF-β in OI patients. Figure 5, graph C further shows the PK prediction based on the PBPK model for an OI patient, with 0.35 mg / kg and 2.5 mg / kg IV doses of Ab1 administered every 3 and 6 months, respectively. These doses were found to reduce TGF-β levels to physiological values ​​for the administration scheme followed. Graph D of Figure 5 further shows the corresponding TGF-β target levels after 0.35 mg / kg and 2.5 mg / kg IV doses of Ab1 administered every 3 and 6 months. The PBPK model predicts doses of 0.35 mg / kg every 3 months and 2.5 mg / kg every 6 months to reduce TGF-β levels to their homeostatic values ​​(Figure 5).

[0084] In the final modeling effort, a PBPK approach was implemented to predict the effect of Ab1 on the levels of TGF-β in bone. PBPK models are well known to have an optimal mathematical framework in which the distribution of drugs in different tissues is predicted depending on physiologically based mass balance and transport phenomena (Jones and Rowland-Yeo., CPT Pharmacometrics Syst Pharmacol. (2013) 2: e63). The inputs to PBPK can generally be divided into drug-specific and organism-specific parameters. Drug-specific parameters relate to the physicochemical properties of the compound, such as molecular weight, affinity for FcRn, affinity for the target of interest, etc. Organ-specific parameters relate to physiological properties of the body, such as tissue volume and tissue blood flow, which are mainly based on the literature and are incorporated into commonly used model platforms. Given their importance in model-based drug development, there are several commercially available platforms that integrate physiologically-based methodologies, such as Simcyp (certara website), GastroPlus (simulations-plus website), SimBiology (mathworks website) and PK-Sim (open systems-pharmacology website). The PK-Sim platform was used because of the relative ease of incorporating target binding to the tissue of interest. The distribution model used to describe the kinetics of Ab1 was based on a two-pore format and was previously described (Niederalt et al., J Pharmacokinet Pharmacodyn. (2018) 45(2):235-57). Required input parameters were the baseline concentrations of TGF-β in plasma and bone, the binding affinities for TGF-β and FcRn and the molecular weight of Ab1. To evaluate the predictive ability of the model, the simulations were compared to the available PK data of Ab1 (Figure 5, graph A). The model-based predictions could well describe the available data, increasing the confidence in the model predictions, especially since the model was not pre-trained on Ab1 PK data.Although there was no bone Ab1 PK available to compare the predicted distribution, our PBPK predictions align well with the literature (Figure 5, graph B), which shows an average 7% distribution of large molecules to bone (Shah and Betts, MAbs. (2013) 5(2):297-305). After establishing the reliability of the PBPK model predictions, a scenario of increased TGF-β concentration was performed. Based on available literature evidence, OI patients showed nearly three-fold higher concentrations of TGF-β in plasma and bone compared to healthy individuals (Grafe et al., Nat Med. (2014) 20(6):670-5; Gebken et al., Pathobiology (2000) 68(3):106-12; and Pfeilschifter et al., ibid.). Based on this evidence, TGF-β expression was increased in plasma and bone to simulate the OI scenario. Therefore, we sought to evaluate the dose of Ab1 that would return OI-associated free TGF-β levels to their healthy values. The underlying assumption is that changes in body physiology associated with OI (i.e., loss of bone volume) do not affect Ab1 PK and may remain constant. Based on PBPK analysis, administration of 0.35 mg / kg every 3 months or 2.5 mg / kg every 6 months (Figure 5, graph C) returns free TGF-β levels in bone to their physiological values ​​(Figure 5, graph D). Interestingly, PBPK analysis showed that once Ab1 concentrations reach their peak, twice-yearly administration of 2.5 mg / kg nearly eliminates the total amount of free TGF-β in bone. This further reveals possible dosing design constraints, which should be optimized to account for this implication.

[0085] In summary, a multi-model approach was implemented to evaluate the concentration-response relationship of anti-TGF-β antibodies and BMD and bone strength as well as TGF-β kinetics in bone of patients with OI. The three modeling approaches provided similar dose predictions for clinically relevant PD effects. The three modeling approaches implemented in this study provided similar dose estimates for clinically relevant PD effects. Notably, the first approach using fresolimumab, Ab1 clinical PK / PD data, predicted 0.4 mg / kg every 3 months or 2 mg / kg twice a year to increase BMD by 5%. The second approach, which further used preclinical data of 1D11, predicted 0.5 mg / kg every 3 months and 2.5 mg / kg twice a year to increase bone volume fraction by 5%. Finally, PBPK modeling predicted 0.35 mg / kg every 3 months or 2.5 mg / kg twice a year to reduce OI-associated TGF-β levels back to their physiological values. Alignment of the three approaches increased confidence in the translation of Ab1 PK / PD relationships and provided a robust model-based assessment for predicting clinical efficacy.

