Methods for treating myelofibrosis and related conditions

Abstract

This invention provides a method for treating anemia in subjects with myelofibrosis. [Solution] A method is provided for treating anemia in a subject with myelofibrosis, comprising administering an effective amount of a hemoduvelin-induced BMP signaling antagonist, a hepcidin antagonist, to the subject.

Description

[Technical Field]

[0001] Related applications This application claims the benefits of U.S. Provisional Patent Application No. 62 / 907,227, filed on 27 September 2019, titled "Hepcidin Antagonists for the Treatment of Myelofibrosis and Related Conditions," No. 63 / 063,761, filed on 10 August 2020, titled "Methods for the Treatment of Myelofibrosis and Related Conditions," and No. 63 / 072,057, filed on 28 August 2020, both of which are incorporated herein by reference in their entirety under 35 U.S. SC § 119(e). [Background technology]

[0002] Iron is a vital component of oxygen-carrying and storage molecules such as hemoglobin and myoglobin. Iron deficiency leads to anemia, while iron overload leads to tissue damage and fibrosis. Hepcidin is an important peptide hormone regulator of systemic iron homeostasis. It exerts its regulatory function by binding to ferroportin, a transmembrane protein present in hepatocytes, duodenal intestinal cells, macrophages, and adipocytes, which is a cellular iron efflux protein. Hepcidin binding promotes the breakdown of ferroportin, preventing iron efflux from cells and release into the plasma. [Overview of the Initiative]

[0003] Aspects of this disclosure provide methods for treating hyperhepcidin disorders resulting in functional iron deficiency, such as myelofibrosis, myeloma, Wanderström hypergammaglobulinemia, chronic kidney disease, anemia of chronic disease, or iron-restricted anemia. Hepcidin expression in hepatocytes involves two main signaling pathways. Hepcidin expression is regulated by bone morphogenetic protein (BMP) signaling pathways (e.g., the BMP6-induced signaling pathway). Hepcidin expression via the BMP signaling pathway is promoted by the membrane-bound coreceptor, hemoduvelin. Hepcidin expression is also regulated by inflammatory pathways (e.g., the IL-6-mediated JAK-STAT pathway). Conditions involving abnormal fibrotic and / or inflammatory responses can result in high hepcidin levels.

[0004] In certain embodiments, a method is provided for treating myelofibrosis, which is generally characterized as a myeloproliferative disorder associated with chronic inflammation and progressive bone marrow fibrosis. Anemia is a major clinical challenge in myelofibrosis and is associated with a negative outcome. Such anemia is generally caused by or associated with bone marrow failure, splenomegaly, and / or functional iron deficiency, and can contribute to inflammation. Further, in myelofibrosis, pro-inflammatory cytokines such as IL-6 and oncostatin-M that induce hepcidin synthesis are typically increased and are associated with iron sequestration, macrophage iron loading, and bone marrow proliferation and macrophage activation (see, e.g., FIG. 1). The resulting increase in hepcidin levels is associated with anemia and negative outcomes. Thus, aspects of the present disclosure relate to treating a subject having elevated hepcidin levels (e.g., myelofibrosis) by inhibiting the BMP signaling pathway (e.g., the hemojuvelin-induced BMP signaling pathway) and / or the inflammatory response (e.g., the IL-6-mediated JAK-STAT pathway). In some embodiments, the subject is treated with an antagonist of the HJV-induced BMP-6 signaling pathway. In some embodiments, the subject is treated with a JAK-STAT inhibitor (e.g., a JAK2 inhibitor). In some embodiments, the subject is treated with a combination of an antagonist of the HJV-induced BMP signaling pathway and a JAK inhibitor. The combination therapy using an antagonist of the HJV-induced BMP signaling pathway and a JAK inhibitor can significantly improve bone marrow failure and splenomegaly and reduce the risk of anemia caused by elevated hepcidin levels.

[0005] In some aspects, the present disclosure provides a method of treating anemia in a subject having myelofibrosis, the method comprising administering to the subject an effective amount of a hepcidin antagonist. In some embodiments, the subject has an iron utilization disorder / functional iron deficiency.

[0006] In some embodiments, the hepcidin antagonist is a hemojuvelin-induced BMP signaling antagonist. In some embodiments, the hemojuvelin-induced BMP signaling antagonist is a BMP antagonist. In some embodiments, the BMP antagonist is a BMP2, BMP4, BMP5 or BMP6 antagonist. In some embodiments, the BMP antagonist is a BMP6 antagonist.

[0007] In some embodiments, the hemojuvelin-induced BMP signaling antagonist selectively inhibits its target molecule. In some embodiments, the target molecule is a BMP receptor. In some embodiments, the hemojuvelin-induced BMP signaling antagonist selectively inhibits its target molecule as compared to a reference molecule. In some embodiments, the reference molecule is JAK2. In some embodiments, the hemojuvelin-induced BMP signaling antagonist selectively inhibits its target molecule as compared to a reference molecule and has an inhibitory concentration (IC 50 ) of the reference molecule that is at least 10-fold higher (e.g., in the range of 10 1 to 10 6 -fold higher) than the half-maximal inhibitory concentration of the target molecule when measured in a kinase potency assay. 50

[0008] In some embodiments, the hemojuvelin-induced BMP signaling antagonist is sHJV or a soluble hemojuvelin-Fc fusion protein. In some embodiments, the soluble HJV-Fc fusion protein is FMX8.

[0009] In some embodiments, the hemojuvelin-induced BMP signaling antagonist is a BMP6 neutralizing antibody. In some embodiments, the BMP6 neutralizing antibody is LY311359, CSJ137, or KY1070.

[0010] In some embodiments, the hemojuvelin-induced BMP signaling is a modified heparin selected from SST0001, RO-82, RO-68, NAc-91, and NacRO-00.

[0011] In some embodiments, the hemoduvelin-induced BMP signaling antagonist is recombinant SMAD6 or SMAD7.

[0012] In some embodiments, the hepcidin antagonist is a hepcidin neutralizer.

[0013] In some embodiments, the hepcidin neutralizer is NOX-94, a PEGylated L-stereoisomerized RNA aptamer that binds to and neutralizes hepcidin. In some embodiments, the hepcidin neutralizer is PRS-080, an anticharin against hepcidin. In some embodiments, the hepcidin neutralizer is LY2787106, a monoclonal antibody that targets hepcidin.

[0014] In some embodiments, the hemoduverin-induced BMP signaling antagonist is an ALK2 antagonist. In some embodiments, the ALK2 antagonist is INCB000928, KER-047, or BLU-782.

[0015] In some embodiments, the hepcidin antagonist is a hemoduverine antagonist. In some embodiments, the hemoduverine antagonist is an anti-hemoduverine antibody. In some embodiments, the anti-hemoduverine antibody preferentially binds to RGMa and RGMb over RGMac. In some embodiments, the anti-hemoduverine antibody has an equilibrium dissociation constant (K) lower than 100 nM. D It binds to RGMc at ). In some embodiments, the anti-HJV antibody is HJV-35202. In some embodiments, the anti-HJV antibody is the anti-HJV antibody listed in Table 1.

[0016] In some embodiments, the anti-hemoduvelin antibody comprises: (a) a variable heavy chain region comprising CDR1 containing the amino acid sequence of SEQ ID NO: 1, CDR2 containing the amino acid sequence of SEQ ID NO: 2, and CDR3 containing the amino acid sequence of SEQ ID NO: 3; and / or (b) a variable light chain region comprising CDR1 containing the amino acid sequence of SEQ ID NO: 4, CDR2 containing the amino acid sequence of SEQ ID NO: 5, and CDR3 containing the amino acid sequence of SEQ ID NO: 6.

[0017] In some embodiments, the anti-hemoduvelin antibody comprises: (a) a variable heavy chain region comprising CDR1 containing the amino acid sequence of SEQ ID NO: 1, CDR2 containing the amino acid sequence of SEQ ID NO: 2, and CDR3 containing the amino acid sequence of SEQ ID NO: 3; and / or (b) a variable light chain region comprising CDR1 containing the amino acid sequence of SEQ ID NO: 7, CDR2 containing the amino acid sequence of SEQ ID NO: 8, and CDR3 containing the amino acid sequence of SEQ ID NO: 9.

[0018] In some embodiments, the anti-hemoduvelin antibody comprises: (a) a variable heavy chain region comprising CDR1 containing the amino acid sequence of SEQ ID NO: 1, CDR2 containing the amino acid sequence of SEQ ID NO: 2, and CDR3 containing the amino acid sequence of SEQ ID NO: 3; and / or (b) a variable light chain region comprising CDR1 containing the amino acid sequence of SEQ ID NO: 10, CDR2 containing the amino acid sequence of SEQ ID NO: 11, and CDR3 containing the amino acid sequence of SEQ ID NO: 12.

[0019] In some embodiments, the anti-hemoduvelin antibody comprises: (a) a variable heavy chain region comprising CDR1 comprising the amino acid sequence of SEQ ID NO: 1, CDR2 comprising the amino acid sequence of SEQ ID NO: 2, and CDR3 comprising the amino acid sequence of SEQ ID NO: 3; and / or (b) a variable light chain region comprising CDR1 comprising the amino acid sequence of SEQ ID NO: 13, CDR2 comprising the amino acid sequence of SEQ ID NO: 14, and CDR3 comprising the amino acid sequence of SEQ ID NO: 15.

[0020] In some embodiments, the anti-hemoduvelin antibody comprises: (a) a variable heavy chain region comprising CDR1 containing the amino acid sequence of SEQ ID NO: 1, CDR2 containing the amino acid sequence of SEQ ID NO: 2, and CDR3 containing the amino acid sequence of SEQ ID NO: 3; and / or (b) a variable light chain region comprising CDR1 containing the amino acid sequence of SEQ ID NO: 16, CDR2 containing the amino acid sequence of SEQ ID NO: 17, and CDR3 containing the amino acid sequence of SEQ ID NO: 18.

[0021] In some embodiments, the anti-hemoduvelin antibody comprises: (a) a variable heavy chain region comprising CDR1 comprising the amino acid sequence of SEQ ID NO: 19, CDR2 comprising the amino acid sequence of SEQ ID NO: 20, and CDR3 comprising the amino acid sequence of SEQ ID NO: 21; and / or (b) a variable light chain region comprising CDR1 comprising the amino acid sequence of SEQ ID NO: 22, CDR2 comprising the amino acid sequence of SEQ ID NO: 23, and CDR3 comprising the amino acid sequence of SEQ ID NO: 24.

[0022] In some embodiments, subjects have myelofibrosis-inducing mutations in JAK2, LNK, PPM1D, MPL, ASXL1, TET2, NFE2, SH2B3, SF3B1, or CALR. In some embodiments, subjects have mutations in genes involved in epigenetic regulation or splicing, namely ASXL1, DNMT3A, TET2, SRSF2, U2AF1, EZH2, or SF3B1. In some embodiments, subjects have mutations in IDH1 / 2, which are associated with a risk of progression to MBN-BP. In some embodiments, subjects contain a human JAK2 gene with an inducement mutation in exon 12 or exon 14. In some embodiments, the inducement mutation in the JAK2 gene is in exon 14, resulting in a V617F substitution. In some embodiments, myelofibrosis is associated with increased levels of pro-inflammatory cytokines (e.g., IL-6, oncostatin-M) in the subjects.

[0023] In some embodiments, subjects have or are at risk of having systemic or microvascular symptoms associated with MPN. In some embodiments, subjects have or are at risk of having thromboembolic or hemorrhagic complications. In some embodiments, subjects have or are at risk of having MPN-acute transformation acute myeloid leukemia (AML). In some embodiments, subjects exhibit ribosomal disease in megakaryocytes. In some embodiments, subjects exhibit decreased GATA1 expression, particularly in megakaryocytes. In some embodiments, subjects exhibit deficiencies in megakaryocyte function or maturation.

[0024] In some embodiments, the subjects do not have nutritional iron deficiency. In some embodiments, the subjects have ferritin levels greater than 100 μg / L. In some embodiments, the subjects have a reticulocyte hemoglobin content less than 26 pg / cell. In some embodiments, the subjects have a transferrin saturation level lower than 50%. In some embodiments, the subjects have liver iron levels higher than 2000 μg / g dry weight. In some embodiments, the subjects have serum iron levels in the range of less than 50 μg / dL. In some embodiments, the subjects have total iron-binding capacity in the range of less than 400 μg / dL. In some embodiments, the subjects have hepcidin levels in the range of more than 55 ng / ml. In some embodiments, the subjects have IL-6 levels higher than 1.8 pg / mL. In some embodiments, the subjects have serum creatinine levels higher than 2 mg / dL. In some embodiments, subjects are identified as having hemoglobin levels in the range of 1.5–2.0 g / dL or 2.0–4.0 g / dL, or hemoglobin levels below normal. In some embodiments, subjects exhibit serum hemoglobin levels lower than 10 g / dL. In some embodiments, subjects exhibit serum hemoglobin levels lower than 8 g / dL.

[0025] In some embodiments, the subjects exhibit thrombocytopenia, anemia, and / or neutropenia. In some embodiments, the subjects have received one or more blood transfusions. In some embodiments, the subjects have transfusion-dependent anemia. In some embodiments, the subjects have received multiple blood transfusions over a 12-week period.

[0026] In some embodiments, subjects had previously received one or more doses of a JAK / STAT antagonist as treatment for Philadelphia chromosome-negative myeloproliferative neoplasm (MPN). In some embodiments, subjects received a JAK / STAT antagonist as treatment for polycythemia vera (PV), essential thrombocythemia (ET), or pre-fibrotic / early primary myelofibrosis (pre-MF). In some embodiments, subjects received a JAK / STAT antagonist as treatment for myeloid fibrosis. In some embodiments, subjects were treated with a JAK / STAT antagonist for 2 to 6 weeks. In some embodiments, the JAK / STAT antagonist is selective for JAK1 or JAK2. In some embodiments, the JAK / STAT antagonist is not active against ACVR1 / ALK2. In some embodiments, the JAK / STAT antagonist is ruxolitinib, fedratinib, pacritinib, baricitinib, tofacitinib, oclacitinib, or NSC13626. In some embodiments, the JAK / STAT antagonist inhibits IL-6-mediated STAT3 activation. In some embodiments, the JAK / STAT antagonist is GS-0387 or CYT-387.

[0027] In some embodiments, the method further includes administering one or more additional therapeutic agents to the target. In some embodiments, the additional therapeutic agents are selected from GDF traps, bromodomain and excess terminal domain (BET) inhibitors, erythropoiesis stimulants, or immunomodulators / erythropoietin stimulants. In some embodiments, the GDF trap is sotatercept, raspatercept, or KER-050. In some embodiments, the BET inhibitor is CPI-0610. In some embodiments, the immunomodulator / erythropoietin stimulant is pomalidomide. In some embodiments, the erythropoiesis stimulant is erythropoietin (EPO).

[0028] In some embodiments, the Disclosure provides a method for treating anemia in a subject having myelofibrosis, comprising administering an effective dose of a hepcidin antagonist and one or more additional therapeutic agents to the subject.

[0029] In some embodiments, the hepcidin antagonist is an HJV-inducing BMP signaling antagonist or a hepcidin neutralizer. In some embodiments, the HJV-inducing BMP signaling antagonist is a BMP antagonist, an HJV antagonist, a modified heparin that targets BMP6, or recombinant SMAD6 or SMAD7. In some embodiments, the BMP antagonist is a BMP6 neutralizing antibody selected from LY311359, CSJ137, and KY1070. In some embodiments, the HJV-inducing BMP signaling antagonist is an HJV antagonist. In some embodiments, the HJV antagonist is an anti-HJV antibody. In some embodiments, an additional therapeutic agent is selected from a GDF trap, a JAK / STAT inhibitor, a BET inhibitor, an erythropoiesis stimulant, or an immunomodulator / erythropoietin stimulant. In some embodiments, the additional therapeutic agent is a GDF trap. In some embodiments, the GDF trap is sotatercept, raspatercept, or KER-050. In some embodiments, the JAK / STAT inhibitor is momenotinib. In some embodiments, the BET inhibitor is CPI-0610. In some embodiments, the immunomodulator / erythropoietin stimulant is pomalidomide. In some embodiments, the erythropoiesis stimulant is EPO.

[0030] In certain embodiments, the Disclosure provides a method for treating subjects who have or are at risk of having an adverse reaction to a JAK-STAT antagonist, the method comprising administering an effective dose of a hemoduverin-induced BMP signaling antagonist to the subject.

[0031] Certain embodiments of this disclosure relate to the observation that hemoduvelin (HJV) is a regulatory agent of hepcidin synthesis and that a lack of hemoduvelin function may be associated with iron overload. For example, in some embodiments, homozygous HJV knockdown animals are unable to amplify hepcidin synthesis in response to IL-6 and are unable to initiate an effective hypoirremia response to acute inflammation. Thus, in some embodiments, the method provided herein comprises administering an effective amount of a hepcidin antagonist to a subject in need to treat a hyperhepcidin disorder, which may be a hemoduvelin antagonist. In some embodiments, the hemoduvelin antagonist is an anti-hemoduvelin antibody. In some embodiments, the anti-hemoduvelin antibody binds to RGMc as its primary mode of action (compared to RGMa and RGMb). Thus, in some embodiments, the anti-hemoduvelin antibody preferentially binds to RGMc compared to RGMa and / or RGMb. In some embodiments, the anti-hemoduvelin antibody binds to RGMc at concentrations lower than 100 nanomoles (nM) (K D <100nM) equilibrium dissociation constant (K D ) binds to RGMc. However, in some embodiments, the anti-hemoduvelin antibody binds to RGMc with similar affinity to RGMa and / or RGMb.

[0032] In some embodiments, the subjects treated in accordance with this disclosure are transfusion-dependent. In some embodiments, the subjects treated are transfusion-independent. In some embodiments, the subjects treated occasionally receive transfusions but are not classified as transfusion-dependent.

[0033] In some embodiments, the subject has previously received an erythropoietin stimulant, a JAK-STAT inhibitor, a growth factor ligand trap, or an antifibrotic agent. In some embodiments, the erythropoietin stimulant is selected from the group consisting of danazol, prednisone, thalidomide, lenalidomide, and pomalidomide. In some embodiments, the JAK-STAT inhibitor is selected from the group consisting of ruxolitinib, momerotinib, pacritinib, INCB039110, AG490, and PpYLKTK. In some embodiments, the growth factor ligand trap is sotatercept and raspatercept. In some embodiments, the antifibrotic agent is PRM-151.

[0034] In some embodiments, the method of treating the subject further includes administering one or more erythropoietin stimulants, JAK-STAT inhibitors, growth factor ligand traps, and antifibrotic agents to the subject. In some embodiments, the erythropoietin stimulant is selected from the group consisting of danazol, prednisone, thalidomide, lenalidomide, and pomalidomide. In some embodiments, the JAK-STAT inhibitor is selected from the group consisting of ruxolitinib, momerotinib, pacritinib, INCB039110, AG490, and PpYLKTK. In some embodiments, the growth factor ligand trap is sotatercept. In some embodiments, the antifibrotic agent is PRM-151.

[0035] In some embodiments, the anti-hemoduvelin antibody is an affinity-mature antibody. In some embodiments, the affinity-mature antibody is derived from a mouse monoclonal antibody. In some embodiments, the anti-hemoduvelin antibody is a humanized antibody. In some embodiments, the anti-hemoduvelin antibody comprises at least three complementarity-determining regions (CDRs) transplanted into a heterologous framework. In some embodiments, the heterologous framework comprises a human framework region, and at least three CDRs comprise non-human CDRs. In some embodiments, the non-human CDRs are derived from rodents. In some embodiments, at least three CDRs comprise variable light chain CDRs. In some embodiments, at least three CDRs comprise three variable heavy chain CDRs and three variable light chain CDRs.

[0036] The aforementioned and other embodiments, implementations, actions, functions, features, and embodiments of this teaching can be better understood from the following description in conjunction with the accompanying drawings. [Brief explanation of the drawing]

[0037] The accompanying drawings incorporated herein and constituting part of this specification illustrate certain embodiments and, together with the description provided, provide non-limiting examples of certain aspects of the compositions and methods disclosed herein. [Figure 1] Figure 1 shows the bone marrow proliferation cycle specific to a particular hyperhepcidin disorder. [Figure 2] Figure 2 shows the hepcidin-stimulating pathway and the physiological regulation of iron homeostasis by hepcidin. [Figure 3A] Figures 3A-3G illustrate the role of hepcidin in functional iron deficiency (FID) and examples of controlling hepcidin levels with hepcidin antagonists. Figure 3A shows the mechanism of functional iron deficiency. [Figure 3B]Figures 3A–3G illustrate the role of hepcidin in functional iron deficiency (FID) and examples of controlling hepcidin levels with hepcidin antagonists. Figure 3B shows that functional iron deficiency is a common feature of anemia in inflammatory and chronic diseases, including myelofibrosis (MF), chronic kidney disease (CKD), cancer, and heart failure. [Figure 3C] Figures 3A-3G illustrate the role of hepcidin in functional iron deficiency (FID) and examples of controlling hepcidin levels with hepcidin antagonists. Figure 3C shows that functional iron deficiency is associated with high iron and high hepcidin levels. [Figure 3D] Figures 3A–3G illustrate the role of hepcidin in functional iron deficiency (FID) and an example of controlling hepcidin levels with a hepcidin antagonist. Figure 3D is a schematic diagram showing how hepcidin levels are reduced to normal levels by using a hepcidin antagonist to treat iron restriction disorders. [Figure 3E] Figures 3A-3G illustrate the role of hepcidin in functional iron deficiency (FID) and examples of controlling hepcidin levels with hepcidin antagonists. Figure 3E shows the use of an anti-HJV antibody as an example of inhibiting the HJV-induced BMP signaling pathway and reducing hepcidin to normal levels. [Figure 3F] Figures 3A-3G illustrate the role of hepcidin in functional iron deficiency (FID) and examples of hepcidin levels being controlled by hepcidin antagonists. Figure 3F shows that matryptase 2 negatively regulates hepcidin by cleaving membrane-bound HJV. [Figure 3G] Figures 3A–3G illustrate the role of hepcidin in functional iron deficiency (FID) and examples of controlling hepcidin levels with hepcidin antagonists. Figure 3G shows examples of hepcidin antagonists that may be used to control hepcidin levels. [Figure 4] Figure 4 shows that activin B regulates hepcidin levels in response to inflammation, via both HJV-induced BMP signaling. [Figure 5]Figure 5 shows the current treatment plan for myelofibrosis based on disease severity. [Figure 6] Figure 6 is a graph showing that IL-6 induces hepcidin expression in cynomolgus macaques, and anti-HJV antibody treatment prevents dose-dependent inflammatory (IL-6) hepcidin increase in cynomolgus macaques. [Modes for carrying out the invention]

[0038] In certain embodiments, the Disclosure provides hepcidin antagonists for targeting hepcidin that are effective in inhibiting hepcidin function and / or reducing hepcidin expression in cells, particularly for modulating iron homeostasis for the treatment of myelofibrosis and / or one or more symptoms or complications thereof. Accordingly, in relevant embodiments, the Disclosure provides compositions and methods for treating myelofibrosis, including primary myelofibrosis, myelofibrosis arising from myeloproliferative neoplasms and / or myelofibrosis-associated anemia, inflammation, bone marrow failure, splenomegaly, catabolism, and / or fatigue, one or more symptoms or complications thereof.

[0039] Further aspects of this disclosure, including explanations of the terms defined, are provided below.

[0040] I. Definition Administration: As used herein, the term “administering” or “dosing” means providing a complex to a subject in a physiologically and / or pharmacologically useful manner (e.g., treating a condition in a subject).

[0041] Antibody: As used herein, the term "antibody" refers to a polypeptide that includes at least one immunoglobulin variable domain or at least one antigen determinant, e.g., a paratope that specifically binds to an antigen. In some embodiments, the antibody is a full-length antibody. In some embodiments, the antibody is a chimeric antibody. In some embodiments, the antibody is a humanized antibody. However, in some embodiments, the antibody is a Fab fragment, F(ab’)2 fragment, Fv fragment or scFv fragment. In some embodiments, the antibody is a nanobody derived from a camelid antibody or a nanobody derived from a shark antibody. In some embodiments, the antibody is a bispecific antibody. In some embodiments, the antibody includes a framework having human germline sequences. In another embodiment, the antibody includes a heavy chain constant domain selected from the group consisting of IgG, IgG1, IgG2, IgG2A, IgG2B, IgG2C, IgG3, IgG4, IgA1, IgA2, IgD, IgM, and IgE constant domains. In some embodiments, the antibody includes a heavy (H) chain variable region (abbreviated herein as V H ), and / or a light (L) chain variable region (abbreviated herein as V L ). In some embodiments, the antibody includes a constant domain, e.g., an Fc region. The immunoglobulin constant domain refers to a heavy or light chain constant domain. The amino acid sequences of human IgG heavy and light chain constant domains and their functional variations are known. With respect to the heavy chain, in some embodiments, the heavy chain of the antibody described herein can be an alpha (α), delta (Δ), epsilon (ε), gamma (γ), or mu (μ) heavy chain. In some embodiments, the heavy chain of the antibody described herein can include a human alpha (α), delta (Δ), epsilon (ε), gamma (γ), or mu (μ) heavy chain. In certain embodiments, the antibody described herein includes a human gamma1 CH1, CH2, and / or CH3 domain. In some embodiments, V HThe amino acid sequence of the domain includes the amino acid sequence of the human gamma (γ) heavy chain constant region, for example, any known in the art. Non-limiting examples of human constant region sequences are described in the art; see, for example, U.S. Patent No. 5,693,780 and Kabat EA et al., (1991) above. In some embodiments, V HThe domains include an amino acid sequence that is at least 70%, 75%, 80%, 85%, 90%, 95%, 98%, or at least 99% identical to any of the variable chain constant regions provided herein. In some embodiments, the antibody is modified, for example by glycosylation, phosphorylation, SUMOylation, and / or methylation. In some embodiments, the antibody is a glycosylated antibody conjugated to one or more sugar or carbohydrate molecules. In some embodiments, one or more sugar or carbohydrate molecules are conjugated to the antibody by N-glycosylation, O-glycosylation, C-glycosylation, glyciation (GPI anchoring), and / or phosphoglycosylation. In some embodiments, one or more sugar or carbohydrate molecules are monosaccharides, disaccharides, oligosaccharides, or glycans. In some embodiments, one or more sugar or carbohydrate molecules are branched oligosaccharides or branched glycans. In some embodiments, one or more sugar or carbohydrate molecules include mannose units, glucose units, N-acetylglucosamine units, or phospholipid units. In some embodiments, the antibody is a construct comprising a polypeptide containing one or more antigen-binding fragments of the present disclosure linked to a linker polypeptide or an immunoglobulin constant domain. The linker polypeptide contains two or more amino acid residues joined by a peptide bond and is used to link one or more antigen-binding moieties. Examples of linker polypeptides have been reported (see, for example, Holliger, P., et al. (1993) Proc. Natl. Acad. Sci. USA 90:6444-6448; Poljak, RJ, et al. (1994) Structure 2:1121-1123). Furthermore, the antibody may be part of a larger immunoadhesion molecule formed by covalent or noncovalent bonding of the antibody or antibody moiety with one or more other proteins or peptides.Examples of such immunoadhesion molecules include the use of streptavidin core regions to create tetrameric scFv molecules (Kipriyanov, SM, et al. (1995) Human Antibodies and Hybridomas 6:93-101) and the use of cysteine ​​residues, marker peptides, and C-terminal polyhistidine tags to create divalent and biotinylated scFv molecules (Kipriyanov, SM, et al. (1994) Mol. Immunol. 31:1047-1058).

[0042] Affinity-matured antibody: “Affinity-matured antibody” as used herein refers to an antibody having one or more modifications to one or more CDRs, which results in an improvement in the antibody’s affinity (i.e., KD, kd, or ka) to a target antigen compared to a parent antibody without modifications. Exemplary affinity-matured antibodies may have nanomolar or even picomolar affinity to a target antigen. Various procedures for producing affinity-matured antibodies are known in the art, including screening of combinatory antibody libraries prepared using biodisplay. For example, Marks et al., BioTechnology, 10: 779-783 (1992) describes affinity maturation by VH and VL domain shuffling. Random mutagenesis of CDRs and / or framework residues is described by Barbas et al., Proc. Nat. Acad. Sci. USA, 91: 3809-3813 (1994); Schier et al., Gene, 169: 147-155 (1995); Yelton et al., J. Immunol., 155: 1994-2004 (1995); Jackson et al., J. Immunol., 154(7); 3310-3319 (1995); and Hawkins et al, J. Mol. Biol., 226: 889-896 (1992). Selective mutagenesis and selective or hypermutation at constant or hypermutation sites by activity-enhancing amino acid residues is described in U.S. Patent No. 6914128B1.

[0043] Approximately: As used herein, the terms “approximately” or “about” mean a value that is similar to the reference value stated, when applied to one or more of the values ​​of interest. In certain embodiments, unless otherwise stated or otherwise evident from the context, the terms “approximately” or “about” mean a range of values ​​that are within 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less in any direction of the reference value stated (except where such a number would exceed 100% of the possible value).

[0044] CDR: As used herein, the term “CDR” refers to the complementarity-determining region within the antibody variable sequence. A typical antibody molecule usually contains a heavy chain variable region (VH) and a light chain variable region (VL) that are involved in antigen binding. The VH and VL regions can be further subdivided into hypervariable regions, also known as “complementarity-determining regions” (“CDR”), which are interspersed with more conserved regions known as “framework regions” (“FR”). Each VH and VL typically consists of three CDRs and four FRs, arranged from amino-terminus to carboxy-terminus in the following order: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4. The extent of the framework regions and CDRs can be precisely identified using methods known in the art, for example, by the Kabat definition, IMGT definition, Chothia definition, AbM definition, and / or contact definition, all of which are well known in the art.Furthermore, Kabat, EA, et al. ( 1991 ) Sequences of Proteins of Immunological Interest , Fifth Edition , US Department of Health and Human Services , NIH Publication No . 91-3242;IMGT(Manufacturing Network), the International ImMunoGeneTics Information System(Manufacturing Network) http: / / www.imgt.org, Lefranc, M.-P. http: / / dx.doi.org / 10.1037 / 0021-843X.112.2.213 Ruiz, M. et al., Nucleic Acids Res., 27:209-212 (1999); Nucleic Acids Res., 31:307–310 (2003);Lefranc, M.-P. et al., In Silico Biol., 5, 0006 (2004) [Epub], 5:45-60 (2005);Lefranc, M.-P. et al., Nucleic Acids Res., 33:D593-597 (2005);Lefranc, M.-P. et al., Nucleic Acids Res., 37:D1006-1012 (2009);Lefranc, M.-P. et al., Nucleic Acids Res., 43:D413-422 (2015);Chothia et al., (1989) Nature 342:877; Chothia, C. et al. (1987) J. Mol. Biol. 196:901–917, Al-Lazikani et al (1997) J. Molec. Biol. 273:927-948;and Almagro, J. Mol. Recognize. 17:132-143 (2004) at hgmp.mrc.ac.uk and at bioinf.org.uk / abs.As used herein, CDR may refer to a CDR as defined by any method known in the art. Two antibodies having the same CDR means that the two antibodies have the same amino acid sequence of the CDR, as determined by the same method, e.g., the IMGT definition.

[0045] Typically, there are three CDRs in each variable region of the heavy and light chains, designated as CDR1, CDR2, and CDR3 for each variable region. As used herein, the term “CDR set” refers to a group of three CDRs that occur in a single variable region capable of binding to an antigen. The exact boundaries of these CDRs are defined differently by different systems. The system described by Kabat et al., Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md. (1987) and (1991)) not only provides a clear residue numbering system applicable to any variable region of an antibody, but also provides precise residue boundaries defining three CDRs. These CDRs may be called Kabat CDRs. The sub-parts of the CDRs may be denoted as L1, L2, and L3 or H1, H2, and H3, where "L" and "H" indicate the light chain and heavy chain regions, respectively. These regions may also be called Chothia CDRs and have boundaries that overlap with the Kabat CDRs. Other boundaries defining CDRs that overlap with the Kabat CDRs are described by Padlan (FASEB J. 9:133-139 (1995)) and MacCallum (J Mol Biol 262(5):732-45 (1996)) is described. Further CDR boundary definitions do not have to strictly adhere to one of the above systems, but despite overlapping with Kabat CDRs, they may be shortened or extended to take into account predictions or experimental findings that do not significantly affect antigen binding, such as specific residues, groups of residues, or even the entire CDR. The methods used herein may utilize CDRs defined according to any of these systems, but preferred embodiments use Kabat or Chothia defined CDRs.

[0046] The CDR of an antibody may have different amino acid sequences depending on the definition system used (e.g., IMGT definition, Kabat definition, or Chothia definition). The definition system annotates each amino acid of a given antibody sequence (e.g., VH or VL sequence) with a number, and the numbers corresponding to the heavy and light chain CDRs are provided in Table 3. Those skilled in the art can obtain the CDR sequences of the anti-HJV antibodies provided in Table 2 using the different numbering systems described in Table 3.

[0047] [Table 1]

[0048] CDR transplant antibody: The term "CDR transplant antibody" refers to an antibody that contains heavy and light chain variable region sequences from a single species, but V H and / or V L This refers to antibodies in which one or more CDR region sequences are replaced with CDR sequences of another species, for example, antibodies having mouse heavy and light chain variable regions in which one or more mouse CDRs (e.g., CDR3) are replaced with human CDR sequences.

[0049] Chimeric antibody: The term "chimeric antibody" refers to an antibody that contains a combination of heavy and light chain variable regions from one species and a constant region sequence from another species, for example, an antibody having mouse heavy and light chain variable regions linked to a human constant region.

[0050] Complementarity: As used herein, the term “complementarity” refers to the ability of two nucleotides or two sets of nucleotides to pair precisely. In particular, complementarity is a term that characterizes the degree of hydrogen bond pairs that result in bonding between two nucleotides or two sets of nucleotides. For example, if a base at one position of an oligonucleotide can hydrogen bond with a base at the corresponding position of a target nucleic acid (e.g., mRNA), then the bases are considered complementary to each other at that position. Base pairs can include both classical Watson-Crick base pairs and non-Watson-Crick base pairs (e.g., fluctuation base pairs and Hoogsteen base pairs). For example, in some embodiments, due to complementary base pairing, an adenosine-type base (A) is complementary to a thymidine-type base (T) or a uracil-type base (U), a cytosine-type base (C) is complementary to a guanosine-type base (G), and a universal base, such as 3-nitropyrrole or 5-nitroindole, is considered to hybridize with and be complementary to any of A, C, U, or T. Inosine (I) is also considered a universal base in the art and is considered to be complementary to any of A, C, U, or T.

[0051] Conservative amino acid substitutions: As used herein, “conservative amino acid substitutions” refer to amino acid substitutions that do not alter the relative charge or size characteristics of the protein in which the substitution is made. Variants can be prepared according to methods of modifying polypeptide sequences known to those skilled in the art, for example, references that compile such methods, e.g., Molecular Cloning: A Laboratory Manual, J. Sambrook, et al., eds., Fourth Edition, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, 2012, or Current Protocols in Molecular Biology, FM Ausubel, et al., eds., John Wiley & Sons, Inc., New York. Conservative amino acid substitutions include substitutions made in amino acids within the following groups: (a) M, I, L, V; (b) F, Y, W; (c) K, R, H; (d) A, G; (e) S, T; (f) Q, N; and (g) E, D.

[0052] Tolerance Reactivity: As used herein, and in the context of targeted agents (e.g., antibodies), the term “tolerance reactivity” refers to the property of a drug to specifically bind to one or more antigens of similar types or classes (e.g., diverse homologous, paralogous, or orthologous antigens) with similar affinity or binding activity. For example, in some embodiments, an antibody that is tolerance reactive to similar types or classes of human and non-human primate antigens (e.g., human hemoduvelin and non-human primate hemoduvelin) can bind to human and non-human primate antigens with similar affinity or binding activity. In some embodiments, the antibody is cross-reactive to similar types or classes of human and rodent antigens. In some embodiments, the antibody is cross-reactive to similar types or classes of rodent and non-human primate antigens. In some embodiments, the antibody is cross-reactive to similar types or classes of human, non-human primate, and rodent antigens.

[0053] Effective dose: As used herein, “effective dose” refers to the amount of each activator (e.g., anti-HJV antibody) required to produce a therapeutic effect on a subject, either alone or in combination with one or more other activators. In some embodiments, the therapeutic effect is a decrease in hepcidin levels or activity, an increase in transferrin saturation (TSAT%) levels, and / or a reduction in the disease state (e.g., a reduction in anemia or a reduction in the progression of myelofibrosis).

[0054] Framework: As used herein, the term “framework” or “framework sequence” refers to the sequence of the remaining variable region excluding the CDRs. Since the precise definition of a CDR sequence can be determined by different systems, the meaning of a framework sequence is assumed to be correspondingly subject to different interpretations. The six CDRs (CDR-L1, CDR-L2, and CDR-L3 of the light chain and CDR-H1, CDR-H2, and CDR-H3 of the heavy chain) also divide the framework regions of the light and heavy chains into four sub-regions (FR1, FR2, FR3, and FR4) of each chain, with CDR1 located between FR1 and FR2, CDR2 between FR2 and FR3, and CDR3 between FR3 and FR4. Without specifying a particular sub-region as FR1, FR2, FR3, or FR4, as otherwise mentioned, the framework region represents a combination of FRs within the variable region of a single, spontaneously occurring immunoglobulin chain. As used herein, FR represents one of four subregions, and FRs represents two or more of the four subregions that constitute a framework region. Human heavy and light chain acceptor sequences are known in the art. In one embodiment, acceptor sequences known in the art may be used in the antibodies disclosed herein.

[0055] Hemoduvelin (HJV): As used herein, the term “hemoduvelin (HJV)” (also known as repulsive guidance molecule C (RGMc) or type 2 hemochromatosis protein (HFE2)) refers to a membrane-bound soluble form protein that controls hepcidin production via the BMP / SMAD signaling pathway. The HFE2 gene encodes two known classes of HJV molecules: GPI-anchored and glycosylated, which are targeted to the membrane and undergo different fates. HJV exists in diverse isoforms, including two soluble isoforms and two membrane-bound isoforms. In some embodiments, the dominant membrane-bound isoform is a disulfide-bonded two-strand form consisting of an N-terminal and a C-terminal fragment. In some embodiments, the full-length single-strand isoform binds to the membrane but is released from the cell surface and accumulates in the extracellular fluid. In some embodiments, HJV may be of human (NCBI Gene ID 148738), non-human primate (e.g., NCBI Gene ID 698805), or rodent (e.g., NCBI Gene ID 69585 or NCBI Gene ID 310681) origin. In addition to HJV (RGMc), the repulsive guidance molecule family includes repulsive guidance molecule A (RGMa) and repulsive guidance molecule B (RGMb). RGMa and RGMb are expressed in the central nervous system during development and are thought to be involved in regulating axonal patterning and neuronal survival, while HJV is produced in the liver as well as in cardiac and skeletal muscle.

[0056] Hepcidin antagonist: As used herein, “hepcidin antagonist” refers to a drug that reduces hepcidin expression and / or hepcidin activity (directly or indirectly). In some embodiments, hepcidin antagonists inhibit hepcidin-induced ferroportin degradation. Thus, in some embodiments, hepcidin antagonists indirectly target hepcidin function through the hepcidin-stimulating pathway to reduce hepcidin expression. In some embodiments, hepcidin antagonists directly target hepcidin function, for example, by binding to hepcidin peptides to capture free hepcidin, or by binding to ferroportin to inhibit the hepcidin-ferroportin binding interaction, thereby reducing hepcidin-induced ferroportin degradation. In some embodiments, the hepcidin antagonist is a ferroportin inhibitor that disrupts the ferroportin-hepcidin interaction, as disclosed, for example, Ross SL, et al., Identification of Antibody and Small Molecule Antagonists of Ferroportin-Hepcidin Interaction. Front Pharmacol. 2017 Nov 21;8:838; Fung E., et al., High-Throughput Screening of Small Molecules Identifies Hepcidin Antagonists. Molecular Pharmacology March 2013, 83 (3) 681-690; and Angeliki Katsarou and Kostas Pantopoulos, Hepcidin Therapeutics. Pharmaceuticals (Basel). 2018 Dec; 11(4): 127, the respective relevant contents of which are incorporated herein by reference. In some embodiments, the hepcidin antagonist is an inhibitory nucleic acid (e.g., miRNA, shRNA, siRNA, or AmiRNA). In some embodiments, the hepcidin antagonist is an HJV-induced BMP signaling antagonist.

[0057] HJV-induced BMP signaling: As used herein, the term “HJV-induced BMP signaling” refers to signaling mediated by hemojuvelin (HJV), a membrane-bound coreceptor of bone morphogenetic protein (BMP) signaling, via BMP receptors. As described in Xia Y, et al., Hemojuvelin regulates hepcidin expression via a selective subset of BMP ligands and receptors independently of neogenin, Blood. 2008 May 15; 111(10): 5195-5204, in hepatocytes, HJV-induced BMP signaling positively regulates hepcidin mRNA expression. In some embodiments, HJV binds to BMP2, BMP4, BMP5, or BMP6 and induces BMP signaling, for example, positively regulating hepcidin levels in hepatocytes. In some embodiments, cleavage of HJV by matriptase-2 reduces the amount of cell surface HJV available to participate in BMP signaling. In some embodiments, the induction of BMP signaling by HJV is independent of neogenin. However, in some embodiments, neogenin facilitates the induction of BMP signaling by HJV, as described in Zhao et al, Neogenin Facilitates the Induction of Hepcidin Expression by Hemojuvelin in the Liver, J Biol Chem. 2016 Jun 3; 291(23): 12322-12335. In some embodiments, BMP6 is responsible for the iron-dependent activation of Smad signaling. In some embodiments, BMP6 is secreted from sinusoidal endothelial cells of the liver and binds to BMP receptors (BMPRs) on hepatocytes, thereby activating the SMAD signaling cascade. In such embodiments, HJV contributes as a co-receptor for such BMP6, for example, by positively regulating hepcidin levels on hepatocytes. In some embodiments, BMPs transmit signals by binding to one or a combination of type I and type II serine / threonine kinase receptors.BMP type II receptors include BMPRII, ActRIIA, and ActRIIB. BMP type I receptors include ALK3, ALK6, and ALK2. In some embodiments, upon ligand binding, the constitutively active type II receptor phosphorylates the type I receptor, which in turn phosphorylates intracellular receptor-activating Smad (R-Smad), primarily Smad1, Smad5, and / or Smad8. In such embodiments, the activated R-Smad forms a complex with a common partner, Smad4, and translocates to the nucleus to regulate gene transcription, e.g., induction of hepcidin expression.

[0058] Human Antibodies: As used herein, the term “human antibody” is intended to include antibodies having variable and constant regions derived from human germline immunoglobulin sequences. Human antibodies in this disclosure may include amino acid residues not encoded by human germline immunoglobulin sequences (e.g., mutations introduced by random or site-directed mutagenesis in vitro or by somatic mutagenesis in vivo), for example, in CDRs, particularly CDR3. However, as used herein, the term “human antibody” is not intended to include antibodies in which germline CDR sequences of another mammalian species, such as mouse, have been transplanted into human framework sequences.

[0059] Humanized antibody: The term "humanized antibody" refers to an antibody that contains heavy and light chain variable region sequences from a non-human species (e.g., mouse), but V H and / or V L This refers to antibodies in which at least a portion of the sequence has been modified to be more "human-like," that is, to more closely resemble the human germline variable sequence. One type of humanized antibody is a CDR-transplanted antibody, in which the human CDR sequence has been modified to be more similar to that of a non-human V H and V LThe sequence is introduced and replaces the corresponding non-human CDR sequence. In one embodiment, a humanized anti-hemoduvelin antibody and antigen-binding moiety are provided. Such antibodies may be produced by obtaining a mouse anti-hemoduvelin monoclonal antibody using humanization that employs in vitro genetic engineering after classical hybridoma techniques, such as disclosed in the international application PCT2005 / 123126A2 by Kasaian et al.

[0060] Inhibitory nucleic acids: As used herein, inhibitory nucleic acids refer to nucleic acids that can reduce the expression and / or function of a target gene. Non-limiting examples of inhibitory RNA include microRNAs (miRNAs), small interfering RNAs (siRNAs), short hairpin RNAs (shRNAs), artificial miRNAs (AmiRNAs), gapmers, mixmers, or antagomils. Inhibitory nucleic acids are useful, for example, for translational repression and / or gene silencing via ribonuclease-mediated degradation. Inhibitory nucleic acids can be delivered directly as oligonucleotides (e.g., isolated single-stranded or double-stranded oligonucleotides) and their formulations. In some embodiments, nucleic acids can be delivered in formulations or as conjugates that facilitate intracellular uptake, such as GalNac conjugates. However, in some embodiments, inhibitory nucleic acids can be delivered by viral vectors, such as lentiviruses, retroviruses, or recombinant adeno-associated viruses (rAAVs), which are engineered to express the inhibitory nucleic acid.

[0061] Isolated antibody: As used herein, “isolated antibody” is intended to mean an antibody that substantially does not contain other antibodies with different antigenic properties (for example, an isolated antibody that specifically binds to hemoduveline substantially does not contain antibodies that specifically bind to antigens other than hemoduveline). An isolated antibody that specifically binds to hemoduveline may, however, cross-reactive to other antigens such as other repulsive guidance molecule (RGM) proteins (e.g., RGMa and / or RGMb). Furthermore, an isolated antibody may not substantially contain other cellular material and / or chemical substances.

[0062] JAK / STAT signaling: As used herein, the term “JAK-STAT signaling” refers to a cell receptor-mediated signaling pathway that recruits a Janus kinase (JAK), such as Janus kinase 1 (JAK1) or Janus kinase 2 (JAK2), to activate a signaling and transcriptional activator (STAT), such as STAT3. In some embodiments, as described in Maliken, BD, et al., The Hepcidin Circuits Act: Balancing Iron and Inflammation, Hepatology. 2011 May; 53(5): 1764-1766, JAK-STAT signaling involves the binding of the cytokine interleukin-6 (IL-6) to its homologous cell receptor, which then recruits Janus kinase 2 (JAK2) to phosphorylate STAT3. In some embodiments, STAT3 then translocates to the nucleus (after JAK2 activation / phosphorylation). In some embodiments, activated STAT3 then induces hepcidin transcription, for example, by binding to a STAT3-binding motif in the hepcidin promoter region. Thus, in some embodiments, hepcidin expression is induced via JAK-STAT signaling during inflammation induced by IL-6-mediated STAT3 activation.

[0063] Kabat numbering: The terms “Kabat numbering,” “Kabat definition,” and “Kabat labeling” are used interchangeably herein. As recognized in the art, these terms refer to a system for numbering amino acid residues that are more variable (i.e., hypervariable) than other amino acid residues in the heavy and light chain variable regions of an antibody, or in its antigen-binding moiety (Kabat et al. (1971) Ann. NY Acad, Sci. 190:382-391 and, Kabat, EA, et al. (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, US Department of Health and Human Services, NIH Publication No. 91-3242). For the heavy chain variable region, the hypervariable region is the range of amino acids 31-35 of CDR1, 50-65 of CDR2, and 95-102 of CDR3. Regarding the light chain variable region, the hypervariable region is located in the range of amino acids 24 to 34 of CDR1, 50 to 56 of CDR2, and 89 to 97 of CDR3.

[0064] Myelofibrosis: As used herein, the term “myelofibrosis” refers to a progressive fibrosis characterized by pathological myeloproliferation and abnormal cytokine production resulting in inflammation and / or dysfunction of the bone marrow niche of the subject. Myelofibrosis and related fibrosis often arise from a non-clonal fibroblastic response to inflammatory and fibrogenic cytokines produced by abnormal clonal myeloid cells, such as megakaryocytes. Myelofibrosis typically results in bone marrow failure, splenomegaly, catabolism, and anemia. In some embodiments, myelofibrosis occurs de novo in the subject. In such embodiments, myelofibrosis is considered “primary” myelofibrosis. However, in some embodiments, myelofibrosis arises from a pre-existing myeloproliferative neoplasm. In some embodiments, the pre-existing myeloproliferative neoplasm is polycythemia. In some embodiments, the pre-existing myeloproliferative neoplasm is essential thrombocytopenia.

[0065] Myelofibrosis-associated anemia: As used herein, the term “myelofibrosis-associated anemia” refers to a condition occurring in the context of myelofibrosis or as a complication of myelofibrosis, characterized by a deficiency in the blood’s ability to transport oxygen. In some embodiments, myelofibrosis-associated anemia is a result of a deficiency in red blood cells, a deficiency in hemoglobin, and / or a deficiency in total blood volume. In some embodiments, myelofibrosis-associated anemia is iron deficiency anemia or myeloplasia anemia. In some embodiments, myelofibrosis-associated anemia is further associated with chronic inflammatory diseases. Examples of anemia other than myelofibrosis-associated anemia include anemia associated with rheumatoid arthritis, anemia of infection, autoimmune hemolytic anemia, aplastic anemia, hypoplastic anemia, pure red cell aplasia and anemia resulting from renal failure or endocrine disorders, megaloblastic anemia, anemia resulting from heme or globin synthesis deficiencies, anemia caused by structural defects in red blood cells, such as sickle cell anemia and sideroblastic anemia, anemia associated with chronic infections, such as malaria, trypanosomiasis, HIV, hepatitis viruses or other viruses, anemia caused by bone marrow deficiency in the absence of myelofibrosis, and chemotherapy-induced anemia.