[0086] The above non-limiting examples are provided for illustrative purposes only to facilitate a more detailed understanding of the disclosed subject matter, and should not be construed as limiting any of the embodiments described herein, including those relating to the antibodies, pharmaceutical compositions, or methods and uses for treating cancer, neurodegenerative, or infectious diseases.

[0087] array The following table shows the amino acid sequences referred to in this disclosure.

[0088] [Table 5]

Claims

1. 1. Use of an anti-TGF-β antibody in the manufacture of a medicament for use in a method of treating osteogenesis imperfecta (OI) in a human subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of an anti-TGF-β antibody; The antibody comprises heavy chain complementarity determining regions (CDRs) 1-3 comprising SEQ ID NOS: 4-6, respectively, and light chain CDRs 1-3 comprising SEQ ID NOS: 7-9, respectively, and the antibody is a human IgG1 with a proline at position 228 (Eu numbering). 4 including a constant region, The antibody 1-8 mg / kg, optionally 2, 2.5, or 5 mg / kg twice a year; or 0.1-1 mg / kg, optionally 0.35, 0.4, or 0.5 mg / kg every 3 months (Q3M) The use, wherein the therapeutically effective amount is administered.

2. The use of claim 1 , wherein the antibody comprises a heavy chain variable domain comprising SEQ ID NO: 10 and a light chain variable domain comprising SEQ ID NO:

11.

3. The antibody is a human IgG 4 The use according to claim 1 , comprising a constant region and / or a human kappa light chain constant region.

4. The use according to claim 3 , wherein the antibody comprises a heavy chain comprising SEQ ID NO: 3 and a light chain comprising SEQ ID NO:

2.

5. The use of claim 1 , wherein the antibody comprises a bone-targeting moiety, and optionally, the bone-targeting moiety is a polyarginine peptide.

6. The use of claim 5 , wherein the antibody comprises one or more polyarginine peptides.

7. The use according to claim 6, wherein the antibody is fused to a polyarginine peptide at the N-terminus or C-terminus or both termini of the heavy chain of the antibody and / or at the C-terminus of the light chain of the antibody.

8. The use according to claim 5, wherein the polyarginine peptide is D10 (SEQ ID NO: 14).

9. The use according to claim 1, wherein the OI is moderate to severe OI or type IV OI.

10. The use according to claim 1 , wherein the OI is type I, type II or type III OI.

11. 2. The use of claim 1, wherein the human subject is an adult patient (age 18 or older) or a pediatric patient (age under 18).

12. 2. The use of claim 1, wherein the human subject has a mutation in the COL1A1 gene or the COL1A2 gene, and optionally the mutation is a glycine substitution mutation in the COL1A1 gene or the COL1A2 gene or a valine deletion in the COL1A2 gene.

13. 2. The use of claim 1, wherein said administration improves a bone parameter selected from the group consisting of bone mineral density (BMD), bone volume density (BV / TV), total bone surface (BS), bone surface density (BS / BV), trabecular number (Tb.N), trabecular thickness (Tb.Th), trabecular spacing (Tb.Sp) and total volume (Dens TV).

14. 2. The use of claim 1, wherein the administration reduces bone turnover and / or bone cell density, and optionally the reduced bone turnover is indicated by a decrease in serum CTX or an increase in serum osteocalcin (OCN).

15. 10. The use of claim 1, wherein the antibody is administered at 2 mg / kg twice yearly or 0.4 mg / kg Q3M, and optionally, the administration results in about a 5% increase in BMD in the subject.

16. 10. The use of claim 1, wherein the antibody is administered at 5 mg / kg twice yearly or 0.5 mg / kg Q3M, and optionally, the administration results in about a 5% increase in BV in the subject.

17. 2. The use of claim 1, wherein the antibody is administered at 2.5 mg / kg twice yearly or 0.35 mg / kg Q3M, and optionally, the administration results in a reduction of TGF-β levels in the subject to homeostatic values.

18. The use of claim 1 , wherein the antibody is administered by intravenous infusion.

19. 2. The use of claim 1, wherein the antibody is administered in combination with a bisphosphonate, parathyroid hormone, calcitonin, teriparatide, or an anti-sclerostin agent.

20. 20. The use of claim 19, wherein the antibody is administered in combination with a bisphosphonate selected from alendronate, pamidronate, zoledronate, and risedronate.

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