[0066] Oligonucleotides: As used herein, the term “oligonucleotide” refers to oligomeric nucleic acid compounds up to 200 nucleotides in length. Examples of oligonucleotides include, but are not limited to, RNAi oligonucleotides (e.g., siRNA, shRNA), microRNAs, gapmers, mixedmers, phosphorodiamidate morpholinos, peptide nucleic acids, aptamers, and guide nucleic acids (e.g., Cas9 guide RNA). Oligonucleotides may be single-stranded or double-stranded. In some embodiments, oligonucleotides may contain one or more modified nucleotides (e.g., 2'-O-methylglycosulfate, purine, or pyrimidine modification). In some embodiments, oligonucleotides may contain one or more modified internucleotide bonds. In some embodiments, oligonucleotides may contain one or more phosphorothioate bonds, which may have an Rp or Sp stereochemical structure.

[0067] Recombinant Adeno-Associated Virus (rAAV): The term “recombinant adeno-associated virus (rAAV)” refers to AAV that is artificially produced or obtained using recombinant methods. Recombinant AAV (rAAV) preferably has tissue-specific targeting ability, so that the rAAV transgene is specifically delivered to one or more predetermined tissues (e.g., eye tissue). rAAV typically contains an AAV capsid protein that encloses the recombinant AAV vector. A “recombinant AAV (rAAV) vector” typically consists of, at a minimum, the transgene and its regulatory sequences, as well as 5' and 3' AAV terminal inversion sequences (ITRs). The AAV capsid is a crucial element in determining these tissue-specific targeting abilities (e.g., tissue tropism). In some embodiments, rAAV with a capsid appropriate for the targeted tissue may be used. In some embodiments, rAAV contains an AAV capsid protein specific for liver delivery. In some embodiments, the AAV capsid protein is of the AAV2, AAV3B, AAV8, or LK03 serotype. In some embodiments, the rAAV vector includes a liver-specific promoter that drives the expression of an inhibitory nucleic acid targeting BMP-6. Non-limiting examples of liver-specific promoters include the human serum albumin promoter, the alpha-1 antitrypsin promoter, the apolipoprotein E / CI liver regulatory region / human alpha-1 antitrypsin chimeric promoter, or the alpha-1 microglobulin / bicinin enhancer / human thyroxine-binding globulin (TBG) chimeric promoter. AAV capsid proteins and liver-specific promoters for liver specificity are described in the Art, for example, Kattenhorn et al., Adeno-Associated Virus Gene Therapy for Liver, Human Gene Therapy, Vol. 27, No. 12.

[0068] Recombinant Antibodies: As used herein, the term “recombinant human antibodies” includes all human antibodies (as described in detail herein) prepared, expressed, created, or isolated by recombinant means, such as antibodies expressed using recombinant expression vectors transfected into host cells; antibodies isolated from recombinant, combinatorial human antibody libraries (Hoogenboom HR, (1997) TIB Tech. 15:62-70; Azzazy H., and Highsmith WE, (2002) Clin. Biochem. 35:425-445; Gavilondo JV, and Larrick JW (2002) BioTechniques 29:128-145; Hoogenboom H., and Chames P. (2000) Immunology Today 21:371-378); antibodies isolated from animals (e.g., mice) that are transgenic of human immunoglobulin genes (e.g., Taylor, LD, et al. (1992) Nucl. Acids It is intended to include antibodies prepared, expressed, created or isolated by any other means, including splicing of human immunoglobulin gene sequences to other DNA sequences (see Res. 20:6287-6295; Kellermann SA., and Green LL (2002) Current Opinion in Biotechnology 13:593-597; Little M. et al (2000) Immunology Today 21:364-370), or by any other means including splicing of human immunoglobulin gene sequences to other DNA sequences. Such recombinant human antibodies have variable and constant regions derived from human germline immunoglobulin sequences. However, in certain embodiments, such recombinant human antibodies are subjected to in vitro mutagenesis (or in vivo somatic mutagenesis, if transgenic animals of human Ig sequences are used), and thus the recombinant antibody is V H and V L The amino acid sequence of the region is human germline V H and V LThe sequence is sequence-derived or related, but does not necessarily have to be naturally present in the human antibody germline repertoire in vivo. One embodiment of the present disclosure provides a complete human antibody capable of binding to human hemoduverin, which can be produced using techniques well known in the art, such as using a human Ig phage library, for example, disclosed in international application PCT2005 / 007699A2 by Jermutus et al.

[0069] Selective: As used herein, the terms “selective” or “selectively” refer to the ability of a molecule to produce an effect associated with its target molecule compared to a reference molecule. For example, a molecule that selectively inhibits its target molecule means that the molecule can inhibit the target molecule to a degree that is distinguishable from the reference molecule in an inhibition assay or other inhibitory environment. For example, with respect to an inhibitor, the term “selectively inhibits” refers to the ability of an inhibitor to inhibit its target molecule to a degree that is distinguishable from a reference molecule that is substantially not inhibited in an inhibition assay, to the extent that it enables selective inhibition of the target molecule, as described herein. For example, the half-number inhibitory concentrations (IC50) of the target molecule and / or reference molecule can be tested by a kinase inhibition assay (e.g., kinase efficacy assay by Carna Biosciences), as described in Asshoff, M. et al., Momelotinib inhibits ACVR1 / ALK2, decreases hepcidin production, and ameliorates anemia of chronic disease in rodents. Blood. 2017 Mar 30; 129(13): 1823-1830. In this assay, an inhibitor solution (e.g., a solution containing the selective inhibitor to be tested) / kinase substrate is mixed with a target molecule solution (e.g., ALK2) or a reference molecule solution (e.g., JAK1 or JAK2) and incubated at room temperature for 1 hour. After the reaction is complete, the signal produced by the enzymatic activity to the substrate can be measured. The half-number inhibitory concentrations of the target molecule and / or reference molecule can be calculated. In some embodiments, the molecules described herein selectively bind to the target molecule. In some embodiments, the molecules described herein selectively inhibit the target molecule. In some embodiments, the molecules described herein selectively antagonistize target molecules. In some embodiments, the molecules described herein selectively neutralize target molecules.

[0070] Specific binding: As used herein, the term “specific binding” refers to the ability of a molecule to bind to a binding partner with a certain degree of affinity or binding strength so that the molecule can be used to distinguish the binding partner from a suitable control in a binding assay or other binding environment. With respect to an antibody, the term “specific binding” refers to the ability of an antibody to bind to a particular antigen with a certain degree of affinity or binding strength so that the antibody can be used to distinguish the particular antigen from others to the extent that it enables selective targeting of certain cells, such as muscle cells, through binding to the antigen, as described herein, for example. In some embodiments, the antibody binds at least about 10 -4 M, 10 -5 M, 10 -6 M, 10 -7 M, 10 -8 M, 10 -9 M, 10 -10 M, 10 -11 M, 10 -12 M, 10 -13 K for binding to M or smaller targets D If present, the antibody binds specifically to the target. In some embodiments, the antibody binds specifically to hemoduveline.

[0071] Subject: As used herein, the term “subject” refers to mammals. In some embodiments, the subject is a non-human primate or rodent. In some embodiments, the subject is a human. In some embodiments, the subject is a patient, e.g., a human patient having or suspected of having a disease. In some embodiments, the subject is a human patient having or suspected of having myelofibrosis and / or one or more conditions resulting from myelofibrosis.

[0072] Treatment: As used herein, the terms “treating” or “treatment” mean the application or administration of a composition comprising one or more active agents to a subject having a target disease or disorder, symptoms of a disease / disorder, or a predisposition to a disease / disorder, for the purpose of treating, curing, alleviating, reducing, altering, relieving, improving, or influencing a disorder, symptoms of a disease, or predisposition to a disease or disorder. Alleviating a target disease / disorder includes delaying or preventing the onset or progression of the disease, or reducing the severity of the disease.

[0073] II. Hepcidin Antagonists Among other embodiments, this disclosure relates to hepcidin antagonists and related methods for the treatment of myelofibrosis, such as anemia associated with myelofibrosis. Figure 2 shows the hepcidin-stimulating pathway and the physiological regulation of iron homeostasis by hepcidin. As shown, hepcidin manipulates iron-releasing target cells (e.g., hepatocytes, duodenal intestinal cells, tissue macrophages, and other cell types) by binding to the iron efflux protein, ferroportin. Hepcidin binding blocks iron efflux and induces ubiquitination, internalization, and lysosomal degradation of ferroportin. This results in a decrease in intracellular iron retention and ultimately a decrease in systemic iron levels. Thus, in some embodiments, the hepcidin antagonists of this disclosure are hepcidin inhibitors that antagonistize hepcidin function by blocking hepcidin or stabilizing ferroportin to inhibit the binding of hepcidin to ferroportin.

[0074] The HAMP gene encodes a hepcidin precursor protein, which is expressed primarily in hepatocytes of the liver at lower levels than in other cells of extrahepatic tissues. The precursor protein is subsequently cleaved to produce physiologically active hepcidin. In some embodiments, the hepcidin antagonists of this disclosure are HAMP antagonists that antagonistize hepcidin function by binding to HAMP or its transcripts or translation products, or by inhibiting HAMP transcription or translation regulators to reduce HAMP expression.

[0075] Further examples of HAMP transcription regulators include, but are not limited to, SMAD1 / 5 / 8 (e.g., the BMP-SMAD signaling pathway) and STAT3 (e.g., the JAK-STAT signaling pathway). Therefore, in some embodiments, HAMP antagonists are BMP-SMAD signaling pathway inhibitors or JAK-STAT signaling pathway inhibitors.

[0076] i. Hemoduvelin-induced BMP signaling antagonists In some embodiments, hemoduvelin-induced BMP signaling antagonists are provided herein to inhibit BMP-SMAD signaling in order to reduce hepcidin expression and / or function, for example, to modulate iron homeostasis for the treatment of myelofibrosis and / or one or more conditions resulting from myelofibrosis. In some embodiments, such methods are based on the recognition that an increase in serum or tissue iron triggers transcriptional induction of hepcidin via the BMP-SMAD signaling pathway. In some embodiments, HJV contributes as a BMP coreceptor and positively regulates hepcidin levels. In certain cells, e.g., hepatocytes, HJV-induced BMP signaling positively regulates hepcidin mRNA expression. In such embodiments, HJV binds to BMP2, BMP4, BMP5, and / or BMP6 and mediates BMP signaling, for example, positively regulating hepcidin levels in hepatocytes. In some embodiments, BMPs transmit signals by binding to one or a combination of type I and type II serine / threonine kinase receptors. In some embodiments, upon ligand binding, the constitutively active type II receptor phosphorylates the type I receptor, which in turn phosphorylates intracellular receptor-activating Smad (R-Smad), primarily Smad1, Smad5, and / or Smad8. In such embodiments, the activated R-Smad forms a complex with a common partner, Smad4, and translocates to the nucleus to regulate gene transcription, e.g., the induction of hepcidin expression.

[0077] In some embodiments, the methods provided herein utilize HJV-induced BMP signaling antagonists for the treatment of anemia associated with myelofibrosis. In some embodiments, the HJV-induced signaling antagonist is a BMP antagonist that directly or indirectly inhibits BMP signaling (e.g., BMP antibodies, BMP inhibitory nucleic acids, soluble BMP receptors, soluble hemoduvelin, etc.). In some embodiments, the BMP antagonist is an anti-BMP antibody that inhibits signaling. In some embodiments, recombinant noggin is provided as the BMP antagonist. In some embodiments, the anti-BMP antibody specifically binds to and inhibits a particular BMP, e.g., BMP6. However, in some embodiments, the anti-BMP binds to and inhibits a variety of BMPs. In some embodiments, the anti-BMP antibody is an antibody against a BMP that binds to HJV.

[0078] In some embodiments, the anti-BMP antibody is an anti-BMP2 antibody that specifically binds to BMP2 and inhibits downstream signaling. Preferred anti-BMP2 antibodies are disclosed, for example, in Gorrell RE, et al., Identification of a bone morphogenetic protein type 2 receptor neutralizing antibody. BMC Res Notes. 2019; 12: 331.; and Kang MH, et al., BMP2 accelerates the motility and invasiveness of gastric cancer cells via activation of the phosphatidylinositol 3-kinase (PI3K) / Akt pathway. Exp Cell Res. 2010 Jan 1;316(1):24-37, the contents of which are incorporated herein by reference.

[0079] In some embodiments, the anti-BMP antibody is an anti-BMP4 antibody that specifically binds to BMP4 and inhibits downstream signaling. Preferred anti-BMP4 antibodies are disclosed, for example, in Calpe S. et al., Comparison of newly developed anti-bone morphogenetic protein 4 llama-derived antibodies with commercially available BMP4 inhibitors. MAbs. 2016 May-Jun; 8(4): 678-688, the contents of which are incorporated herein by reference.

[0080] In some embodiments, BMP-2 and / or BMP4 antagonists are referenced in U.S. Patent Publication No. 8338377, published December 25, 2012, titled “BMP-ALK3 antagonists and uses for promoting bone growth”; U.S. Patent No. 9738636, published August 22, 2017, titled “Fused heterocyclic compounds as selective BMP inhibitors”; U.S. Patent Application Publication No. 2019218214, published May 21, 2019, titled “Inhibition of BMP Signaling Compounds, Compositions and Uses Thereof”; U.S. Patent Application Publication No. 2019284183, published September 19, 2019, titled “Inhibition of bmp signaling, compounds, compositions and uses thereof”; and U.S. Patent Application Publication No. 2019284183, published February 20, 2020, titled “Fused heterocyclic compounds as selective bmp These are BMP2 and / or BMP4 antagonists disclosed in U.S. Patent Application Publication No. 2020054643 of “inhibitors”.

[0081] In some embodiments, the anti-BMP antibody is an anti-BMP5 antibody that specifically binds to BMP5 and inhibits downstream signaling. In some embodiments, the anti-BMP5 antibody is, for example, human BMP-5 antibody AF615 (R&D Systems) or human BMP-5 antibody MAB7151 (R&D Systems).

[0082] In some embodiments, the anti-BMP antibody is an anti-BMP6 antibody that specifically binds to BMP6 and inhibits downstream signaling. In some embodiments, the anti-BMP6 antibodies for use in the methods provided herein are U.S. Patent No. 8,795665B2, titled “BMP-6 antibodies,” issued on August 5, 2014; U.S. Patent No. 8,980582B2, titled “BMP-6 antibodies and DNA encoding the same,” issued on March 17, 2015; U.S. Patent No. 9,439963B2, titled “Methods of treating anaemia,” issued on September 13, 2016; U.S. Patent No. 9,862764B2, titled “Compositions and methods for antibodies targeting BMP6,” issued on January 19, 2018; and U.S. Patent No. 9,862764B2, titled “Methods for treating disease using inhibitors of bone morphogenetic protein 6,” published on December 21, 2017. The anti-BMP-6 antibody is disclosed in International Publication No. 2017216724A1 of “(bmp6)”; International Publication No. 2017191437A1 of “Methods, regimens, combinations & antagonists” published on November 9, 2017; and International Publication No. 2020065252 of “Antagonists” published on April 2, 2020. In some embodiments, the anti-BMP-6 antibody is LY3113593. In some embodiments, the anti-BMP-6 antibody is CSJ137. In some embodiments, the anti-BMP-6 antibody is KY1070.

[0083] In some embodiments, the BMP antagonist is an inhibitory nucleic acid that inhibits BMP expression (e.g., a dsRNA, siRNA, miRNA, shRNA, AmiRNA, antisense oligonucleotide (ASO), or aptamer that targets BMP2, BMP4, BMP5, or BMP6). In some embodiments, the BMP-targeting inhibitory nucleic acid may be used herein in the treatment of myelofibrosis and related conditions. In some embodiments, the BMP6-targeting inhibitory nucleic acid is, for example, the BMP6-targeting inhibitory nucleic acid disclosed in U.S. Patent No. 9,228188, issued on January 5, 2016, entitled “Compositions and method for inhibiting hepcidin antimicrobial peptide (HAMP) or HAMP-related gene expression,” whose entire contents are incorporated herein by reference. In some embodiments, the BMP-6-targeting inhibitory nucleic acid is an inhibitory nucleic acid. In some embodiments, the inhibitory nucleic acid is a miRNA that targets BMP-6. In some embodiments, the inhibitory nucleic acid is an shRNA that targets BMP-6. In some embodiments, the inhibitory nucleic acid is an siRNA that targets BMP-6. In some embodiments, the inhibitory nucleic acid is an AmiRNA that targets BMP-6.

[0084] In some embodiments, further examples of BMP6 antagonists include, but are not limited to, TP-0184, FKBP12, twist gastrulation protein, dolsomorphin, noggin, chordin, ventroptin, follistatin, follistatin-related genes (FLRG), heparin (e.g., SST0001, RO-82, RO-68, NAc-91, and NacRO-00), sulfated glycosaminoglycans, and sclerostin domain-containing protein 1 (SOSTDC1). Further examples of BMP6 antagonists that may be useful in certain ways provided herein are offered. In some embodiments, the BMP6 antagonist is the subject of U.S. Patent No. 8,318,167, issued November 27, 2012, titled “METHODS AND COMPOSITIONS FOR REGULATING IRON HOMEOSTASIS BY MODULATION OF BMP-6”; U.S. Patent No. 9,556,251, issued January 31, 2017, titled “METHODS AND COMPOSITIONS TO REGULATE HEPCIDIN EXPRESSION”; U.S. Patent No. 9,862,764, issued January 9, 2018, titled “COMPOSITIONS AND METHODS FOR ANTIBODIES TARGETING BMP6”; U.S. Patent No. 9,682,983, issued June 20, 2017, titled “BMP INHIBITORS AND METHODS OF USE THEREOF”; and the subject of the “INHIBITORS” patent issued August 13, 2013, the entire contents of which are incorporated herein by reference. U.S. Patent No. 8507501 for “O OF THE BMP SIGNALING PATHWAY”; U.S. Patent No. 9,738636 for “FUSED HETEROCYCLIC COMPOUNDS AS SELECTIVE BMP INHIBITORS” issued on August 22, 2017; and U.S. Patent No. 8,795665 for “BMP-6 ANTIBODIES” issued on August 5, 2014;U.S. Patent Application Publication No. 2010 / 0093760, published on April 15, 2010, with the title "METHODS FOR IDENTIFYING COMPOUNDS THAT MODULATE CELL SIGNALING AND METHODS EMPLOYING SUCH COMPOUND"; U.S. Patent Application Publication No. 2014 / 0199314, published on July 17, 2014, with the title "METHODS AND COMPOSITIONS FOR REGULATING IRON HOMEOSTASIS BY MODULATION OF BMP-6"; U.S. Patent Application Publication No. 2014 / 0086919, published on March 27, 2014, with the title "METHODS AND COMPOSITIONS FOR REGULATING IRON HOMEOSTASIS BY MODULATION OF BMP-6"; U.S. Patent Application Publication No. 2014 / 0086919, published on September 15, 2016, with the title "COMPOSITIONS AND METHODS FOR U.S. Patent Application Publication No. 2016 / 0263117 for “CARDIOVASCULAR DISEASE”; U.S. Patent Application Publication No. 2016 / 0115167 for “BMP INHIBITORS AND METHODS OF USE THEREOF” published on April 28, 2016; U.S. Patent Application Publication No. 2017 / 0197968 for “COMPOSITIONS AND METHODS FOR INHIBITING BMP” published on July 13, 2017; U.S. Patent Application Publication No. 2017 / 0190705 for “COMPOSITIONS AND METHODS FOR INHIBITING BMP” published on July 6, 2017; U.S. Patent Application Publication No. 2017 / 0190705 for “COMPOSITIONS AND METHODS FOR INHIBITING BMP” published on October 26, 2017 U.S. Patent Application Publication No. 2017 / 0305883 for “BMP”; U.S. Patent Application Publication No. 2018 / 0021340, published on January 25, 2018, with the title “METHODS AND COMPOSITIONS FOR THE TREATMENT OR PREVENTION OF ABNORMAL BONE FORMATION IN A SOFT TISSUE”;International application PCT2017 / 216724, titled “METHODS FOR TREATING DISEASE USING INHIBITORS OF BONE MORPHOGENETIC PROTEIN 6 (BMP6),” published on December 21, 2017; International publication 2018 / 136634, titled “FUSED HETEROCYCLIC COMPOUNDS AS SELECTIVE BMP INHIBITORS,” published on July 26, 2018; International publication 2018 / 053234, titled “TWISTED GASTRULATION POLYPEPTIDES AND USES THEREOF,” published on March 22, 2018; International publication 2018 / 053234, titled “REGULATOR OF BMP-SMAD SIGNALING AND USES The BMP6 antagonist is disclosed in International Publication No. 2018 / 185341 of “THEREOF” and International Publication No. 2016 / 146651 of “MACROCYCLIC ACTIVIN-LIKE RECEPTOR KINASE INHIBITORS,” published on September 22, 2016.

[0085] In some embodiments, the hemoduvelin-induced BMP signaling antagonist is a BMP receptor antagonist. In some embodiments, the BMP receptor antagonist is a neutralizing antibody against the BMP receptor. In some embodiments, BMP signals by binding to one or a combination of type I and type II serine / threonine kinase receptors. BMP type II receptors include BMPRII, ActRIIA, and ActRIIB. BMP type I receptors include ALK3, ALK6, and ALK2. In some embodiments, the BMP receptor antagonist neutralizes an antibody that targets the BMP receptor. In some embodiments, the BMP receptor neutralizing antibody is an anti-BMPRII antibody, an anti-ActRIIA antibody, an anti-ActRIIB antibody, an anti-ALK3 antibody, an anti-ALK6 antibody, or an anti-ALK2 antibody. In some embodiments, the BMP receptor neutralizing antibody is an anti-ALK2 antibody. In some embodiments, the anti-ALK2 antibody is the anti-ALK2 antibody disclosed in U.S. Patent No. 1,0428148B2, titled “Anti-ALK2 antibody,” issued on 1 October 2019; International Publication No. 2020086730A1, titled “Alk2 antibodies and methods of use thereof,” published on 30 April 2020; and U.S. Patent Application Publication No. 2018 / 0118835, titled “ANTI-ALK2 ANTIBODY,” published on 3 May 2018, the contents of which are incorporated herein by reference.

[0086] In some embodiments, the BMP receptor antagonist is an inhibitory nucleic acid that inhibits the expression of the BMP receptor (e.g., BMP type I receptor or BMP type II receptor). In some embodiments, the inhibitory nucleic acid is an inhibitory nucleic acid that inhibits ALK2 expression. Therefore, in some embodiments, inhibitory nucleic acids that inhibit the expression of the BMP receptor may be used herein for the treatment of myelofibrosis and related conditions.

[0087] In some embodiments, the hemoduverin-induced BMP signaling antagonist selectively inhibits its target molecule. In some embodiments, the target molecule is a BMP receptor. In some embodiments, the hemoduverin-induced BMP signaling antagonist selectively inhibits its target molecule compared to a reference molecule. In some embodiments, the reference molecule is JAK2. In some embodiments, the target molecule is ALK2. In some embodiments, the hemoduverin-induced BMP signaling antagonist selectively inhibits its target molecule compared to a reference molecule, resulting in at least 10 times (e.g., at least 10 times, at least 20 times, at least 30 times, at least 40 times, at least 50 times, at least 60 times, at least 70 times, at least 80 times, at least 90 times, or higher) the inhibition compared to the target molecule. 2 times (for example, at least 100 times, at least 200 times, at least 300 times, at least 400 times, at least 500 times, at least 600 times, at least 700 times, at least 800 times, at least 900 times, or higher), 10 3 times (for example, at least 1000 times, at least 2000 times, at least 3000 times, at least 4000 times, at least 5000 times, at least 6000 times, at least 7000 times, at least 8000 times, at least 9000 times, or higher), 10 4 times (for example, at least 1x10) 4 Double, at least 2x10 4 Double, at least 3x10 4 Double, at least 4x10 4 Double, at least 5x10 4 Double, at least 6x10 4 Double, at least 7x10 4 Double, at least 8x10 4 Double, at least 9x10 4 (times, or more), 10 5 times (for example, at least 1x10) 5 Double, at least 2x10 5 Double, at least 3x10 5 Double, at least 4x10 5 Double, at least 5x10 5Double, at least 6x10 5 Double, at least 7x10 5 Double, at least 8x10 5 Double, at least 9x10 5 (times, or more), 10 6 times (for example, at least 1x10) 6 Double, at least 2x10 6 Double, at least 3x10 6 Double, at least 4x10 6 Double, at least 5x10 6 Double, at least 6x10 6 Double, at least 7x10 6 Double, at least 8x10 6 Double, at least 9x10 6 It has a reference molecule half-block inhibitory concentration (IC50) that is 10 times or more. In some embodiments, hemoduvelin-induced BMP signaling antagonists selectively inhibit their target molecule compared to the reference molecule, resulting in a 10- to 10-fold inhibitory effect compared to the target molecule. 2 times, 10 times~10 3 double, or 10 times ~ 10 4 times, 50 times ~ 10 5 double or 100 times ~ 10 6 It has a reference molecule half-block inhibitory concentration (IC50) in the range of 1x or higher. In some embodiments, the IC50 is determined according to a kinase inhibitory assay (e.g., an assay described in Asshoff, M. et al., Momelotinib inhibits ACVR1 / ALK2, decreases hepcidin production, and ameliorates anemia of chronic disease in rodents. Blood. 2017 Mar 30; 129(13): 1823-1830 (e.g., kinase inhibitory assay by Carna Biosciences)). In some embodiments, the selective BMP receptor inhibitor is a selective ALK2 inhibitor as determined by the kinase inhibitory assay. In some embodiments, the selective BMP receptor inhibitor does not inhibit JAK1 / JAK2. In some embodiments, the selective ALK2 inhibitor is not momerotinib.

[0088] In some embodiments, the BMP receptor antagonist is a small molecule inhibitor of the BMP receptor. In some embodiments, the BMP receptor antagonist is a small molecule ALK2 inhibitor.In some embodiments, the ALK2 inhibitor is one of the following, the contents of which are incorporated herein by reference: U.S. Patent No. 10233186, “Inhibitors of activin receptor-like kinase,” issued March 19, 2019; U.S. Patent No. 10202356, “JAK2 AND ALK2 INHIBITORS AND METHODS FOR THEIR USE,” issued February 12, 2019; U.S. Patent No. 10669277B2, “Inhibitors of activin receptor-like kinase,” issued June 2, 2020; International Publication No. 2019079649, “Substituted pyrrolopyridines as inhibitors of activin receptor-like kinase,” published April 25, 2019; and “NOVEL ALK2 INHIBITORS AND METHODS FOR INHIBITING BMP,” published November 1, 2018. International Publication No. 2018 / 200855 for “SIGNALING”; International Publication No. 2020086730 for “Alk2 antibodies and methods of use thereof” published on April 30, 2020; International Publication No. 2020086963 for “Crystal forms of an alk2 inhibitor” published on April 30, 2020; International Publication No. 2020068729 for “Pyrazolo[4,3-d]pyrimidine compounds as alk2 and / or fgfr modulators” published on April 2, 2020; US Patent Application Publication No. 2020095250 for “Pyrazolopyrimidine compounds and uses thereof” published on March 26, 2020; “Imidazopyridazine and imidazopyridine compounds and uses” published on June 25, 2020 It is an ALK2 inhibitor disclosed in U.S. Patent Application Publication No. 2020199131 of “them”.Further suitable ALK2 inhibitors are disclosed in Hudson, L. et al., Novel Quinazolinone Inhibitors of ALK2 Flip between Alternate Binding Modes: Structure-Activity Relationship, Structural Characterization, Kinase Profiling, and Cellular Proof of Concept. Med. Chem. 2018, 61, 16, 7261-7272 and Carvalho D, et al., ALK2 inhibitors display beneficial effects in preclinical models of ACVR1 mutant diffuse intrinsic pontine glioma. Communications Biology volume 2, Article number: 156 (2019), the relevant content of each thereof is incorporated herein by reference. In some embodiments, a suitable ALK-2 inhibitor for use in the methods provided herein is KER-047. In some embodiments, a suitable ALK-2 inhibitor for use in the methods provided herein is BLU-782. In some embodiments, the ALK-2 inhibitor suitable for use in the methods provided herein is INCB000928. In some embodiments, the ALK2 inhibitor is LDN-212854, LDN-193189, or LDN-214117.

[0089] In some embodiments, the BMP antagonist is a BMP ligand trap. In some embodiments, the BMP ligand trap is a soluble BMP receptor. In some embodiments, the soluble BMP receptor is fused to the Fc portion of an immunoglobulin (e.g., ActRIIa-Fc ligand trap or Dalantercept, activin receptor-like kinase-1 ligand trap, ActRIIb-Fc ligand trap). Inhibition of BMP signaling by inhibiting the BMP receptor is described, for example, in Gomez-Puerto MC, et al., Bone morphogenetic protein receptor signal transduction in human disease. J Pathol. 2019 Jan; 247(1): 9-20. In some embodiments, the BMP ligand trap is incorporated herein by reference by the following patent applications: U.S. Patent No. 7,709605B2, published May 4, 2010, titled “ActRII receptor polypeptides, methods and compositions”, U.S. Patent No. 9,526759, published December 27, 2016, titled “Activin-actriia antagonists and uses for treating or preventing breast cancer”, U.S. Patent No. 8,058229, published November 15, 2011, titled “A method of increasing red blood cell levels or treating anemia in a patient”, U.S. Patent Application Publication No. 2013243743, published September 19, 2013, titled “Methods and compositions for treating ineffective erythropoiesis”, and U.S. Patent Application Publication No. 2013243743, published June 4, 2019, titled “Activin Type 2 Receptor U.S. Patent No. 10307455, issued on August 2, 2011, is titled "Activin-ActRII antagonists and uses for increasing red blood cellU.S. Patent No. 7,988,973, titled "levels", U.S. Patent No. 7,612,041, published November 3, 2009, titled "An isolated activating-binding ActRIIA polypeptide comprising the SEQ ID NO: 7 and uses for promoting bone growth", U.S. Patent Publication No. 2011070233 A1, published March 24, 2011, titled "Actriib antagonists and dosing and uses thereof", U.S. Patent No. 7,960343, published June 14, 2011, titled "Activin-actriia antagonists and uses for decreasing or inhibiting FSH secretion", U.S. Patent Publication No. 2019282663, published September 19, 2019, titled "Activin receptor type iia variants and methods of use thereof", and published May 16, 2019, titled "Activin receptor type iia variants and methods of use International Publication No. 2019094751 of “thereof”; U.S. Patent No. 7,842663, published November 30, 2010, with the title “Variants derived from ACTRIIB and uses therefor”; U.S. Patent Application Publication No. 2010008918, published January 14, 2010; U.S. Patent No. 8,058229, published November 15, 2011, with the title “A method of increasing red blood cell levels or treating anemia in a patient”; U.S. Patent No. 8,293881, published October 23, 2012, with the title “An isolated nucleic acid encoding a truncated actriib fusion protein”; and “Method of detection of neutralizing anti-actriib” published July 1, 2014.U.S. Patent No. 8,765,385 for "antibodies", U.S. Patent Publication No. 2015361163, published December 17, 2015, titled "Methods for increasing red blood cell levels and treating sickle-cell disease", U.S. Patent Publication No. 2017274077, published September 28, 2017, titled "Methods for increasing red blood cell levels and treating ineffective erythropoiesis", U.S. Patent Publication No. 2018050085, published February 22, 2018, titled "Methods and compositions for treating myelofibrosis", International Publication No. 2018067740, published April 12, 2018, titled "Compositions and method for treating kidney disease", and "Variant actriib proteins and uses" published February 20, 2020. U.S. Patent Application Publication No. 2020055919 for “thereof”; International Publication No. 2020092523, published on May 7, 2020, for “Treatment of anemia due to very low, low, or intermediate risk myelodysplastic syndromes in subjects with ring sideroblasts using activating-actrii ligand traps”; U.S. Patent No. 10189882, published on January 29, 2019, for “Methods for treating myelodysplastic syndromes and sideroblastic anemias”; International Publication No. 2019 / 140283, published on July 18, 2019, for “Activin receptor type iib variants and methods of use thereof”; and “Actriib proteins and variants and uses” published on April 29, 2014.Therefore, the BMP ligand traps disclosed in U.S. Patent No. 8710016, “relating to utrophin induction for muscular dystrophy therapy”, U.S. Patent Application Publication No. 2020 / 101134, “Methods for treating myeloproliferative neoplasm-associated myelofibrosis and anemia,” published on April 2, 2020, U.S. Patent Application Publication No. 2018 / 148491, “Novel Hybrid ActRIIB Ligand Trap Proteins For Treating Muscle Wasting Diseases,” published on May 31, 2018, and U.S. Patent No. 9884900, “Methods for treating janus kinase-associated disorders by administering soluble transforming growth factor beta type II receptor,” issued on February 6, 2018.

[0090] In some embodiments, the BMP antagonist is a deadBMP receptor. In some embodiments, the deadBMP receptor is a dominant-negative BMP receptor. In some embodiments, overexpression of the deadBMP receptor interferes with BMP-induced Smad activity. Any dominant-negative BMP receptor may be used herein, for example, Pouliot et al., Overexpression of a Dominant Negative Type II Bone Morphogenetic Protein Receptor Inhibits the Growth of Human Breast Cancer Cells, Cancer Res. 2003 Jan 15;63(2):277-81; Kawakami Y et al, BMP signaling during bone pattern determination in the developing limb. Development. 1996 Nov; 122(11):3557-66; Chen et al., Differential roles for bone morphogenetic protein (BMP) receptor type IB and IA in differentiation and specification of mesenchymal precursor cells to osteoblast and adipocyte lineages, J Cell Biol. 1998 Jul 13; 142(1):295-305.

[0091] In some embodiments, the HJV-induced BMP signaling antagonist of this disclosure is a hemoduvelin antagonist. In some embodiments, the hemoduvelin antagonist binds to one or more proteins of the repulsive guidance molecule (RGM) family, including RGMa, RGMb, and RGMc (HJV). In some embodiments, the hemoduvelin antagonist binds more selectively to hemoduvelin (RGMc) than to RGMa and RGMb. In some embodiments, the hemoduvelin antagonist is an antisense oligonucleotide that reduces hemoduvelin expression (see, for example, US7534764 of “Competitive regulation of hepcidin mRNA by soluble and cell-associated hemojuvelin” published on 19 May 2019, incorporated herein by reference; US2014127325 of “Competitive regulation of hepcidin mRNA by soluble and cell-associated hemojuvelin” published on 19 May 2009; and WO2016180784 of “Improved treatments using oligonucleotides” published on 17 November 2016). In some embodiments, the hemoduvelin antagonist is a small molecule compound that inhibits hemoduvelin, for example, by competitive binding and / or chemical modification of hemoduvelin.

[0092] In some embodiments, the HJV-induced BMP signaling antagonist is an HJV antagonist. In some embodiments, the HJV antagonist is soluble HJV. In some embodiments, the soluble HJV is a soluble HJV-Fc fusion protein.In some embodiments, soluble HJV is used in relation to U.S. Patent No. 8,318167B2, issued November 27, 2012, titled “Methods and compositions for regulating iron homeostasis by modulation of BMP-6”; U.S. Patent No. 9,708379B2, issued July 18, 2017, titled “COMPOSITIONS FOR REGULATING IRON HOMEOSTASIS AND METHODS OF USING SAME”; U.S. Patent No. 1,0273273B2, issued April 30, 2019, titled “COMPOSITIONS AND REGULATING IRON HOMEOSTASIS AND METHODS OF USING SAME”; and U.S. Patent No. 2, issued June 28, 2011, titled “METHODS AND COMPOSITIONS TO REGULATE IRON U.S. patent numbers include: U.S. No. 7,968091B2 for “METABOLISM”, U.S. No. 8,637023B2 for “HEMOJUVELIN FUSION PROTEINS” issued on January 28, 2014, U.S. No. 8,865168B2 for “METHODS AND COMPOSITIONS TO REGULATE HEPCIDIN EXPRESSION” issued on October 21, 2014, U.S. No. 9,556251B2 for “METHODS AND COMPOSITIONS TO REGULATE HEPCIDIN EXPRESSION” issued on January 31, 2017; U.S. No. 8,895002B2 for “Hemojuvelin fusion proteins and uses thereof” issued on November 25, 2014; and “Juvenile hemochromatosis gene (HFE2A) cleavage products and uses” issued on March 31, 2009. It is soluble HJV as disclosed in U.S. Patent No. 7511018B2. In some embodiments, the sHJV-Fc fusion protein is Ferruxmax. In some embodiments, the sHJV-Fc fusion protein is FMX-8.

[0093] In some embodiments, the hemoduvelin antagonist is a hemoduvelin-specific antibody and / or one or more proteins from the RGM protein family (e.g., RGMa, RGMb). In some embodiments, hemomodoverin-specific antibodies and / or one or more RGM proteins are incorporated herein by reference by the following patents: U.S. Patent No. 10118958, issued November 6, 2018, titled “Composition and method for the diagnosis and treatment of iron-related disorders”; U.S. Patent No. 9636398, issued May 2, 2017, titled “Composition and method for the diagnosis and treatment of iron-related disorders”; and U.S. Patent No. 8507435, issued August 13, 2013, titled “Juvenile hemochromatosis gene (HFE2A) cleavage products and uses thereof”; U.S. Patent No. 10118958, issued November 6, 2018, titled “Composition and method for the diagnosis and treatment of iron-related disorders”; and U.S. Patent No. 8507435, published December 23, 2010, titled “Bone morphogenetic protein (BMP)-binding domains of proteins of the U.S. Patent Application Publication No. 2010 / 0322941 for “repulsive guidance molecule (RGM) protein family and functional fragments thereof, and use of same”; U.S. Patent No. 9040052 issued on May 26, 2015, for “Precision Medicine By Targeting Rare Human PCSK9 Variants for Cholesterol Treatment”;and U.S. Patent Application Publication No. 2017 / 0029499, published on February 2, 2017, with the title "Methods for treating hepcidin-mediated disorders"; and International Publication No. 2007039256, published on April 12, 2007, with the title "Binding domains of proteins of the repulsive guidance molecule (rgm) protein family and functional fragments thereof, and their use"; International Publication No. 2015171691, published on November 12, 2015, with the title "Compositions and methods for growth factor modulation"; International Publication No. 2018 / 009624, published on January 11, 2018, with the title "Tgf-beta superfamily heteromultimers and uses thereof"; and "Rgmc-selective inhibitors and use" published on April 30, 2020. The anti-HJV antibody and / or one or more RGM proteins disclosed in International Publication No. 2020 / 086736 of “them”.

[0094] In some embodiments, the anti-HJV antibody is one of the anti-HJV antibodies listed in Table 1. Table 1 contains exemplary amino acid sequences of CDRs of anti-HJV antibodies. In some embodiments, the HJV antagonist of this application is an anti-HJV antibody comprising a CDR containing an amino acid sequence selected from Table 1.

[0095] In some embodiments, the anti-HJV antibody of this disclosure comprises one or more heavy chain CDR (e.g., CDR-H1, CDR-H2, or CDR-H3) amino acid sequences from any one of the anti-HJV antibodies selected from Table 1. In some embodiments, the anti-HJV antibody of this disclosure comprises CDR-H1, CDR-H2, and CDR-H3 according to any one of the antibodies selected from Table 1. In some embodiments, the anti-HJV antibody of this disclosure comprises one or more light chain CDR (e.g., CDR-L1, CDR-L2, or CDR-L3) amino acid sequences from any one of the anti-HJV antibodies selected from Table 1. In some embodiments, the anti-HJV antibody of this disclosure comprises CDR-L1, CDR-L2, and CDR-L3 according to any one of the anti-HJV antibodies selected from Table 1.

[0096] In some embodiments, the anti-HJV antibodies of this disclosure comprise CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and CDR-L3 according to any one of the anti-HJV antibodies selected from Table 1. In some embodiments, the antibody heavy chain and light chain CDR3 domains may play a particularly important role in the antibody's binding specificity / affinity to the antigen. Therefore, the anti-HJV antibodies of this disclosure may comprise at least the heavy chain and / or light chain CDR3 of any one of the anti-HJV antibodies selected from Table 1.

[0097] Also within the scope of this disclosure are any exemplary functional variants of anti-HJV antibodies disclosed herein. A functional variant is a functional variant compared to a reference antibody. H and / or V LThe heavy chain CDR may contain one or more amino acid residue variations, but may retain substantially similar binding and biological activity (e.g., substantially similar binding affinity, binding specificity, inhibitory activity, anti-inflammatory activity, or a combination thereof) to the reference antibody. In some embodiments, the functional variants of the anti-HJV antibody described herein contain one or more amino acid variations in the heavy chain CDR and / or one or more amino acid variations in the light chain CDR compared to the reference antibody, but retain substantially similar binding and biological activity (e.g., substantially similar binding affinity, binding specificity, inhibitory activity, anti-inflammatory activity, or a combination thereof). In some embodiments, the functional variants of the anti-HJV antibody described herein contain one or more amino acid variations in the heavy chain framework region and / or one or more amino acid variations in the light chain framework region compared to the reference antibody, but retain substantially similar binding and biological activity (e.g., substantially similar binding affinity, binding specificity, inhibitory activity, anti-inflammatory activity, or a combination thereof). When used herein in the context of function (e.g., binding affinity and / or biological function), substantially, it means that an antibody variant (e.g., an anti-HJV antibody variant) has at least 80%, at least 85%, at least 90%, at least 91%, at least 91%, at least 91%, at least 91%, at least 91%, at least 91%, at least 91%, at least 91%, or at least 100% of the function (e.g., binding affinity and / or biological function) compared to a reference antibody (any anti-HJV antibody listed in Tables 1 and 2).

[0098] In some embodiments, any anti-HJV antibody of this disclosure has one or more CDR (e.g., heavy chain CDR or light chain CDR) sequences substantially similar to any of the CDR-H1, CDR-H2, CDR-H3, CDR-L1, CDR-L2, and / or CDR-L3 sequences from one of the anti-HJV antibodies selected from Table 1. In some embodiments, the positions of one or more CDRs in the VH (e.g., CDR-H1, CDR-H2, or CDR-H3) and / or VL (e.g., CDR-L1, CDR-L2, or CDR-L3) regions of the antibodies described herein may vary by one, two, three, four, five, or six amino acid positions, as long as immunospecific binding to hemoduverin (e.g., human hemoduverin) is maintained (e.g., substantially maintaining at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it originates). For example, in some embodiments, the position defining the CDR of any antibody described herein may be altered by shifting the N-terminal and / or C-terminal boundary of the CDR by one, two, three, four, five, or six amino acids compared to any one CDR position of any antibody described herein, as long as immunospecific binding to hemoduvelin (e.g., human hemoduvelin) is maintained (e.g., substantially maintaining at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it originates). In another embodiment, the length of one or more CDRs in the VH (e.g., CDR-H1, CDR-H2, or CDR-H3) and / or VL (e.g., CDR-L1, CDR-L2, or CDR-L3) regions of the antibodies described herein may vary by one, two, three, four, five, or more amino acids (e.g., short or long), as long as immunospecific binding to hemoduvelin (e.g., human hemoduvelin) is maintained (e.g., substantially maintained at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived).

[0099] Therefore, in some embodiments, the HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 described herein may have one, two, three, four, five, or more amino acids shorter than one or more of the CDRs described herein (e.g., CDRs from any of the anti-HJV antibodies selected from Table 1), as long as immunospecific binding to hemoduvelin (e.g., human hemoduvelin) is maintained (e.g., substantially maintained by at least 80%, at least 90%, or at least 95% compared to binding to the original antibody from which it is derived). In some embodiments, the HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 described herein may have one, two, three, four, five, or more amino acids longer than one or more of the CDRs described herein (e.g., CDRs from any of the anti-HJV antibodies selected from Table 1), provided that immunospecific binding to hemoduvelin (e.g., human hemoduvelin) is maintained (e.g., substantially maintained by at least 80%, at least 90%, or at least 95% compared to binding to the original antibody from which it is derived). In some embodiments, the amino portions of HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 described herein may have one, two, three, four, five, or more amino acids extended compared to one or more CDRs described herein (e.g., CDRs from any of the anti-HJV antibodies selected from Table 1), provided that immunospecific binding to hemoduvelin (e.g., human hemoduvelin) is maintained (e.g., substantially maintained by at least 80%, at least 90%, or at least 95% compared to binding to the original antibody from which it is derived).In some embodiments, the carboxyl portions of HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 described herein may be elongated by one, two, three, four, five, or more amino acids compared to one or more CDRs described herein (e.g., CDRs from any of the anti-HJV antibodies selected from Table 1), provided that immunospecific binding to hemoduvelin (e.g., human hemoduvelin) is maintained (e.g., substantially maintained by at least 80%, at least 90%, or at least 95% compared to binding to the original antibody from which it is derived). In some embodiments, the amino portions of HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 described herein may be shortened by one, two, three, four, five, or more amino acids compared to one or more CDRs described herein (e.g., CDRs from any of the anti-HJV antibodies selected from Table 1), provided that immunospecific binding to hemoduvelin (e.g., human hemoduvelin) is maintained (e.g., substantially maintained by at least 80%, at least 90%, or at least 95% compared to binding to the original antibody from which it is derived). In some embodiments, the carboxyl portions of HC CDR1, HC CDR2, HC CDR3, LC CDR1, LC CDR2, and / or LC CDR3 described herein may be shortened by one, two, three, four, five, or more amino acids compared to one or more CDRs described herein (e.g., CDRs from any of the anti-HJV antibodies selected from Table 1), provided that immunospecific binding to hemoduvelin (e.g., human hemoduvelin) is maintained (e.g., substantially maintained by at least 80%, at least 90%, or at least 95% compared to binding to the original antibody from which it is derived). Any suitable known method can be used to determine whether immunospecific binding to hemoduvelin (e.g., human hemoduvelin) is maintained, for example, by using binding assays and conditions described in the Art.

[0100] In some examples, any anti-HJV antibody of this disclosure has one or more CDR (e.g., HC CDR or LC CDR) sequences substantially similar to any one of the anti-HJV antibodies selected from Table 1. For example, an antibody may contain one or more CDR sequences from any anti-HJV antibody selected from Table 1, containing up to five, four, three, two, or one amino acid residue variation compared to the corresponding CDR region of any one of the CDRs provided herein (e.g., a CDR from any of the anti-HJV antibodies selected from Table 1), provided that immunospecific binding to hemoduverin (e.g., human hemoduverin) is maintained (e.g., substantially maintained by at least 80%, at least 90%, or at least 95% compared to the binding of the original antibody from which it is derived). In some embodiments, any amino acid variation in any CDR provided herein may be a conserved variation. A conserved variation may be introduced into the CDR at a position where the residue would not be involved in interaction with the hemoduverin protein (e.g., human hemoduverin protein) if determined, for example, based on the crystal structure. Some aspects of this disclosure provide anti-HJV antibodies comprising one or more heavy-chain variable (VH) and / or light-chain variable (VL) domains provided herein. In some embodiments, any VH domain provided herein comprises one or more HC CDR sequences provided herein (e.g., HC CDR1, HC CDR2, and HC CDR3), for example, any CDR-H sequence provided for any one of the anti-HJVs selected from Table 1. In some embodiments, any VL domain provided herein comprises one or more CDR-L sequences provided herein (e.g., LC CDR1, LC CDR2, and LC CDR3), for example, any LC CDR sequence provided for any one of the anti-HJV antibodies selected from Table 1.

[0101] In some embodiments, the anti-HJV antibody includes a variable heavy chain region comprising CDR1 containing the amino acid sequence of SEQ ID NO: 1, CDR2 containing the amino acid sequence of SEQ ID NO: 2, and CDR3 containing the amino acid sequence of SEQ ID NO: 3. In some embodiments, the anti-HJV antibody includes a variable light chain region comprising CDR1 containing an amino acid sequence selected from any one of SEQ ID NOs: 4, 7, 10, 13, and 16, CDR2 containing an amino acid sequence selected from any one of SEQ ID NOs: 5, 8, 11, 14, and 17, and CDR3 containing an amino acid sequence selected from any one of SEQ ID NOs: 6, 9, 12, 15, and 18.

[0102] In some embodiments, the anti-HJV antibody includes a variable heavy chain region comprising CDR1 containing the amino acid sequence of SEQ ID NO: 19, CDR2 containing the amino acid sequence of SEQ ID NO: 20, and CDR3 containing the amino acid sequence of SEQ ID NO: 21. In some embodiments, the anti-HJV antibody includes a variable light chain region comprising CDR1 containing the amino acid sequence of SEQ ID NO: 22, CDR2 containing the amino acid sequence of SEQ ID NO: 23, and CDR3 containing the amino acid sequence of SEQ ID NO: 24.

[0103] [Table 2]

[0104] In some embodiments, the anti-HJV antibody of this disclosure comprises any antibody comprising the heavy chain variable domain and / or light chain variable domain of any one anti-HJV antibody selected from Table 2, as well as any variants thereof. In some embodiments, the anti-HJV antibody of this disclosure comprises any antibody comprising the heavy chain variable and light chain variable pairs of any anti-HJV antibody selected from Table 2.

[0105] Aspects of this disclosure provide anti-HJV antibodies having heavy-chain variable (VH) and / or light-chain variable (VL) domain amino acid sequences homologous to any of those described herein. In some embodiments, the anti-HJV antibody includes a heavy-chain variable sequence or a light-chain variable sequence that is at least 75% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to any one of the heavy-chain variable sequences and / or light-chain variable sequences of any anti-HJV antibody selected from Table 2. In some embodiments, the homologous heavy-chain variable and / or light-chain variable amino acid sequences are not different within any of the CDR sequences provided herein. For example, in some embodiments, the degree of sequence variation (e.g., 75%, 80%, 85%, 90%, 95%, 98%, or 99%) may occur within the heavy-chain variable and / or light-chain variable sequences other than any of the CDR sequences provided herein. In some embodiments, any anti-HJV antibody provided herein includes a heavy chain variable sequence and a light chain variable sequence that include a framework sequence which is at least 75%, 80%, 85%, 90%, 95%, 98%, or 99% identical to the framework sequence of any anti-HJV antibody selected from Table 2.

[0106] Table 2 contains exemplary amino acid sequences of variable heavy and variable light chains of anti-HJV antibodies. In some embodiments, the hepcidin antagonist of this application is an anti-HJV antibody comprising a variable heavy and / or variable light chain comprising an amino acid sequence selected from Table 2.

[0107] In some embodiments, the anti-HJV antibody of the present disclosure comprises heavy chain variable domains CDR-H1, CDR-H2, and CDR-H3 having the amino acid sequence of SEQ ID NO: 25. Alternatively, the anti-HJV antibody of the present disclosure further comprises light chain variable domains CDR-L1, CDR-L2, and CDR-L3 having the amino acid sequence of SEQ ID NO: 26.

[0108] In some embodiments, the Kabat definition system defines the anti-HJV antibody of this disclosure as comprising CDR-H1 having the amino acid sequence of SEQ ID NO: 1, CDR-H2 having the amino acid sequence of SEQ ID NO: 2, CDR-H3 having the amino acid sequence of SEQ ID NO: 3; and / or CDR-L1 having the amino acid sequence of SEQ ID NO: 4, CDR-L2 having the amino acid sequence of SEQ ID NO: 5, and CDR-L3 having the amino acid sequence of SEQ ID NO: 6.

[0109] In some embodiments, the anti-HJV antibody includes a variable heavy chain region containing the amino acid sequence of SEQ ID NO: 25 and / or a variable light chain region containing the amino acid sequence of SEQ ID NO: 26.

[0110] In some embodiments, the anti-HJV antibody of the present disclosure includes a VH containing 20 or fewer amino acid variations compared to the VH described in SEQ ID NO: 25 (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations). Alternatively, the anti-HJV antibody of the present disclosure includes a VL containing 20 or fewer amino acid variations compared to the VL described in SEQ ID NO: 26 (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations). In some embodiments, the anti-HJV antibody of this disclosure comprises a VH having an amino acid sequence that is at least 80% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to the VH described in SEQ ID NO: 25. Alternatively, the anti-HJV antibody of this disclosure comprises a VL having an amino acid sequence that is at least 80% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to the VL described in SEQ ID NO: 26. In some embodiments, the function of the anti-HJV antibody having amino acid variations in the FR region of the VH (e.g., based on the Kabat definition) compared to the VH described in SEQ ID NO: 25 is maintained (e.g., substantially maintaining at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived). In some embodiments, the function of an anti-HJV antibody having five or fewer amino acid variations (e.g., five, four, three, two, or one) or three or fewer amino acid variations (e.g., three, two, or one) in the FR region of VH (e.g., based on the Kabat definition) compared to VH described in SEQ ID NO: 25 is maintained (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained). Alternatively, in some embodiments, the function of an anti-HJV antibody having amino acid variations in the FR region of VL (e.g., based on the Kabat definition) compared to VL described in SEQ ID NO: 26 is maintained (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained).In some embodiments, the function of an anti-HJV antibody having five or fewer amino acid variations (e.g., five, four, three, two, or one or fewer) or three or fewer amino acid variations (e.g., three, two, or one or fewer) in the FR region of the VL (e.g., based on the Kabat definition) is maintained compared to the VL described in SEQ ID NO: 26 (e.g., at least 80%, at least 90%, and at least 95% of the binding of the original antibody from which it is derived is substantially maintained).

[0111] In some embodiments, the anti-HJV antibody of the present disclosure comprises heavy chain variable domains CDR-H1, CDR-H2, and CDR-H3 having the amino acid sequence of SEQ ID NO: 27. Alternatively, the anti-HJV antibody of the present disclosure comprises light chain variable domains CDR-L1, CDR-L2, and CDR-L3 having the amino acid sequence of SEQ ID NO: 28.

[0112] In some embodiments, the anti-HJV antibody of the present disclosure is defined by the Kabat definition system as including CDR-H1 having the amino acid sequence of SEQ ID NO: 1, CDR-H2 having the amino acid sequence of SEQ ID NO: 2, CDR-H3 having the amino acid sequence of SEQ ID NO: 3; and / or CDR-L1 having the amino acid sequence of SEQ ID NO: 7, CDR-L2 having the amino acid sequence of SEQ ID NO: 8, and CDR-L3 having the amino acid sequence of SEQ ID NO: 9.

[0113] In some embodiments, the anti-HJV antibody includes a variable heavy chain region containing the amino acid sequence of SEQ ID NO: 27 and / or a variable light chain region containing the amino acid sequence of SEQ ID NO: 28.

[0114] In some embodiments, the anti-HJV antibody of the present disclosure includes a VH containing 20 or fewer amino acid variations compared to the VH described in SEQ ID NO: 27 (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations). Alternatively, the anti-HJV antibody of the present disclosure includes a VL containing 20 or fewer amino acid variations compared to the VL described in SEQ ID NO: 28 (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations). In some embodiments, the anti-HJV antibody of this disclosure comprises a VH having an amino acid sequence that is at least 80% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to the VH described in SEQ ID NO: 27. Alternatively, the anti-HJV antibody of this disclosure comprises a VL having an amino acid sequence that is at least 80% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to the VL described in SEQ ID NO: 28. In some embodiments, the function of the anti-HJV antibody having amino acid variations in the FR region of the VH (e.g., based on the Kabat definition) compared to the VH described in SEQ ID NO: 27 is maintained (e.g., at least 80%, at least 90%, at least 95% of the binding of the original antibody from which it is derived is substantially maintained). In some embodiments, the function of an anti-HJV antibody having five or fewer amino acid variations (e.g., five, four, three, two, or one) or three or fewer amino acid variations (e.g., three, two, or one) in the FR region of VH (e.g., based on the Kabat definition) compared to VH described in SEQ ID NO: 27 is maintained (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained). Alternatively, in some embodiments, the function of an anti-HJV antibody having amino acid variations in the FR region of VL (e.g., based on the Kabat definition) compared to VL described in SEQ ID NO: 28 is maintained (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained).In some embodiments, the function of an anti-HJV antibody having five or fewer amino acid variations (e.g., five, four, three, two, or one or fewer) or three or fewer amino acid variations (e.g., three, two, or one or fewer) in the FR region of the VL (e.g., based on the Kabat definition) is maintained compared to the VL described in SEQ ID NO: 28 (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained).

[0115] In some embodiments, the anti-HJV antibody of the present disclosure comprises heavy chain variable domains CDR-H1, CDR-H2, and CDR-H3 having the amino acid sequence of SEQ ID NO: 29. Alternatively, the anti-HJV antibody of the present disclosure further comprises light chain variable domains CDR-L1, CDR-L2, and CDR-L3 having the amino acid sequence of SEQ ID NO: 30.

[0116] In some embodiments, the Kabat definition system defines the anti-HJV antibody of this disclosure as comprising CDR-H1 having the amino acid sequence of SEQ ID NO: 1, CDR-H2 having the amino acid sequence of SEQ ID NO: 2, CDR-H3 having the amino acid sequence of SEQ ID NO: 3; and / or CDR-L1 having the amino acid sequence of SEQ ID NO: 10, CDR-L2 having the amino acid sequence of SEQ ID NO: 11, and CDR-L3 having the amino acid sequence of SEQ ID NO: 12.

[0117] In some embodiments, the anti-HJV antibody includes a variable heavy chain region containing the amino acid sequence of SEQ ID NO: 29 and / or a variable light chain region containing the amino acid sequence of SEQ ID NO: 30.

[0118] In some embodiments, the anti-HJV antibody of the present disclosure includes a VH containing 20 or fewer amino acid variations compared to the VH described in SEQ ID NO: 29 (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations). Alternatively, the anti-HJV antibody of the present disclosure includes a VL containing 20 or fewer amino acid variations compared to the VL described in SEQ ID NO: 30 (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations). In some embodiments, the anti-HJV antibody of this disclosure comprises a VH having an amino acid sequence that is at least 80% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to the VH described in SEQ ID NO: 29. Alternatively, the anti-HJV antibody of this disclosure comprises a VL having an amino acid sequence that is at least 80% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to the VL described in SEQ ID NO: 30. In some embodiments, the function of the anti-HJV antibody having amino acid variations in the FR region of the VH (e.g., based on the Kabat definition) compared to the VH described in SEQ ID NO: 29 is maintained (e.g., at least 80%, at least 90%, at least 95% of the binding of the original antibody from which it is derived is substantially maintained). In some embodiments, the function of an anti-HJV antibody having five or fewer amino acid variations (e.g., five, four, three, two, or one) or three or fewer amino acid variations (e.g., three, two, or one) in the FR region of VH (e.g., based on the Kabat definition) compared to VH described in SEQ ID NO: 29 is maintained (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained). Alternatively, in some embodiments, the function of an anti-HJV antibody having amino acid variations in the FR region (e.g., based on the Kabat definition) of VL compared to VL described in SEQ ID NO: 30 is maintained (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained).In some embodiments, the function of an anti-HJV antibody having five or fewer amino acid variations (e.g., five, four, three, two, or one or fewer) or three or fewer amino acid variations (e.g., three, two, or one or fewer) in the FR region of the VL (e.g., based on the Kabat definition) is maintained compared to the VL described in SEQ ID NO: 30 (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained).

[0119] In some embodiments, the anti-HJV antibody of the present disclosure comprises heavy chain variable domains CDR-H1, CDR-H2, and CDR-H3 having the amino acid sequence of SEQ ID NO: 31. Alternatively, the anti-HJV antibody of the present disclosure comprises light chain variable domains CDR-L1, CDR-L2, and CDR-L3 having the amino acid sequence of SEQ ID NO: 32.

[0120] In some embodiments, the Kabat definition system defines the anti-HJV antibody of this disclosure as comprising CDR-H1 having the amino acid sequence of SEQ ID NO: 1, CDR-H2 having the amino acid sequence of SEQ ID NO: 2, CDR-H3 having the amino acid sequence of SEQ ID NO: 3; and / or CDR-L1 having the amino acid sequence of SEQ ID NO: 13, CDR-L2 having the amino acid sequence of SEQ ID NO: 14, and CDR-L3 having the amino acid sequence of SEQ ID NO: 15.

[0121] In some embodiments, the anti-HJV antibody includes a variable heavy chain region containing the amino acid sequence of SEQ ID NO: 31 and / or a variable light chain region containing the amino acid sequence of SEQ ID NO: 32.

[0122] In some embodiments, the anti-HJV antibody of the present disclosure comprises a VH containing 20 or fewer amino acid variations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations) compared to the VH described in SEQ ID NO: 31. Alternatively, the anti-HJV antibody of the present disclosure comprises a VL containing 20 or fewer amino acid variations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations) compared to the VL described in SEQ ID NO: 32. In some embodiments, the anti-HJV antibody of this disclosure comprises a VH having an amino acid sequence that is at least 80% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to the VH described in SEQ ID NO: 31. Alternatively, the anti-HJV antibody of this disclosure comprises a VL having an amino acid sequence that is at least 80% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to the VL described in SEQ ID NO: 32. In some embodiments, the function of the anti-HJV antibody having amino acid variations in the FR region of the VH (e.g., based on the Kabat definition) compared to the VH described in SEQ ID NO: 31 is maintained (e.g., at least 80%, at least 90%, at least 95% of the binding of the original antibody from which it is derived is substantially maintained). In some embodiments, the function of an anti-HJV antibody having five or fewer amino acid variations (e.g., five, four, three, two, or one) or three or fewer amino acid variations (e.g., three, two, or one) in the FR region of VH (e.g., based on the Kabat definition) compared to VH described in SEQ ID NO: 31 is maintained (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained). Alternatively, in some embodiments, the function of an anti-HJV antibody having amino acid variations in the FR region of VL (e.g., based on the Kabat definition) compared to VL described in SEQ ID NO: 32 is maintained (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained).In some embodiments, the function of an anti-HJV antibody having five or fewer amino acid variations (e.g., five, four, three, two, or one or fewer) or three or fewer amino acid variations (e.g., three, two, or one or fewer) in the FR region of the VL (e.g., based on the Kabat definition) is maintained compared to the VL described in SEQ ID NO: 32 (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained).

[0123] In some embodiments, the anti-HJV antibody of the present disclosure comprises heavy chain variable domains CDR-H1, CDR-H2, and CDR-H3 having the amino acid sequence of SEQ ID NO: 33. Alternatively, the anti-HJV antibody of the present disclosure comprises light chain variable domains CDR-L1, CDR-L2, and CDR-L3 having the amino acid sequence of SEQ ID NO: 34.

[0124] In some embodiments, the Kabat definition system defines the anti-HJV antibody of this disclosure as comprising CDR-H1 having the amino acid sequence of SEQ ID NO: 1, CDR-H2 having the amino acid sequence of SEQ ID NO: 2, CDR-H3 having the amino acid sequence of SEQ ID NO: 3; and / or CDR-L1 having the amino acid sequence of SEQ ID NO: 16, CDR-L2 having the amino acid sequence of SEQ ID NO: 17, and CDR-L3 having the amino acid sequence of SEQ ID NO: 18.

[0125] In some embodiments, the anti-HJV antibody includes a variable heavy chain region containing the amino acid sequence of SEQ ID NO: 33 and / or a variable light chain region containing the amino acid sequence of SEQ ID NO: 34.

[0126] In some embodiments, the anti-HJV antibody of the present disclosure includes a VH containing 20 or fewer amino acid variations compared to the VH described in SEQ ID NO: 33 (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations). Alternatively, the anti-HJV antibody of the present disclosure includes a VL containing 20 or fewer amino acid variations compared to the VL described in SEQ ID NO: 34 (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations). In some embodiments, the anti-HJV antibody of this disclosure comprises a VH having an amino acid sequence that is at least 80% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to the VH described in SEQ ID NO: 33. Alternatively, the anti-HJV antibody of this disclosure comprises a VL having an amino acid sequence that is at least 80% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to the VL described in SEQ ID NO: 34. In some embodiments, the function of the anti-HJV antibody having amino acid variations in the FR region of the VH (e.g., based on the Kabat definition) compared to the VH described in SEQ ID NO: 33 is maintained (e.g., at least 80%, at least 90%, at least 95% of the binding of the original antibody from which it is derived is substantially maintained). In some embodiments, the function of an anti-HJV antibody having five or fewer amino acid variations (e.g., five, four, three, two, or one) or three or fewer amino acid variations (e.g., three, two, or one) in the FR region of VH (e.g., based on the Kabat definition) compared to VH described in SEQ ID NO: 33 is maintained (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained). Alternatively, in some embodiments, the function of an anti-HJV antibody having amino acid variations in the FR region of VL (e.g., based on the Kabat definition) compared to VL described in SEQ ID NO: 34 is maintained (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained).In some embodiments, the function of an anti-HJV antibody having five or fewer amino acid variations (e.g., five, four, three, two, or one or fewer) or three or fewer amino acid variations (e.g., three, two, or one or fewer) in the FR region of the VL (e.g., based on the Kabat definition) is maintained compared to the VL described in SEQ ID NO: 34 (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained).

[0127] In some embodiments, the anti-HJV antibody of the present disclosure comprises heavy chain variable domains CDR-H1, CDR-H2, and CDR-H3 having the amino acid sequence of SEQ ID NO: 35. Alternatively, the anti-HJV antibody of the present disclosure further comprises light chain variable domains CDR-L1, CDR-L2, and CDR-L3 having the amino acid sequence of SEQ ID NO: 36.

[0128] In some embodiments, the Kabat definition system defines the anti-HJV antibody of this disclosure as comprising CDR-H1 having the amino acid sequence of SEQ ID NO: 19, CDR-H2 having the amino acid sequence of SEQ ID NO: 20, CDR-H3 having the amino acid sequence of SEQ ID NO: 21; and / or CDR-L1 having the amino acid sequence of SEQ ID NO: 22, CDR-L2 having the amino acid sequence of SEQ ID NO: 23, and CDR-L3 having the amino acid sequence of SEQ ID NO: 24.

[0129] In some embodiments, the anti-HJV antibody includes a variable heavy chain region containing the amino acid sequence of SEQ ID NO: 35 and / or a variable light chain region containing the amino acid sequence of SEQ ID NO: 36.

[0130] In some embodiments, the anti-HJV antibody of the present disclosure comprises a VH containing 20 or fewer amino acid variations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations) compared to the VH described in SEQ ID NO: 35. Alternatively, the anti-HJV antibody of the present disclosure comprises a VL containing 20 or fewer amino acid variations (e.g., 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 9, 8, 7, 6, 5, 4, 3, 2, or 1 or fewer amino acid variations) compared to the VL described in SEQ ID NO: 36. In some embodiments, the anti-HJV antibody of this disclosure comprises a VH having an amino acid sequence that is at least 80% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to the VH described in SEQ ID NO: 35. Alternatively, the anti-HJV antibody of this disclosure comprises a VL having an amino acid sequence that is at least 80% (e.g., 80%, 85%, 90%, 95%, 98%, or 99%) identical to the VL described in SEQ ID NO: 36. In some embodiments, the function of the anti-HJV antibody having amino acid variations in the FR region of the VH (e.g., based on the Kabat definition) compared to the VH described in SEQ ID NO: 35 is maintained (e.g., at least 80%, at least 90%, at least 95% of the binding of the original antibody from which it is derived is substantially maintained). In some embodiments, the function of an anti-HJV antibody having five or fewer amino acid variations (e.g., five, four, three, two, or one) or three or fewer amino acid variations (e.g., three, two, or one) in the FR region of VH (e.g., based on the Kabat definition) compared to VH described in SEQ ID NO: 35 is maintained (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained). Alternatively, in some embodiments, the function of an anti-HJV antibody having amino acid variations in the FR region of VL (e.g., based on the Kabat definition) compared to VL described in SEQ ID NO: 36 is maintained (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained).In some embodiments, the function of an anti-HJV antibody having five or fewer amino acid variations (e.g., five, four, three, two, or one or fewer) or three or fewer amino acid variations (e.g., three, two, or one or fewer) in the FR region of the VL (e.g., based on the Kabat definition) is maintained compared to the VL described in SEQ ID NO: 36 (e.g., at least 80%, at least 90%, or at least 95% of the binding of the original antibody from which it is derived is substantially maintained).

[0131] [Table 3]

[0132] In some embodiments, the anti-HJV antibodies described herein can inhibit hepcidin expression by blocking the BMP signaling pathway (e.g., the BMP6 signaling pathway). In some embodiments, the anti-HJV antibodies described herein can inhibit hepcidin expression by blocking the JAK / STAT pathway (e.g., the IL-6 signaling pathway). In some embodiments, the anti-HJV antibodies described herein can inhibit hepcidin expression by blocking both the BMP signaling pathway (e.g., the BMP6 signaling pathway) and the JAK-STAT pathway (e.g., the IL-6 signaling pathway).

[0133] In some embodiments, the HJV antagonist is an inhibitory nucleic acid that targets HJV (e.g., HJV-targeting dsRNA, siRNA, miRNA, shRNA, AmiRNA, antisense oligonucleotide (ASO), or aptamer). In some embodiments, HJV-targeting inhibitory nucleic acids may be used herein in the treatment of myelofibrosis and related conditions.

[0134] In some embodiments, the anti-HJV antagonist is recombinant matryptase-2 (TMPRSS6). Matryptase-2 is a transmembrane serine protease capable of cleaving HJV, and overexpression of the matryptase-2 protein in cells suppresses the activation of hepcidin expression (Du X, She E, Gelbart T, et al. The serine protease TMPRSS6 is required to sense iron deficiency, Science, 2008, vol. 320 5879 (pg. 1088-1092)).

[0135] In some embodiments, upon ligand binding, the constitutively active type II receptor phosphorylates the type I receptor, which then phosphorylates intracellular receptor-activating Smad (R-Smad), primarily Smad1, Smad5, and / or Smad8. In such embodiments, activated R-Smad forms a complex with a common partner, Smad4, and translocates to the nucleus to regulate gene transcription, e.g., induction of hepcidin expression. In some embodiments, the HJV-induced BMP signaling antagonist is an intracellular inhibitor of R-Smad (e.g., Smad1, Smad5, and Smad8) and its partner Smad4. In some embodiments, the intracellular inhibitors of R-Smad and Smad4 are intracellular antibodies.

[0136] In some embodiments, the intracellular inhibitor of Smad is an inhibitory nucleic acid that targets R-Smad (e.g., Smad1, Smad5, or Smad8) and Smad4. In some embodiments, the inhibitory nucleic acid that targets R-Smad (e.g., Smad1, Smad5, or Smad8) and Smad4 is an inhibitory RNA. In some embodiments, the inhibitory nucleic acid is a miRNA that targets R-Smad (e.g., Smad1, Smad5, or Smad8) and Smad4. In some embodiments, the inhibitory nucleic acid is an shRNA that targets R-Smad (e.g., Smad1, Smad5, or Smad8) and Smad4. In some embodiments, the inhibitory nucleic acid is an siRNA that targets R-Smad (e.g., Smad1, Smad5, or Smad8) and Smad4. In some embodiments, the inhibitory nucleic acid is an AmiRNA that targets R-Smad (e.g., Smad1, Smad5, or Smad8) and Smad4.

[0137] In some embodiments, the intracellular inhibitors of R-Smad and Smad4 are recombination inhibitory Smad (I-Smad) (e.g., Smad6 or Smad7). In some embodiments, Smad6 preferentially inhibits Smad signaling initiated by bone morphogenetic protein (BMP) type I receptors ALK-3 and ALK-6, while Smad7 inhibits both transforming growth factor β (TGF-β) and BMP-induced Smad signaling.

[0138] Efficient iron signaling via the BMP-SMAD signaling pathway involves cofactors such as diiron-transferrin (Tf) sensor transferrin receptor (TfR) that stimulate hepcidin expression (Figure 2). In some embodiments, the hepcidin antagonist of this disclosure is a transferrin antagonist that antagonistizes hepcidin function by binding to transferrin and / or transferrin receptor 2, thereby inhibiting activation of the BMP-SMAD signaling pathway. In some embodiments, the transferrin antagonist is an antisense oligonucleotide that targets Tf and / or TfR, such as siTFR2 (see, for example, U.S. Patent No. 9,228,188 incorporated herein by reference).

[0139] ii. Other hepcidin antagonists In some embodiments, a hepcidin antagonist is a hepcidin antagonist. A hepcidin antagonist refers to a drug that directly neutralizes hepcidin. In some embodiments, the hepcidin antagonists of this disclosure are drugs that bind to HAMP or their transcripts or translation products. Examples of such hepcidin antagonists include, but are not limited to, antisense oligonucleotides, small molecule inhibitor compounds, and antibodies, antikalins, or aptamers that are specific to HAMP transcripts or translation products (e.g., hepcidin).

[0140] In some embodiments, the hepcidin antagonist is a hepcidin inhibitor. In some embodiments, the hepcidin inhibitor is a molecule that specifically binds to hepcidin (e.g., an antibody, antikalin, or aptamer). Examples of molecules that specifically bind to hepcidin include, but are not limited to, PRS-080, LY2787106, NOX-H94 (Lexaptepid Pegol), 12B9m, LS-B4534, lipocalin mutant proteins, and hNGAL mutant proteins (see also U.S. Patent Nos. 8,629250; 9315577; 9051382; 9657098; 9610356; 8530619, and U.S. Patent Publication Nos. 2015 / 0291675, 2018 / 0057812, and 2017 / 0247448, incorporated herein by reference).

[0141] In some embodiments, the hepcidin neutralizer is an anti-hepcidin antibody. In some embodiments, the anti-hepcidin antibody is incorporated herein by reference by U.S. Patent No. 1,0323088B2, titled “Humanized anti-hepcidin antibodies and uses thereof,” issued on 31 August 2017; U.S. Patent No. 8,609817B2, titled “Anti-hepcidin-25 selective antibodies and uses thereof,” issued on 17 December 2013; U.S. Patent No. 8,304258B2, titled “Methods of producing monoclonal antibodies specific for human hepcidin,” issued on 6 November 2012; U.S. Patent No. 8,629250B2, titled “Hepcidin, hepcidin antagonists and methods of use,” issued on 14 January 2014; and U.S. Patent No. 8,629250B2, titled “Anti-hepcidin antibodies and uses,” issued on 23 May 2017. These are anti-hepcidin antibodies as described in U.S. Patent No. 9,657,098,B2, titled "Anti-hepcidin antibodies and uses thereof," issued on October 31, 2017, U.S. Patent No. 9,803,011,B2, titled "Anti-hepcidin antibodies and uses thereof," issued on March 26, 2019, or U.S. Patent No. 9,315,577,B2, titled "Anti-hepcidin antibodies and methods of use," issued on April 19, 2016. In some embodiments, the anti-hepcidin antibody is LY2787106.

[0142] In some embodiments, the hepcidin neutralizer is an inhibitory nucleic acid that targets hepcidin. As a non-limiting set of examples, the inhibitory nucleic acid may be, but is not limited to, small interfering RNA (siRNA), microRNA (miRNA), short hairpin RNA (shRNA), Dicer substrate interfering RNA (dsiRNA), short siRNA, or single-stranded siRNA. In some embodiments, a double-stranded antisense oligonucleotide is an RNAi oligonucleotide. In some embodiments, inhibitory nucleic acids targeting hepcidin include, but are not limited to, siHepcidin and XEN701 disclosed in U.S. Patent Application Publication No. 20160186172, published June 30, 2016, titled “Compositions and methods for inhibiting hepcidin antimicrobial peptide (HAMP) or HAMP-related gene expression,” and U.S. Patent Application Publication No. 20120115930, published May 10, 2012, titled “Compositions and their uses directed to hepcidin,” the respective contents of which are incorporated herein by reference.

[0143] In some embodiments, the hepcidin neutralizer is anticarin for hepcidin. Anticarin protein is an artificial protein capable of binding to an antigen. Anticarin protein is an engineered lipocalin, an endogenous low-molecular-weight human protein typically found in plasma and other bodily fluids that naturally binds, stores, and transports a wide range of molecules. In some embodiments, the lipocalin for hepcidin is the lipocalin disclosed in U.S. Patent No. 9051382B2, issued June 6, 2015, titled “Human neutrophil gelatinase-associated lipocalin (hNGAL) muteins that bind hepcidin and nucleic acid encoding such,” and U.S. Patent Application Publication No. 20150369821, published December 14, 2015, titled “Novel lipocalin-mutein assays for measuring hepcidin concentration,” the respective contents of which are incorporated herein by reference. In some embodiments, the antikarin for hepcidin is PRS-080.

[0144] In some embodiments, the hepcidin neutralizer is a PEGylated L-stereoisomerized RNA aptamer that binds to and neutralizes hepcidin. In some embodiments, the PEGylated L-stereoisomerized RNA aptamer for hepcidin is the PEGylated L-stereoisomerized RNA aptamer for hepcidin described in U.S. Patent No. 8,841431B2, issued September 23, 2014, titled “Hepcidin binding nucleic acids”; and International Publication No. 2012055573A1, published May 3, 2012, titled “Use of hepcidin binding nucleic acids for depletion of hepcidin from the body,” the respective contents of which are incorporated herein by reference. In some embodiments, the PEGylated L-stereoisomerized RNA aptamer for hepcidin is NOX-94. Other molecules that specifically bind to hepcidin include, but are not limited to, 12B9m, LS-B4534, lipocalin mutant proteins, and hNGAL mutant proteins. In some embodiments, other molecules that specifically bind to hepcidin are those described in U.S. Patent No. 8,629,250, issued January 14, 2014, titled “Hepcidin, hepcidin antagonists and methods of use”; U.S. Patent No. 9,315,577, issued April 19, 2016, titled “Anti-hepcidin antibodies and methods of use”; U.S. Patent No. 9,051,382, issued June 9, 2015, titled “Human neutrophil gelatinase-associated lipocalin (hNGAL) muteins that bind hepcidin and nucleic acid encoding such”; U.S. Patent No. 9,657,098, issued May 23, 2017, titled “Anti-hepcidin antibodies and uses thereof”; and “Methods for preventing or treating disorders by increasingU.S. Patent Publication No. 9,610356, “bioavailability of iron and related pharmaceutical formulation”; U.S. Patent Publication No. 8,530619, “Identification of the hepcidin binding site on ferroportin,” issued on September 10, 2013; U.S. Patent Publication No. 20150291675, “Human neutrophil gelatinase-associated lipocalin (hngal) muteins that bind hepcidin and nucleic acid encoding such,” published on October 15, 2015; and U.S. Patent Publication No. 20180057812, “Hepcidin antagonists for use in the treatment of inflammation,” published on March 1, 2018. In some embodiments, the hepcidin antagonists are molecules described in “Anti-hepcidin antibodies and uses,” issued on October 26, 2010, the contents of which are incorporated herein by reference. U.S. Patent No. 7820163B2 titled "thereof"; U.S. Patent No. 8328308B2, published December 11, 2012, titled "Fluid ejecting apparatus, fluid ejecting head control method in fluid ejecting apparatus, and driving waveform generating apparatus for fluid ejecting head"; Honduran Patent Application Publication No. 2010000752A, published August 7, 2012, titled "Anti-hepcidin antibodies and uses thereof"; U.S. Patent No. 8609817B2, published December 17, 2013, titled "Anti-hepcidin-25 selective antibodies and uses thereof"; and "Organic compositions to treat" published April 9, 2015.International Publication No. 2015 / 051135A2 for “hepcidin-related diseases”, U.S. Patent No. 8841431B2 for “Hepcidin Binding Nucleic Acids” published on September 23, 2014, U.S. Patent Application Publication No. 2014 / 057970 for “Use of Hepcidin Binding Nucleic Acids for Depletion of Hepcidin From the Body” published on February 27, 2014, U.S. Patent Application Publication No. 2015 / 0369821A1 for “Novel lipocalin-mutein assays for measuring hepcidin concentration” published on December 24, 2015, U.S. Patent No. 9051382B2 for “Binding proteins for hepcidin” published on June 9, 2015, and “Methods for preventing or treating disorders by” published on April 4, 2017 U.S. Patent No. 9610356B2 for “increasing bioavailability of iron and related pharmaceutical formulation”, U.S. Patent No. 9228188B2 for “Compositions and Method for Inhibiting Hepcidin Antimicrobial Peptide (HAMP) or HAMP-Related Gene Expression” issued on January 5, 2016, U.S. Patent No. 9315577B2 for “Anti-hepcidin antibodies and methods of use” issued on April 19, 2016, U.S. Patent No. 8629250B2 for “Hepcidin, hepcidin antagonists and methods of use” issued on January 14, 2014, International Publication No. 2018 / 128828A1 for “Novel hepcidin mimetics and uses thereof” published on July 12, 2018, and “Assessment of chronic” published on September 13, 2018.International Publication No. 2018 / 165186A1 for “iron deficiency”, U.S. Patent No. 7411048B2 issued on August 12, 2008, titled “Diagnostic method for diseases by screening for hepcidin in human or animal tissues, blood or body fluids and therapeutic uses therefor”, U.S. Patent No. 8915875B2 issued on December 23, 2014, titled “Adsorbents for the adsorption of hepcidin”, Chinese Patent Application Publication No. 101816674A issued on September 1, 2010, titled “Hepcidin inhibitor and application thereof”, U.S. Patent No. 9657098B2 issued on May 23, 2017, titled “Anti-hepcidin antibodies and uses thereof”, and “Humanized anti-hepcidin antibodies and uses U.S. Patent No. 10323088B2 titled "thereof its", U.S. Patent No. 4628027A published on December 9, 1986, titled "Vitro diagnostic methods using monoclonal antibodies against connective tissue proteins", Japanese Patent Application Publication No. 2019147772A published on September 5, 2019, titled "Hepcidin expression inhibitor, and food and drink for improvement and / or prevention of iron-deficiency anemia", and "Sulphated glycosaminoglycans, including heparin and derivatives thereof, for use in inhibiting the expression of hepcidin and for the therapeutic treatment of anemia" published on October 9, 2013.European Patent No. 2335708B1, titled "with high levels of hepcidin", US Patent Application Publication No. 2016 / 0122409A1, published on May 5, 2016, titled "Erythroferrone and erfe polypeptides and methods of regulating iron metabolism", US Patent Application Publication No. 20120214803A1, published on August 23, 2012, titled "Novel Sulfonaminoquinoline Hepcidin Antagonists", International Publication No. 2011 / 023722A1, published on March 3, 2011, titled "Novel quinoxalinone hepcidin antagonists", and "Novel thiazol and oxazol hepcidin", published on March 17, 2011. This is a hepcidine antidote described in International Publication No. 2011 / 029832A1 of “antagonists”, U.S. Patent Application Publication No. 2012 / 0196853A1, published on August 2, 2012, with the title “Novel Quinoline-Hepcidine Antagonists”, U.S. Patent Application Publication No. 2012 / 0214798A1, published on August 23, 2012, with the title “Novel Ethanediamone Hepcidine Antagonists”, U.S. Patent Application Publication No. 2012 / 0202806A1, published on August 9, 2012, with the title “Novel Pyrimidine- And Triazine-Hepcidine Antagonists”, and Chinese Patent Application Publication No. 103655542B, published on April 13, 2016, with the title “Ampelopsin suppresses the application in the preparation of ferrum tune element expression in preparation”.

[0145] In some embodiments, the hepcidin inhibitor is a molecule that specifically binds to ferroportin (e.g., an antibody, antikalin, or aptamer). In some embodiments, the molecule that specifically binds to ferroportin is LY2928057. Molecules that bind to ferroportin and inhibit hepcidin binding without affecting ferroportin activity are described (see also U.S. Patent No. 8,183346 and U.S. Patent No. 9,175078, and International Publication No. 2010065496A1, the full contents of each of these are incorporated herein by reference). In some embodiments, the hepcidin inhibitor is a chemical modifier compound that modifies hepcidin or ferroportin to inhibit the hepcidin-ferroportin binding interaction. For example, in some embodiments, the hepcidin inhibitor is fursultiamine (see, e.g., Fung and Nemeth. Haematologica. 2013 Nov; 98(11):1667-76).

[0146] iii. JAK / STAT signaling antagonists In some embodiments, hepcidin antagonists bind to and inhibit molecules involved in the JAK-STAT signaling pathway. Therefore, in some embodiments, hepcidin antagonists are JAK-STAT signaling pathway inhibitors. Examples of such antagonists include, but are not limited to, IL-6, the IL-6 receptor, JAK1 / 2, and STAT3. In some embodiments, the JAK-STAT signaling pathway inhibitor is either a JAK inhibitor or a STAT inhibitor. In some embodiments, the JAK inhibitor is selective for one or both of the JAK1 and JAK2 subtypes (e.g., a JAK1 / 2 inhibitor). In some embodiments, the STAT inhibitor is a STAT3 inhibitor.

[0147] The JAK-STAT3 signaling pathway is activated by the inflammatory cytokine IL-6. Binding of IL-6 to the IL-6 receptor (IL-6R) induces receptor dimerization in hepatocytes, leading to activation of the JAK-STAT3 signaling pathway (Figure 2). Therefore, in some embodiments, JAK-STAT signaling pathway inhibitors are IL-6 antagonists that bind to IL-6 and / or the IL-6 receptor, thereby antagonizing hepcidin function by inhibiting activation of the JAK-STAT3 signaling pathway. In some embodiments, the IL-6 antagonist is selected from the group consisting of infliximab, curcumin, 3,3'-diindolylmethane, tocilizumab, and siltuximab. In some embodiments, IL-6 and IL-6R inhibitors are, for example, U.S. Patent Application Publication No. 20170029499A1, published February 2, 2017, titled “Methods for treating hepcidin-mediated disorders,” International Publication No. 2008144757A1, published November 27, 2008, titled “Novel rabbit antibody humanization methods and humanized rabbit antibodies,” U.S. Patent Application Publication No. 20090104187A1, published April 23, 2009, titled “Novel Rabbit Antibody Humanization Methods and Humanized Rabbit Antibodies,” and “Antagonists of IL-6 to prevent or treat,” published June 10, 2010. International publication no. 2010065077A2 of “thrombosis”, international publication no. 2011066369A2 of “Antagonists of il-6 to raise albumin and / or lower crp” published on June 3, 2011, and international publication no. 2011066369A2 of “Antagonists of il-6 to raise albumin and / or lowerU.S. Patent No. 9701747B2 for "crp", U.S. Patent No. 8420089B2 issued on April 16, 2013, for "Antagonists of IL-6 to raise albumin and / or lower crp", U.S. Patent No. 9265825B2 issued on February 23, 2016, for "Antagonists of IL-6 to raise albumin and / or lower crp", U.S. Patent No. 8277804B2 issued on October 2, 2012, for "Antagonists of IL-6 to prevent or treat thrombosis", U.S. Patent No. 9085615B2 issued on July 21, 2015, for "Antibodies to IL-6 and use thereof", and published on June 3, 2011, for "Antibodies to IL-6 and use International Publication No. 2011066371A2 for “thereof”; U.S. Patent No. 8323649B2 issued on December 4, 2012, for “Antibodies to IL-6 and use thereof”; U.S. Patent No. 9452227B2 issued on September 27, 2016, for “Antibodies to IL-6 and use thereof”; International Publication No. 2010065079A2 issued on June 10, 2010, for “Antibodies to IL-6 and use thereof”; U.S. Patent No. 9724410B2 issued on August 8, 2017, for “Antagonists of IL-6 to prevent or treat cachexia, weakness, fatigue and / or fever”; and “Novel rabbit antibody humanization methods and humanized rabbit” published on September 24, 2009. U.S. Patent Application Publication No. 20090238825A1 for "antibodies", U.S. Patent No. 9993480B2 issued on June 12, 2018, with the title "mTOR / JAK INHIBITOR COMBINATION THERAPY", and "Methods for treating" published on February 2, 2017.These are anti-IL-6 or IL-6R inhibitors described in U.S. Patent Application Publication No. 20170029499A1 for "hepcidin-mediated disorders" and U.S. Patent Application Publication No. 20190241650A1, published on August 8, 2019, with the title "Methods for treating IL-6 mediated inflammation without immunosuppression".

[0148] In some embodiments, the JAK-STAT antagonist is a selective JAK1 inhibitor (determined, for example, by a kinase efficacy assay described herein). In some embodiments, the JAK-STAT antagonist is a JAK2 inhibitor (determined, for example, by a kinase efficacy assay described herein). In some embodiments, the JAK-STAT antagonist is inactive against ACVR1 / ALK2. In some embodiments, the JAK-STAT antagonist is ruxolitinib, fedratinib, pacritinib, baricitinib, tofacitinib, oclacitinib, or NSC13626. In some embodiments, the JAK / STAT antagonist is GS-0387 or CYT-387.

[0149] In some embodiments, the JAK / STAT antagonist is a selective JAK1 / JAK2 inhibitor. In some embodiments, the selective JAK1 / JAK2 inhibitor is ruxolitinib. Suitable JAK1 / JAK2 inhibitors for use in the treatment of myelofibrosis include, for example, U.S. Patent No. 7,598,257, issued October 6, 2009, titled “Heteroaryl substituted pyrrolo[2,3-b]pyridines and pyrrolo[2,3-b]pyrimidines as Janus kinase inhibitors,” issued April 9, 2013, titled “Pyrazolyl substituted pyrrolo[2,3-b]pyrimidines as Janus kinase inhibitors,” and “Salts of the Janus kinase inhibitor,” issued May 13, 2014. U.S. Patent No. 8722693 for “(R)-3-(4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-cyclopentylpropanenitrile”; U.S. Patent No. 8822481 for “Salts of the janus kinase inhibitor (R)-3-(4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-cyclopentylpropanenitrile” issued on September 2, 2014;U.S. Patent No. 8829013, issued on September 9, 2014, with the title "Salts of the Janus kinase inhibitor (R)-3-(4-(7H-pyrrolo[2,3-D]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-cyclopentylpropanenitrile"; U.S. Patent No. 9079912, issued on September 2, 2014, with the title "Salts of the janus kinase inhibitor (R)-3-(4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-cyclopentylpropanenitrile"; and U.S. Patent No. 9079912, issued on November 14, 2017, with the title "Heteroaryl substituted pyrrolo[2,3-B] pyridines and pyrrolo[2,3-B] pyrimidines as janus kinase This is described in U.S. Patent No. 9,814,722, “inhibitors” and U.S. Patent No. 1,001,6429,B2, issued on July 10, 2018, with the title “Salts of the janus kinase inhibitor (R)-3-(4-(7H-pyrrolo[2,3-D]pyrimidin-4-yl)-1H-pyrazol-1-yl)-3-cyclopentylpropanenitrile”. In some embodiments, the JAK1 / JAK2 inhibitor is ruxolitinib (RUX).

[0150] In some embodiments, the JAK-STAT inhibitor is a selective JAK2 inhibitor. Suitable JAK2 inhibitors for use in the treatment of myelofibrosis are described, for example, in U.S. Patent No. 7,528,143, titled “Bi-aryl meta-pyrimidine inhibitors of kinases,” issued on 5 May 2009; U.S. Patent No. 7,825,246, titled “Bi-aryl meta-pyrimidine inhibitors of kinases,” issued on 2 November 2010; U.S. Patent No. 8,138,199, titled “Use of bi-aryl meta-pyrimidine inhibitors of kinases,” issued on 20 March 2012; and U.S. Patent No. 1,039,1094, titled “Compositions and methods for treating myelofibrosis,” issued on 27 August 2019, whose entire contents are incorporated herein by reference. In some embodiments, the selective JAK2 inhibitor is fedratinib.

[0151] In some embodiments, the JAK-STAT inhibitor is not a selective JAK inhibitor. In some embodiments, the JAK-STAT inhibitor is an inhibitor of JAK1 / JAK2 and ACVRI (also known as ALK2) (e.g., momerotinib). Suitable JAK1 / JAK2 and ACVRI (ALK2) inhibitors for use in the methods provided herein are described, for example, in U.S. Patent No. 8,486,941B2, issued July 16, 2013, titled “Phenyl amino pyrimidine compounds and uses thereof” and in U.S. Patent No. 1,024,5268B2, issued April 2, 2019, titled “Momelotinib for treating of acvr1-mediated diseases,” the entire contents of which are incorporated herein by reference. In some embodiments, the non-selective JAK-STAT inhibitor is momelotinib (see, for example, ASSHOFF MALTE ET AL: "The Jak1 / Jak2 Inhibitor Momelotinib Inhibits Alk2, Decreases Hepcidin Production and Ameliorates Anemia of Chronic Disease (ACD) in Rodents", BLOOD, vol. 126, no. 23, December 2015 (2015-12-01)).

[0152] In some embodiments, the JAK inhibitors are those described in U.S. Patent No. 8,202881B2, published June 19, 2020, titled “JAK2 inhibitors and their use for the treatment of myeloproliferative diseases and cancer,” U.S. Patent No. 8,193189B2, published June 5, 2012, titled “Quinoxaline derivatives as tyrosine kinase activity inhibitors,” each of which is incorporated herein by reference in its entirety; U.S. Patent No. 8,629168B2, published January 14, 2014, titled “Benzoxazoles and oxazolopyridines being useful as janus kinases inhibitors,” and “Treatment of jak2-mediated,” published March 13, 2014. U.S. Patent Application Publication No. 2014 / 073643 for “conditions”, U.S. Patent No. 9469613 B2 for “(N-(cyanomethyl)-4-(2-(4-morpholinophenylamino)pyrimidin-4-yl)benzamide” issued on October 18, 2016, International Publication No. 2020 / 041466 A1 for “Platelet count-agnostic methods of treating myelofibrosis” issued on February 27, 2020, International Publication No. 2018 / 096525 A2 for “Heteroaryl compounds and uses thereof” issued on May 31, 2018, and “N-(hetero)aryl-pyrrolidine derivatives of pyrazol-4-yl-pyrrolo[2,3-d]pyrimidines and” issued on May 6, 2014 U.S. Patent No. 8716303B2, issued November 16, 2010, for "pyrrol-3-yl-pyrrolo[2,3-d]pyrimidines as Janus kinase inhibitors," titled "METABOLITES OF THE JANUS KINASE INHIBITORS."U.S. Patent No. 7,834,022B2, titled "(R)-3-(4-(7H-PYRROLO[2,3-d]PYRIMIDIN-4-YL)-1H-PYRAZOL-1-YL)-3-CYCLOPENTYLPROPANENITRILE", U.S. Patent Publication No. 20070149506A1, published June 28, 2007, titled "Azepine inhibitors of Janus kinases", U.S. Patent No. 8,513,270B2, published August 20, 2013, titled "Substituted heterocycles as janus kinase inhibitors", U.S. Patent No. 9,359,358B2, published June 7, 2016, titled "Cyclohexyl Azetidine Derivatives as JAK Inhibitors", and "HYDROXYL, KETO, AND U.S. Patent No. 8486902B2 for “GLUCURONIDE DERIVATIVES OF 3-(4-(7H-PYRROLO[2,3-d] PYRIMIDIN-4-YL)-1H-PYRAZOL-1-YL)-3-CYCLOPENTYLPROPANENITRILE”, U.S. Patent No. 9358229B2 for “JAK PI3K / mTOR COMBINATION THERAPY” issued on June 7, 2016, U.S. Patent No. 8933085B2 for “Cyclobutyl substituted pyrrolopyridine and pyrrolopyrimidine derivatives as jak inhibitors” issued on January 13, 2015, and “Macrocyclic compounds and their use as kinase” issued on July 1, 2014 U.S. Patent No. 8,765,727B2 for “inhibitors”, U.S. Patent No. 9,034,884B2 for “Heterocyclic-substituted pyrrolopyridines and pyrrolopyrimidines as jak inhibitors” issued on May 19, 2015, and “SALTS OF THE JANUS KINASE INHIBITOR” published on December 18, 2008.US20080312259A1 for “(R)-3-(4-(7H-PYRROLO[2,3-d]PYRIMIDIN-4-YL)-1H-PYRAZOL-1-YL)-3-CYCLOPENTYLPROPANENITRILE”, US Patent Application Publication No. 20150246046A1, published on September 3, 2015, with the title “Jak1 inhibitors for the treatment of myelodysplastic syndromes”, US Patent No. 8158616B2, issued on April 17, 2012, with the title “Azetidine and cyclobutane derivatives as jak inhibitors”, and US Patent Publication No. 8158616B2, issued on February 26, 2008, with the title “Pyrrolo[2,3-b]pyridin-4-yl-amines and pyrrolo[2,3-b]pyrimidin-4-yl-amines as janus kinase U.S. Patent No. 7,335667B2, titled "inhibitors", U.S. Patent No. 1,0064866B2, published on September 4, 2018, titled "Treatment of b-cell malignancies by a combination jak and pi3k inhibitors", U.S. Patent Publication No. 20110207754A1, published on August 25, 2011, titled "Cyclobutane and methylcyclobutane derivatives as janus kinase inhibitors", U.S. Patent No. 9,193733B2, published on November 24, 2015, titled "Piperidinylcyclobutyl substituted pyrrolopyridine and pyrrolopyrimidine derivatives as jak inhibitors", U.S. Patent No. 9,382231B2, published on July 5, 2016, titled "Bipyrazole derivatives as jak inhibitors", and "Azetidinyl phenyl, pyridyl or pyrazinyl carboxamide derivatives as jakU.S. Patent No. 8691807B2 for “inhibitors”, U.S. Patent No. 9181271B2 issued on November 10, 2015, with the title “Tricyclic fused thiophene derivatives as jak inhibitors”, U.S. Patent No. 8604043B2 issued on December 10, 2013, with the title “3-[4-(7h-pyrrolo[2,3-d]pyrimidin-4-yl)-1h-pyrazol-1-yl]octane- or heptane-nitrile as jak inhibitors”, and U.S. Patent No. 8604043B2 issued on February 2, 2016, with the title “HETEROCYCLIC DERIVATIVES OF PYRAZOL-4-YL-PYRROLO[2,3-d]PYRIMIDINES AS JANUS KINASE”. U.S. Patent No. 9,249145B2, titled "INHIBITORS", U.S. Patent No. 8,765734B2, published July 1, 2014, titled "Piperidin-4-yl azetidine derivatives as jak1 inhibitors", U.S. Patent Application Publication No. 20060106020A1, published May 18, 2006, titled "Tetracyclic inhibitors of Janus kinases", U.S. Patent No. 9,993480B2, published June 12, 2018, titled "mTOR / JAK INHIBITOR COMBINATION THERAPY", and "4-pyrazolyl-n-arylpyrimidin-2-amines and 4-pyrazolyl-n-heteroarylpyrimidin-2-amines as janus kinase" published November 13, 2012. U.S. Patent No. 8309718B2 titled "inhibitors", U.S. Patent Publication No. 20060153852A1 titled "Novel human Jak2 kinase" published on July 13, 2006, U.S. Patent No. 8871753B2 titled "Macrocyclic compounds and their use as kinase inhibitors" issued on October 28, 2014, and "4-(3-Aminopyrazole) Pyrimidine" published on November 20, 2008.U.S. Patent Publication No. 20080287475A1, titled “Derivatives for Use as Tyrosine Kinase Inhibitors in the Treatment of Cancer”, U.S. Patent Publication No. 20090062302A1, published March 5, 2009, titled “Jak2 Tyrosine Kinase Inhibition”, U.S. Patent No. 8648069B2, published February 11, 2014, titled “5-substituted indazoles as kinase inhibitors”, International Publication No. 2017196261A1, published November 16, 2017, titled “Jak and hdac dual-inhibitor compounds”, and “Therapeutic Combinations of a BTK Inhibitor, a PI3K Inhibitor and / or a JAK-2 U.S. Patent Application Publication No. 9949971B2 for “Inhibitor”, published August 10, 2017; U.S. Patent Application Publication No. 20170224819A1 for “Therapeutic Combinations of a BTK Inhibitor, a PI3K Inhibitor, a JAK-2 Inhibitor, and / or a CDK 4 / 6 Inhibitor”, published August 24, 2017; U.S. Patent Application Publication No. 9949971B2 for “Inhibitor”, published August 10, 2017; U.S. Patent Application Publication No. 20170224819A1 for “Therapeutic Combinations of a BTK Inhibitor, a PI3K Inhibitor, a JAK-2 Inhibitor, a PD-1 Inhibitor, and / or a PD-L1 U.S. Patent Application Publication No. 20170239351A1 titled “Inhibitor”, U.S. Patent Application Publication No. 20180317602A1 titled “Combinations”, published on November 8, 2018, and “Pyrazolyl pyrrolo[2,3-b]pyrmidine-5-carboxylate analogs and methods of making theU.S. Patent Application Publication No. 20190135807A1 for “same”, U.S. Patent Application Publication No. 20130089512A1 for “Heteroaryl imidazolone derivatives as jak inhibitors” published on April 11, 2013, U.S. Patent No. 9206183B2 for “Pyrazole derivatives as jak inhibitors” issued on December 8, 2015, U.S. Patent No. 9034311B2 for “Pyridin-2(1h)-one derivatives as jak inhibitors” issued on May 19, 2015, and “Pyridin-2(1h)-one derivatives useful as medicaments for the treatment of myeloproliferative disorders, transplant rejection, immune-mediated and inflammatory” published on June 19, 2014. U.S. Patent Publication No. 20140170110A1 for “diseases”, International Publication No. 2015091531A1 for “Imidazolopyrimidin-2-yl derivatives as jak inhibitors” published on June 25, 2015, U.S. Patent Publication No. 20130216498A1 for “Imidazopyridine derivatives as jak inhibitors” published on August 22, 2013, U.S. Patent No. 9133200B2 for “Imidazo [1,2-b] pyridazine and imidazo [4,5-b] pyridine derivatives as jak inhibitors” issued on September 15, 2015, U.S. Patent No. 8349851B2 for “JAK kinase modulating compounds and methods of use thereof” issued on January 8, 2013, and “Substituted pyrrolo[2,3-b]pyridines as ITK and JAKU.S. Patent No. 9206188B2 for “inhibitors”, International Publication No. 2010 / 020810 A1 for “2-(imidaz0lylamin0)-pyridine derivatives and their use as jak kinase inhibitors” published on February 25, 2010, U.S. Patent Application Publication No. 2010 / 324040 A1 for “9-(pyrazol-3-yl)-9h-purine-2-amine and 3-(pyrazol-3-yl)-3h-imidazo[4,5-b]pyridin-5-amine derivatives and their use for the treatment of cancer” published on July 16, 2013, and International Publication No. 2009 / 027736 for “2,4 diaminopyrimides for the treatment of myeloproliferative disorders and cancer” published on March 5, 2009. U.S. Patent Application Publication No. 2011 / 201628 A1, titled "Heterocyclic jak kinase inhibitors," published on August 18, 2011; International Publication No. 2008 / 117050 A1, titled "Pyrazolyl-amino-substituted pyrazines and their use for the treatment of cancer," published on October 2, 2008; U.S. Patent Application Publication No. 2010 / 204246 A1, titled "5-aminopyrazol-3-yl-3h-imidazo (4,5-b) pyridine derivatives and their use for the treatment of cancer," published on August 12, 2010. International Publication No. 2008 / 132502 A1, published November 6, 2008, with the title "Pyrazolyl-amino-substituted pyrimidines and their use for the treatment of cancer"; International Publication No. 2009 / 016410 A3, published February 5, 2009, with the title "Chemical compounds 831"; International Publication No. 2009 / 095712 A2, published August 6, 2009, with the title "4-(3-amin0pyraz0le) pyrimidine derivatives for use as jak kinase inhibitors in the treatment of cancer"; U.S. Patent No. 8440679B2, issued May 14, 2013, with the title "Bicyclic compounds and their uses as dual c-SRC / JAK inhibitors"; and "Pyrrolo[2,3-D]pyrimidine" issued May 16, 2015. U.S. Patent No. 9035074B2 for “derivatives”, International Publication No. 2019057112 A1 for “2-substituted pyrazole amino-4-substituted amino-5-pyrimidine formamide compound, composition, and application thereof” published on March 28, 2019, U.S. Patent No. 8815840B2 for “Carbazole and carboline kinase inhibitors” published on August 26, 2014, U.S. Patent No. 8957065B2 for “Fused pyrimidine derivatives for inhibition of tyrosine kinase activity” published on February 17, 2015, and International Publication No. 2014020531 for “Imidazo[1,2-b]pyridazin-6-amine derivatives as kinase jak-2 inhibitors” published on February 6, 2014International publication No. 2015118434 A1, published on August 13, 2015, with the title "PYRAZOLO[1,5-a]PYRIMIDINE DERIVATIVES AS KINASE JAK-2 INHIBITORS"; U.S. Patent Application Publication No. 2008021013 A1, published on January 24, 2008, with the title "JAK inhibitors for treatment of myeloproliferative disorders"; U.S. Patent No. 8937065B2, issued on January 20, 2015, with the title "Compositions and methods for modulating a kinase"; U.S. Patent Application Publication No. 2015 / 197525 A1, published on July 16, 2015, with the title "Deuterated derivatives of ruxolitinib"; issued on January 10, 2017, with the title "Deuterated U.S. Patent No. 9540367B2 for “baricitinib”, U.S. Patent No. 9518027B2 for “Deuterated momelotinib” issued on December 13, 2016, U.S. Patent No. 8354408B2 for “N-containing heterocyclic compound” issued on January 15, 2013, International Publication No. 2020097396 A1 for “Benzimidazole derivatives and aza-benzimidazole derivatives as janus kinase 2 inhibitors and uses thereof” published on May 14, 2020, International Publication No. 2020097398 A1 for “Benzothiazole derivatives and 7-aza-benzothiazole derivatives as janus kinase 2 inhibitors and uses thereof” published on May 14, 2020, and “Compositions and methods of use thereof for treatment of metabolic diseases” published on January 9, 2020. International publication No. 202009740 of “and related disorders”U.S. Patent No. 10111897B2, published on October 30, 2018, with the title "Compositions and methods for treating cancer with JAK2 activity"; U.S. Patent No. 8440663B2, published on May 14, 2013, with the title "4-aryl-2-amino-pyrimidines or 4-aryl-2-aminoalkyl-pyrimidines as JAK-2 modulators and methods of use"; U.S. Patent No. 8088767B2, published on January 3, 2012, with the title "JAK-2 modulators and methods of use"; International Publication No. 2009017838A2, published on February 5, 2009, with the title "Combinations of jak-2 inhibitors and other agents"; and U.S. Patent Application Publication No. 2010 / 035875, published on February 11, 2010, with the title "Triazolopyridine jak inhibitor compound and methods". U.S. Patent Application Publication No. 2010 / 048557, A1, published on February 25, 2010, with the title "Triazolopyridine JAK Inhibitor Compounds and Methods"; U.S. Patent No. 10307426B2, issued on June 4, 2019, with the title "Therapeutic compounds and compositions, and methods of use thereof"; U.S. Patent No. 8637526B2, issued on January 28, 2014, with the title "Pyrazolopyrimidine JAK inhibitor compound and methods"; U.S. Patent No. 8999998B2, issued on April 7, 2015, with the title "Pyrazolopyrimidine JAK inhibitor compound and methods"; U.S. Patent Application Publication No. 2019 / 040068, published on February 7, 2019, with the title "Pyrrolopyrimidine five-membered azacyclic derivative and application thereof";U.S. Patent Publication No. 2019 / 328857 A1, published on October 31, 2019, with the title "Calr and jak2 vaccine compositions"; International Publication No. 11153586 A1, published on December 15, 2011, with the title "Kinase inhibitors"; U.S. Patent No. 10294226B2, issued on May 21, 2019, with the title "Small molecule inhibitors of the JAK family of kinases"; U.S. Patent Publication No. 2019 / 322665 A1, published on October 24, 2019, with the title "Imidazopyrrolopyridine as inhibitors of the jak family of kinases"; and "Aminopyrimidine kinase" issued on December 2, 2014. U.S. Patent No. 8901145B2 for “inhibitors”, U.S. Patent No. 8563539B2 for “Aminopyrimidine kinase inhibitors” published on October 22, 2013, U.S. Patent Publication No. 2018 / 002328 A1 for “Substituted imidazo[1,2-a]pyridin-2-ylamine compounds, and pharmaceutical compositions and methods of use thereof” published on January 4, 2018, International Publication No. 2017 / 143014 A1 for “Jak inhibitors and uses thereof” published on August 24, 2017, International Publication No. 2019 / 107943 A1 for “Jak inhibitor compound and preparation method therefor” published on June 6, 2019, International Publication No. 2008 / 047831 for “JAK inhibitor” published on February 25, 2010 International publication number 2010 / 141062 for "Inhibitors of constitutively active janus kinases and uses thereof," published on December 9, 2010, in publication A1.U.S. patent no. A1, U.S. patent no. 8278335B2 issued on October 2, 2012, with the title "Inhibitors of Janus kinases", U.S. patent no. 8349865B2 issued on January 8, 2013, U.S. patent no. 8344144B2 issued on January 1, 2013, with the title "Inhibitors of Janus kinases", U.S. patent no. US8420695B2 issued on April 16, 2013, with the title "Inhibitors of Janus kinases", U.S. patent no. 8367706B2 issued on February 5, 2013, with the title "Inhibitors of Janus kinases", and U.S. patent no. 8367706B2 issued on April 30, 2014 U.S. Patent No. 8431569B2 for "kinases", U.S. Patent No. 8415346B2 for "Inhibitors of Janus kinases" issued on April 9, 2013, U.S. Patent No. 8183245B2 for "Pyrazine substituted pyrrolopyridines as inhibitors of JAK and PDK1" issued on May 22, 2012, U.S. Patent No. 7763634B2 for "Inhibitors of janus kinases" issued on July 27, 2010, U.S. Patent No. 9283224B2 for "Substituted pyrimidinyl-pyrroles active as kinase inhibitors" issued on March 15, 2016, and "Alkynyl substituted pyrimidinyl-pyrroles active as kinases" issued on December 16, 2014 U.S. Patent No. 8912200B2 for “inhibitors”, U.S. Patent No. 9688661B2 issued on June 27, 2017, with the title “Substituted pyrroles active as kinases inhibitors”, and U.S. Patent No. 9688661B2 issued on March 18, 2014, with the title “Aminopyrazine derivative andU.S. Patent No. 8673891B2 for “medicine”, U.S. Patent No. 8629168B2 for “Benzoxazoles and oxazolopyridines being useful as janus kinases inhibitors” issued on January 14, 2014, U.S. Patent No. 8501735B2 for “N-containing heteroaryl derivatives as JAK3 kinase inhibitors” issued on August 6, 2013, International Publication No. 2010 / 072823A1 for “PYRAZOLE[1,5a]PYRIDINE DERIVATIVES” published on July 1, 2010, U.S. Patent No. 8633206B2 for “Pyrrolo[2,3-D]pyrimidine compounds” issued on January 21, 2014, and “Pyrrolo[2,3-d]pyrimidinyl,” issued on April 11, 2017. U.S. Patent No. 9,617258B2 for "pyrrolo[2,3-b]pyrazinyl and pyrrolo[2,3-d]pyridinyl acrylamides", International Publication No. 2009 / 017954A1 for "Inhibitors of jak2 kinase" published on February 5, 2009, U.S. Patent No. 8,258144B2 for "Inhibitors of protein kinases" published on September 4, 2012, U.S. Patent No. 9,676756B2 for "Substituted pyrimidinyl kinase inhibitors" published on June 13, 2017, U.S. Patent No. 9,533986B2 for "Bicyclic dihydropyridone kinase inhibitors" published on January 3, 2017, and "Bicyclic oxa-lactam kinase" published on October 18, 2016. U.S. Patent No. 9,469,654B2, titled "Inhibitors of protein kinases," issued on March 20, 2012; U.S. Patent No. 8,138,339B2, titled "Inhibitors of protein kinases," issued on November 13, 2012; and "Pyrimidine-2-amine compounds and their use asU.S. Patent No. 8309566B2, titled "inhibitors of JAK kinases", U.S. Patent No. 8846908B2, published September 30, 2014, titled "Tricyclic carbamate JAK inhibitors", U.S. Patent No. 10011571B2, published July 3, 2018, titled "Preparation method for aromatic heterocyclic compound used as selective JAK3 and / or JAK1 kinase inhibitor and application of aromatic heterocyclic compound", U.S. Patent Application Publication No. 2020 / 071303 A1, published March 5, 2020, titled "Diphenylaminopyrimidine compound for inhibiting kinase activity", U.S. Patent No. 9371320B2, published June 21, 2016, titled "Heterocyclic compound", and U.S. Patent Publication No. 9371320B2, published May 2, 2017, titled "Heterocyclic U.S. Patent No. 9637483B2 for “compound”, International Publication No. 2019 / 069844 for “Heterocyclic compound” published on April 11, 2019, International Publication No. 2009 / 046416 A1 for “Anilinopyrimidines as jak kinase inhibitors” published on May 9, 2009, International Publication No. 2009 / 049028 for “Pyrrolopyrimidine compounds and their use as janus kinase modulators” published on April 16, 2009, International Publication No. 2009 / 055674 A1 for “Pyrrolopyrimidine alkynyl compounds and methods of making and using same” published on April 30, 2009, U.S. Patent No. 10202356B2, published on February 12, 2019, with the title "JAK2 and ALK2 inhibitors and methods for their use"; U.S. Patent Publication No. 2019 / 169208 A1, published on June 6, 2019, with the title "Macrocycle kinase inhibitors"; U.S. Patent Publication No. 2011 / 243853 A1, published on October 6, 2011, with the title "Models of erythropoiesis"; U.S. Patent No. 8367078B2, published on February 5, 2013, with the title "Kinase inhibitor compounds"; and U.S. Patent Publication No. 2015 / 306112, published on October 29, 2015, with the title "Zhankuic acid A, a JAK2 / 3 tyrosine kinase inhibitor, and a potential therapeutic agent for hepatitis". U.S. Patent Publication No. 2014 / 286964 A1, published on September 25, 2014, with the title "Methods for Identifying Janus Kinase (JAK) Modulators for Therapeutics", U.S. Patent No. 8937064B2, published on January 20, 2015, with the title "Pyrazolo[1,5-a]pyrimidines useful as JAK2 inhibitors", U.S. Patent No. 7767816B2, published on August 3, 2010, with the title "Azaindoles useful as inhibitors of janus kinases", U.S. Patent No. 8580802B2, published on November 12, 2013, with the title "Pyrrolo[2,3-D]pyrimidines as inhibitors of Janus kinases", published on January 21, 2014, with the title "Substituted pyrrolo[2,U.S. Patent No. 8,633205B2, titled "3-d]pyrimidines as inhibitors of protein kinases", U.S. Patent No. 8,741912B2, published June 3, 2014, titled "Deazapurines useful as inhibitors of Janus kinases", U.S. Patent Publication No. 2010 / 160287 A1, published June 24, 2010, titled "Compounds useful as inhibitors of janus kinases", U.S. Patent No. 8,921376B2, published December 30, 2014, titled "Pyrrolopyridines useful as inhibitors of protein kinase", U.S. Patent Publication No. 2014 / 073643 A1, published March 13, 2014, titled "Treatment of jak2-mediated conditions", and "Phenyl amino pyrimidine bicyclic compounds and uses" published August 19, 2014. It is a JAK inhibitor as described in U.S. Patent No. 8809359B2 of “thereof its”

[0153] In some embodiments, the JAK / STAT inhibitor is a STAT inhibitor. STAT inhibitors are previously described, for example, in U.S. Patent No. 8,779001B2, published July 15, 2014, titled “Stat3 inhibitors,” International Publication No. 2019 / 204427A1, published October 24, 2019, titled “Methods for measuring and stabilizing stat3 inhibitors,” each of which is incorporated herein by reference in its entirety. U.S. Patent No. 1,0112933B2, published October 30, 2018, titled “Methods and compositions for treatment of fibrosis,” and U.S. Patent No. 9,650399B2, published May 16, 2017, titled “Salicylic acid derivatives, pharmaceutically acceptable salt thereof, composition thereof and method of use thereof.”

[0154] iv. Chromatin modulator In some embodiments, the chromatin remodeling bromodomain and excess terminus (BET) protein regulate inflammation-involved genes such as MYC, BCL-2, and NF-κB. In some embodiments, the NF-κB pathway downstream of BET is activated in myelofibrosis, for example, via JAK-STAT signaling. The BET protein acts as an epigenetic reader, transmitting signals carried by acetylated lysine residues on histones and transcribing them to various phenotypes. Unregulated BET signaling is involved in many diseases, including myelofibrosis (MF). BET inhibitors are described, for example, in U.S. Patent Application Publication 20170333406A1, which is incorporated herein by reference. In some embodiments, the BET inhibitor for use in this disclosure is CPI-0610. CPI-0610 is a potent and selective small molecule designed to promote antitumor activity by selectively inhibiting the function of the BET protein, which reduces the expression of genes abnormally expressed in cancer. The BET protein binds to acetylated histone lysine residues and acts as a co-activator of gene expression. These compounds work in conjunction with the transcription factor NFκB to activate pro-inflammatory cytokine gene expression. CP-0610 downregulates pro-inflammatory cytokines in a mouse model, and the combination of a BET inhibitor and ruxolitinib synergistically reduces splenomegaly, cytokine expression, myelofibrosis, and mutant gene burden.In some embodiments, BET inhibitors suitable for use in the manner described herein include, each of which is incorporated herein by reference in its entirety: U.S. Patent Application Publication No. 2019152949, titled “Therapeutic compounds and uses thereof,” published on 23 May 2019; U.S. Patent No. 10206931B2, titled “Therapeutic compounds and uses thereof,” issued on 19 February 2019; U.S. Patent Application Publication No. 2016317632, titled “Use of cbp / ep300 bromodomain inhibitors for cancer immunotherapy,” published on 3 November 2016; U.S. Patent Application Publication No. 2017196878, titled “Use of cbp / ep300 and bet inhibitors for treatment of cancer,” published on 13 July 2017; and U.S. Patent Application Publication No. 2017196878, titled “Use of cbp / ep300 and bet inhibitors for treatment of cancer,” published on 22 August 2019. This is a BET inhibitor described in International Publication No. 2019161162 of “inhibitors”, International Publication No. 2020112086 of “Methods of treating myeloproliferative disorders” published on June 4, 2020, and International Publication No. 2019161157 of “P300 / cbp hat inhibitors” published on June 11, 2020.

[0155] v. Immunomodulators / Erythropoietin Stimulants In some embodiments, immunomodulatory agents are provided herein, for example, for the treatment of anemia associated with myelofibrosis. Such immunomodulatory agents include, for example, corticosteroids, androgenic steroids, thalidomide, pomalidomide, lenalidomide, and others. In some embodiments, immunomodulatory agents are beneficial because they have a favorable effect on reducing inflammation and promoting erythropoiesis. For example, danazol is a steroid compound that has hematopoietic stimulating and immunomodulatory effects. For example, in some embodiments, danazol has antagonist activity at glucocorticoid receptors, resulting in an upregulation of erythropoiesis (see, for example, Chai KY, et al., Danazol: An Effective and Underutilised Treatment Option in Diamond-Blackfan Anemia. Case Reports in Hematology. Volume 2019, Article ID 4684156). Similarly, in some embodiments, glucocorticoids such as prednisone, which promote erythropoiesis, may be useful in reducing inflammation, for example, in the fibrous bone marrow of MF patients (see, e.g., Amylon MD et al., Prednisone stimulation of erythropoiesis in leukemic children during remission. American Journal of Hematology. Volume 23, Issue 2, October 1986). Other immunomodulatory / erythropoietin stimulants that affect erythropoiesis include thalidomide and its derivatives or analogs, such as danazol, prednisone, thalidomide, lenalidomide, and pomalidomide. In some embodiments, erythropoietin (EPO) may be used in the manner described herein.

[0156] III. How to Use Aspects of this disclosure relate to compositions and methods for treating myelofibrosis and / or one or more conditions resulting from myelofibrosis in a subject.

[0157] Myelofibrosis (MF) is a myeloproliferative disorder characterized by the abnormal proliferation of hematopoietic stem cells that result in fibrosis of the bone marrow. The production of healthy blood cells (megakaryocytes and erythrocytes, which are responsible for platelet production) is impaired. MF can be classified into primary MF (PMF) and secondary MF (SMF). PMF and SMF have similar clinical profiles, including common main symptoms such as anemia, fatigue, and splenomegaly. Primary myelofibrosis (PMF) is characterized as MF that occurs on its own. Secondary myelofibrosis (SMF) occurs as a result of scar tissue in the bone marrow as a complication of another disease, such as an autoimmune disease. In some embodiments, the subjects described herein have or are suspected of having PMF. In some embodiments, the subjects described herein have or are suspected of having SMF.

[0158] In some embodiments, subjects having or suspected of having myelofibrosis (e.g., PMF and / or SMF) have one or more mutations in one or more genes. In some embodiments, subjects have one or more mutations in the JAK2 gene. JAK2 plays a crucial role in signaling from receptors involved in the proliferation of myeloid cell lines mediated by EPO, TPO, and / or G-CSF (see, e.g., Alshemmari et al., Molecular Pathogenesis and Clinical Significance of Driver Mutations in Primary Myelofibrosis: A Review, Med Princ Pract, 2016;25(6):501-509). In some embodiments, subjects contain a human JAK2 gene with a triggering mutation in exon 12 or exon 14. In some embodiments, the triggering mutation in the JAK2 gene is in exon 14, resulting in a V617F substitution. In some embodiments, the V617F mutation results in hyperactivation of JAK2 and its associated signaling pathways. In some embodiments, JAK2 hyperactivation leads to myelofibrosis (e.g., PMF and / or SMF).

[0159] In some embodiments, the subjects have one or more mutations in the thrombopoietin receptor (MPL) gene. MPL is a cognate receptor of thrombopoietin (TPO), and mutations that result in increased function of the MPL gene lead to impaired megakaryocyte production. In some embodiments, the subjects include the W515L / K mutation in MPL. In some embodiments, subjects with one or more mutations in the MPL gene are more likely to develop anemia compared to subjects with MF as a whole (e.g., more than 10%, more than 20%, more than 30%, more than 40%, more than 50%, more than 60%, more than 70%, more than 80%, more than 90%, more than twice, more than three times, more than four times, more than five times, more than six times, more than seven times, more than eight times, more than nine times, or more than ten times) (Guglielmelli P et al., Anemia characterises patients with myelofibrosis harbouring Mpl mutation. Br J Haematol 2007; 137: 244-247).

[0160] In some embodiments, subjects have one or more mutations in the calreticulin (CALR) gene. The CALR gene encodes calreticulin protein, a multifactorial protein that regulates calcium homeostasis, cell signaling, gene expression, cell adhesion, autoimmunity, and apoptosis. Approximately 140 CALR mutations have been identified in 19 variants and are associated with MF. In some embodiments, subjects have or are suspected of having MF with an exon 9 mutation in the CALR gene.

[0161] Further mutations in other genes associated with MF have been identified. Non-exclusive examples of genes associated with MF include, for example, JAK2, MPL, CLAR, LNK, ASXL1, SRSF2, PPM1D, IDH1 / 2, TET2, EZH2, U2AF1, NFE2, SH2B3, SF3B1, or CBL. In some embodiments, subjects have or are suspected of having MF, which includes one or more mutations in one or more of the genes described herein.

[0162] In some embodiments, the subject has one or more mutations in genes involved in epigenetic regulation or splicing. In some embodiments, the one or more mutations in genes involved in epigenetic regulation or splicing are ASXL1, DNMT3A, TET2, SRSF2, U2AF1, EZH2, or SF3B1. In some embodiments, the subject has mutations in IDH1 / 2, which are associated with a risk of progression to MBN-BP.

[0163] In some embodiments, this disclosure relates to compositions and methods for treating myelofibrosis in subjects. In some embodiments, subjects treated according to this disclosure may be identified based on appropriate diagnostic or prognostic methods. For example, the Dynamic International Prognostic Scoring System (DIPSS) and age-adjusted DIPSS provide models of patient outcomes based on several patient-specific variables, including age, hemoglobin levels, leukocyte count, peripheral hemoblasts, and systemic symptoms (see, for example, Passamonti, F., et al. Blood. 2010 Mar 4;115(9):1703-8, incorporated herein by reference). The DIPSS model enables the calculation of a DIPSS score and the assignment of patients to risk categories for prognostic purposes. A DIPSS score of 0 indicates a “low-risk” patient, a DIPSS score of 1–2 indicates an “intermediate-1 risk” patient, a DIPSS score of 3–4 indicates an “intermediate-2 risk” patient, and a DIPSS score of 5–6 indicates a “high-risk” patient. Therefore, in some embodiments, subjects requiring treatment according to this application may have a DIPSS score of at least 1. In some embodiments, subjects have a DIPSS score of 1–4 (e.g., 1, 2, 3, or 4). In some embodiments, the DIPSS score is 5 or 6 (e.g., 5 or 6).

[0164] In some embodiments, the subjects treated in accordance with this disclosure may be evaluated by appropriate diagnostic or prognostic methods. For example, the Myeloproliferative Neoplasm-Symptom Assessment Form Total Symptom Score (MPN-SAF TSS) provides a 10-item tool designed to assess most representative and clinically relevant symptoms among patients with MPN. The tool records a patient's assessment of the incidence and severity of these disease-related symptoms. It can be used to track symptoms over time and guide subsequent management decisions (see, for example, Emanuel RM, et al. Myeloproliferative neoplasm (MPN) symptom assessment form total symptom score: prospective international assessment of an abbreviated symptom burden scoring system among patients with MPNs, J Clin Oncol. 2012;30(33):4098-4103, incorporated herein by reference). The MPN-SAF TSS includes symptoms such as fatigue, early satiety, lethargy, lack of concentration, abdominal discomfort, night sweats, bone pain, itching, unintentional weight loss, and fever. Each symptom is rated on a severity scale from 0 (none / best possible) to 10 (worst possible / worst possible). The MPN-SAF TSS has a possible range of 0 to 100, with 100 representing the highest level of symptom severity. In some embodiments, the Myelofibrosis Symptom Assessment Form (MFSAF) is derived from the MPN-SAF TSS. The MFSAF is a means of measuring symptoms reported by more than 10% of MF patients and includes quality of life (QoL) measurements.The MFSAF includes a comprehensive assessment of fatigue, an assessment of splenomegaly and associated organic symptoms, and an assessment of other symptoms such as night sweats, itching (pruritus), bone pain, fever, unintentional weight loss, and overall quality of life (see, for example, Mesa et al., The Myelofibrosis Symptom Assessment Form (MFSAF): An Evidence-based Brief Inventory to Measure Quality of Life and Symptomatic Response to Treatment in Myelofibrosis, Leuk Res. 2009 Sep; 33(9): 1199-1203, incorporated herein by reference). Symptoms are assessed on a severity scale from 0 (none / best possible) to 10 (worst possible / worst possible). The MFSAF can be used to track symptoms over time and guide decisions for subsequent management.

[0165] In some embodiments, subjects with MF (e.g., PMF or SMF) develop anemia. Anemia in MF is a result of a multifactorial process. In some embodiments, anemia in MF is caused by increased red blood cell destruction due to ineffective erythropoiesis due to bone marrow suppression and iron metabolism deficiency, splenomegaly, increased plasma volume, an abnormally pro-inflammatory environment in the bone marrow, or a combination thereof. In some embodiments, among other causes, anemia in MF is associated with abnormal iron metabolism. In some embodiments, abnormal iron metabolism in MF patients is functional iron deficiency (FID). FID represents a state of iron-restricted erythropoiesis characterized by an imbalance between the iron required for effective red blood cell production and readily available serum iron. In FID, iron is sequestrated and unavailable for red blood cell production, even when the body has sufficient or increased systemic iron stores. In some embodiments, FID is caused by increased hepcidin compared to iron storage levels. In some embodiments, upregulation of inflammatory cytokines in the bone marrow of MF patients is also associated with upregulation of circulating hepcidin, resulting in FID. In some embodiments, anemia in MF may be treatment-related. In some embodiments, MF patients have been previously treated with a JAK inhibitor (e.g., ruxolitinib or fedratinib). In some embodiments, patients receiving a JAK inhibitor (e.g., ruxolitinib or fedratinib) are more likely to develop MF-associated anemia. Inhibition of the JAK-STAT signaling pathway results in inhibition of erythropoietin-mediated JAK2 signaling, which is essential for erythropoiesis.In some embodiments, initial anemia has been identified as a major adverse event associated with JAK inhibitor (e.g., ruxolitinib) treatment (the full contents of each of these are incorporated herein by reference: Verstovsek S, Kantarjian H, Mesa RA, et al. Safety and efficacy of INCB018424, a JAK1 and JAK2 inhibitor, in myelofibrosis. N Engl J Med. 2010;363(12):1117-1127; Verstovsek S, Mesa RA, Gotlib J, et al. A doubleblind, placebo-controlled trial of ruxolitinib for myelofibrosis. N Engl J Med. 2012;366(9):799-807; Parganas E, Wang D, Stravopodis D, et al. Jak2 is essential for signaling through a variety of cytokine receptors. Cell. (See Cell. 1998;93(3):397-409).

[0166] In some embodiments, subjects have or are at risk of having systemic or microvascular symptoms associated with myeloproliferative neoplasms (MPN). In some embodiments, subjects have or are at risk of having thromboembolic or hemorrhagic complications. In some embodiments, subjects have or are at risk of having acute myeloid leukemia (AML) in the acute transition phase of MPN. In some embodiments, subjects exhibit ribosomal disease in megakaryocytes. In some embodiments, subjects exhibit decreased GATA1 expression, particularly in megakaryocytes. In some embodiments, subjects exhibit a defect in megakaryocyte function or maturation. In some embodiments, subjects do not have nutritional iron deficiency. In some embodiments, subjects exhibit thrombocytopenia, anemia, and / or neutropenia.

[0167] Therefore, in some embodiments, subjects requiring treatment in accordance with this disclosure have previously received therapeutic intervention for a blood disorder. In some embodiments, subjects have previously undergone surgical procedures to treat one or more blood disorders. In some embodiments, subjects have previously undergone splenectomy. In some embodiments, subjects have previously received therapeutic agents to treat one or more blood disorders.

[0168] In some embodiments, the subject has previously received an immunomodulator or erythropoietin stimulant, such as danazol, prednisone, thalidomide, lenalidomide, or pomalidomide.

[0169] In some embodiments, the subject has previously received a JAK-STAT pathway inhibitor; in some embodiments, the JAK-STAT pathway inhibitor is either a JAK inhibitor or a STAT inhibitor. In some embodiments, the JAK inhibitor is selective for one or both of subtypes JAK1 and JAK2 (e.g., a JAK1 / 2 inhibitor). In some embodiments, the STAT inhibitor is a STAT3 inhibitor. In some embodiments, the JAK1 / 2 or STAT3 inhibitor is selected from the group consisting of ruxolitinib, momerotinib, pacritinib, INCB039110, AG490, and PpYLKTK. In some embodiments, the subject received a JAK / STAT antagonist as treatment for polycythemia vera (PV), essential thrombocythemia (ET), or pre-fibrotic / early primary myelofibrosis (pre-MF). In some embodiments, the subject received treatment with a JAK / STAT antagonist for 2–6 weeks. In some embodiments, the subject receiving the JAK-STAT pathway inhibitor has anemia. In some embodiments, anemia in subjects receiving JAK-STAT pathway inhibitors is not relieved by the JAK-STAT inhibitors. In some embodiments, anemia in subjects receiving JAK-STAT pathway inhibitors is more severe than in subjects not receiving JAK-STAT inhibitors.

[0170] In some embodiments, the subject has previously received a growth factor ligand trap. In some embodiments, the growth factor ligand trap is a transformed growth factor beta (TGF-β) ligand trap. In some embodiments, the TGF-β ligand trap is sotatercept or raspatercept. In some embodiments, the subject has previously received an antifibrotic agent. In some embodiments, the antifibrotic agent is PRM-151.

[0171] In some embodiments, the Disclosure provides compositions and methods for treating subjects who are known to have or suspected to have a blood disorder characterized by low systemic iron levels (e.g., MF-associated anemia). In some embodiments, the subjects have one or more conditions resulting from myelofibrosis and / or myelofibrosis as otherwise described herein. In some embodiments, the anemia in the subjects is addressed by red blood cell transfusion. In some embodiments, the subjects are red blood cell transfusion-dependent. "Transfusion-dependent" may refer to a patient with a red blood cell transfusion frequency in which at least two units of packed red blood cells are transfused every four weeks on average over a preceding 12-week period. A transfusion-dependent patient may not receive red blood cell transfusions for four or six consecutive weeks over a preceding 12 or 24-week period. In some embodiments, the subjects are red blood cell transfusion-independent. "Transfusion-independent" may refer to a patient who is anemic (e.g., Hgb levels of ≤11 g / dL, ≤10 g / dL, or ≤9 g / dL). A patient is defined as someone who may receive intermittent transfusions, is anemic, and does not meet either the criteria for transfusion-dependent or transfusion-independent. In some embodiments, the subject receives multiple transfusions over a 12-week period. In some embodiments, the subject receives at least four RBC transfusions over a 12-week period. In some embodiments, the subject receives at least one transfusion of 2 units of concentrated red blood cells over 4, 6, or 8 weeks, and in some embodiments, the subject also receives at least 4, 6, or 8 units of concentrated red blood cell transfusions over a 12-week period. In some embodiments, the subject may have a transfusion load reduced by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or more. In some embodiments, the subject is transfusion-independent (e.g., does not receive repeated red blood cell transfusions over a 12-week period). In some embodiments, the subjects are anemic (e.g., Hgb level below 10 g / dL) and receive blood transfusions as needed, but in smaller quantities than the 6 units of 12-week concentrated red blood cells mentioned earlier.

[0172] In some embodiments, this disclosure relates to compositions and methods for treating myelofibrosis-associated anemia. Any anemic condition described herein may be characterized by one or more hematological criteria described herein. In some embodiments, myelofibrosis-associated anemia may be characterized as mild to moderate or severe anemia according to appropriate diagnostic threshold parameters. For example, in some embodiments, myelofibrosis-associated anemia is characterized based on hemoglobin (Hgb) levels, and the severity of anemia increases with decreasing Hgb levels. In some embodiments, mild to moderate anemia is associated with Hgb levels of at least 8 g / dL and below the lower limit of normal (e.g., between about 8 g / dL and about 14 g / dL, between about 8 g / dL and about 12 g / dL, between about 8 g / dL and about 10 g / dL, between about 10 g / dL and about 14 g / dL, or between about 10 g / dL and about 12 g / dL). In some embodiments, severe anemia is associated with Hgb levels of approximately 8 g / dL or less (e.g., between approximately 2 g / dL and 8 g / dL, between approximately 4 g / dL and 8 g / dL, between approximately 6 g / dL and 8 g / dL, between approximately 2 g / dL and 4 g / dL, or between approximately 1.5 g / dL and 2 g / dL). In some embodiments, severe anemia is associated with red blood cell transfusion dependence. In some embodiments, severe anemia is associated with red blood cell transfusion independence resulting from a therapeutic intervention (e.g., therapeutic recovery from a transfusion-dependent state), and the subject depends on ongoing therapeutic procedures to maintain transfusion independence.

[0173] In some embodiments, myelofibrosis-associated anemia is also characterized based on ferritin levels. Ferritin is an iron-containing blood protein, and its level indicates how much iron is stored in the body. In some embodiments, subjects have serum ferritin levels within or above the normal range. The normal range for ferritin is 24–336 μg / L for men and 11–307 μg / L for women. In some embodiments, the target is higher than 100 μg / L (for example, between approximately 100 μg / L and approximately 110 μg / L, between approximately 100 μg / L and approximately 120 μg / L, between approximately 100 μg / L and approximately 130 μg / L, between approximately 100 μg / L and approximately 140 μg / L, between approximately 100 μg / L and approximately 150 μg / L, between approximately 110 μg / L and approximately 120 μg / L, between approximately 110 μg / L and approximately 130 μg / L, between approximately 110 μg / L and approximately 140 μg / L, between approximately 110 μg / L and approximately Between 150 μg / L, between approximately 120 μg / L and 130 μg / L, between approximately 120 μg / L and 140 μg / L, between approximately 120 μg / L and 150 μg / L, between approximately 130 μg / L and 140 μg / L, between approximately 130 μg / L and 150 μg / L, between approximately 140 μg / L and 150 μg / L, higher than 150 μg / L (for example, between approximately 150 μg / L and 160 μg / L, between approximately 150 μg / L and 170 μg / L, between approximately 150 μg / L and 180 μg Between / L, between approximately 150 μg / L and approximately 190 μg / L, between approximately 150 μg / L and approximately 200 μg / L, between approximately 160 μg / L and approximately 170 μg / L, between approximately 160 μg / L and approximately 180 μg / L, between approximately 160 μg / L and approximately 190 μg / L, between approximately 160 μg / L and approximately 200 μg / L, between approximately 170 μg / L and approximately 180 μg / L, between approximately 170 μg / L and approximately 190 μg / L, between approximately 170 μg / L and approximately 200 μg / L, between approximately 180 μg / L and approximately 190 μg Between / L, between approximately 180μg / L and approximately 200μg / L, between approximately 190μg / L and approximately 200μg / L), higher than 200μg / L (for example, between approximately 200μg / L and approximately 210μg / L, between approximately 200μg / L and approximately 220μg / L, between approximately 200μg / L and approximately 230μg / L, between approximately 200μg / L and approximately 240μg / L, between approximately 200μg / L and approximately 250μg / L, between approximately 210μg / L and approximately 220μg / L, between approximately 210μg / L and approximately 230μg / L,Between approximately 210 μg / L and approximately 240 μg / L, between approximately 210 μg / L and approximately 250 μg / L, between approximately 220 μg / L and approximately 230 μg / L, between approximately 220 μg / L and approximately 240 μg / L, between approximately 220 μg / L and approximately 250 μg / L, between approximately 230 μg / L and approximately 240 μg / L, between approximately 230 μg / L and approximately 250 μg / L, between approximately 240 μg / L and approximately 250 μg / L, higher than 250 μg / L (for example, between approximately 250 μg / L and approximately 260 μg / L, between approximately 250 μg / L and approximately 270 μg / L, between approximately 250 μg / L and approximately 280 μg / L, between approximately 250 μg / L and approximately 2 Between 90 μg / L, between approximately 250 μg / L and 300 μg / L, between approximately 260 μg / L and 270 μg / L, between approximately 260 μg / L and 280 μg / L, between approximately 260 μg / L and 290 μg / L, between approximately 260 μg / L and 300 μg / L, between approximately 270 μg / L and 280 μg / L, between approximately 270 μg / L and 290 μg / L, between approximately 270 μg / L and 300 μg / L, between approximately 280 μg / L and 290 μg / L, between approximately 280 μg / L and 300 μg / L, between approximately 290 μg / L and 300 μg / L), higher than 300 μg / L (for example, approximately 3 Between 00 μg / L and approximately 310 μg / L, between approximately 300 μg / L and approximately 320 μg / L, between approximately 300 μg / L and approximately 330 μg / L, between approximately 300 μg / L and approximately 340 μg / L, between approximately 300 μg / L and approximately 350 μg / L, between approximately 310 μg / L and approximately 320 μg / L, between approximately 310 μg / L and approximately 330 μg / L, between approximately 310 μg / L and approximately 340 μg / L, between approximately 310 μg / L and approximately 350 μg / L, between approximately 320 μg / L and approximately 330 μg / L, between approximately 320 μg / L and approximately 340 μg / L, between approximately 320 μg / L and approximately 350 μg / L, and approximately 330 μg / L. Between approximately 340 μg / L, between approximately 330 μg / L and approximately 350 μg / L, between approximately 340 μg / L and approximately 350 μg / L), higher than 350 μg / L (for example, between approximately 350 μg / L and approximately 360 μg / L, between approximately 350 μg / L and approximately 370 μg / L, between approximately 350 μg / L and approximately 380 μg / L, between approximately 350 μg / L and approximately 390 μg / L, between approximately 350 μg / L and approximately 400 μg / L, between approximately 360 μg / L and approximately 370 μg / L, between approximately 360 μg / L and approximately 380 μg / L, between approximately 360 μg / L and approximately 390 μg / L, between approximately 360 μg / L and approximately 400 μg / L,Between approximately 370 μg / L and 380 μg / L, between approximately 370 μg / L and 390 μg / L, between approximately 370 μg / L and 400 μg / L, between approximately 380 μg / L and 390 μg / L, between approximately 380 μg / L and 400 μg / L, between approximately 390 μg / L and 400 μg / L, higher than 400 μg / L (for example, between approximately 400 μg / L and 410 μg / L, between approximately 400 μg / L and 420 μg / L, between approximately 400 μg / L and 430 μg / L, between approximately 400 μg / L and 440 μg / L, between approximately 400 μg / L and 450 μg / L, between approximately 410 μg / L and 4 Between 20 μg / L, between approximately 410 μg / L and 430 μg / L, between approximately 410 μg / L and 440 μg / L, between approximately 410 μg / L and 450 μg / L, between approximately 420 μg / L and 430 μg / L, between approximately 420 μg / L and 440 μg / L, between approximately 420 μg / L and 450 μg / L, between approximately 430 μg / L and 440 μg / L, between approximately 430 μg / L and 450 μg / L, between approximately 440 μg / L and 450 μg / L), higher than 450 μg / L (for example, between approximately 450 μg / L and 460 μg / L, between approximately 450 μg / L and 470 μg / L, approximately 4 Between 50 μg / L and approximately 480 μg / L, between approximately 450 μg / L and approximately 490 μg / L, between approximately 450 μg / L and approximately 500 μg / L, between approximately 460 μg / L and approximately 470 μg / L, between approximately 460 μg / L and approximately 480 μg / L, between approximately 460 μg / L and approximately 490 μg / L, between approximately 460 μg / L and approximately 500 μg / L, between approximately 470 μg / L and approximately 480 μg / L, between approximately 470 μg / L and approximately 490 μg / L, between approximately 470 μg / L and approximately 500 μg / L, between approximately 480 μg / L and approximately 490 μg / L, between approximately 480 μg / L and approximately 500 μg / L, and approximately 490 μg / L (between approximately 500 μg / L), higher than 500 μg / L (for example, between approximately 500 μg / L and approximately 510 μg / L, between approximately 500 μg / L and approximately 520 μg / L, between approximately 500 μg / L and approximately 530 μg / L, between approximately 500 μg / L and approximately 540 μg / L, between approximately 500 μg / L and approximately 550 μg / L, between approximately 510 μg / L and approximately 520 μg / L, between approximately 510 μg / L and approximately 530 μg / L, between approximately 510 μg / L and approximately 540 μg / L, between approximately 510 μg / L and approximately 550 μg / L, between approximately 520 μg / L and approximately 530 μg / L, between approximately 520 μg / L and approximately 540 μg / L,Between approximately 520 μg / L and approximately 550 μg / L, between approximately 530 μg / L and approximately 540 μg / L, between approximately 530 μg / L and approximately 550 μg / L, between approximately 540 μg / L and approximately 550 μg / L, higher than 550 μg / L (for example, between approximately 550 μg / L and approximately 560 μg / L, between approximately 550 μg / L and approximately 570 μg / L, between approximately 550 μg / L and approximately 580 μg / L, between approximately 550 μg / L and approximately 590 μg / L, between approximately 550 μg / L and approximately 600 μg / L, between approximately 560 μg / L and approximately 570 μg / L, between approximately 560 μg / L and approximately 580 μg / L, between approximately 560 μg / L and approximately 5 Between 90 μg / L, between approximately 560 μg / L and 600 μg / L, between approximately 570 μg / L and 580 μg / L, between approximately 570 μg / L and 590 μg / L, between approximately 570 μg / L and 600 μg / L, between approximately 580 μg / L and 590 μg / L, between approximately 580 μg / L and 600 μg / L, between approximately 590 μg / L and 600 μg / L), higher than 600 μg / L (for example, between approximately 600 μg / L and 610 μg / L, between approximately 600 μg / L and 620 μg / L, between approximately 600 μg / L and 630 μg / L, between approximately 600 μg / L and 640 μg / L, approximately 6 Between 00 μg / L and approximately 650 μg / L, between approximately 610 μg / L and approximately 620 μg / L, between approximately 610 μg / L and approximately 630 μg / L, between approximately 610 μg / L and approximately 640 μg / L, between approximately 610 μg / L and approximately 650 μg / L, between approximately 620 μg / L and approximately 630 μg / L, between approximately 620 μg / L and approximately 640 μg / L, between approximately 620 μg / L and approximately 650 μg / L, between approximately 630 μg / L and approximately 640 μg / L, between approximately 630 μg / L and approximately 650 μg / L, between approximately 640 μg / L and approximately 650 μg / L), higher than 650 μg / L (for example, between approximately 350 μg / L and approximately 360 μg / L) Between μg / L, between approximately 350 μg / L and approximately 370 μg / L, between approximately 350 μg / L and approximately 380 μg / L, between approximately 350 μg / L and approximately 390 μg / L, between approximately 350 μg / L and approximately 400 μg / L, between approximately 360 μg / L and approximately 370 μg / L, between approximately 360 μg / L and approximately 380 μg / L, between approximately 360 μg / L and approximately 390 μg / L, between approximately 360 μg / L and approximately 400 μg / L, between approximately 370 μg / L and approximately 380 μg / L, between approximately 370 μg / L and approximately 390 μg / L, between approximately 370 μg / L and approximately 400 μg / L, between approximately 380 μg / L and approximately 390 μg / L,Between approximately 380 μg / L and approximately 400 μg / L, between approximately 390 μg / L and approximately 400 μg / L), higher than 700 μg / L (for example, between approximately 700 μg / L and approximately 710 μg / L, between approximately 700 μg / L and approximately 720 μg / L, between approximately 700 μg / L and approximately 730 μg / L, between approximately 700 μg / L and approximately 740 μg / L, between approximately 700 μg / L and approximately 750 μg / L, between approximately 710 μg / L and approximately 720 μg / L, between approximately 710 μg / L and approximately 730 μg / L, between approximately 710 μg / L and approximately 740 μg / L, between approximately 710 μg / L and approximately 750 μg / L, between approximately 720 μg / L and approximately 7 Between 30 μg / L, between approximately 720 μg / L and 740 μg / L, between approximately 720 μg / L and 750 μg / L, between approximately 730 μg / L and 740 μg / L, between approximately 730 μg / L and 750 μg / L, between approximately 740 μg / L and 750 μg / L), higher than 750 μg / L (for example, between approximately 750 μg / L and 760 μg / L, between approximately 750 μg / L and 770 μg / L, between approximately 750 μg / L and 780 μg / L, between approximately 750 μg / L and 790 μg / L, between approximately 750 μg / L and 800 μg / L, between approximately 760 μg / L and 770 μg / L, approximately 7 Between 60 μg / L and approximately 780 μg / L, between approximately 760 μg / L and approximately 790 μg / L, between approximately 760 μg / L and approximately 800 μg / L, between approximately 770 μg / L and approximately 780 μg / L, between approximately 770 μg / L and approximately 790 μg / L, between approximately 770 μg / L and approximately 800 μg / L, between approximately 780 μg / L and approximately 790 μg / L, between approximately 780 μg / L and approximately 800 μg / L, between approximately 790 μg / L and approximately 800 μg / L, higher than 800 μg / L (for example, between approximately 800 μg / L and approximately 810 μg / L, between approximately 800 μg / L and approximately 820 μg / L, between approximately 800 μg / L and approximately 830 μg / L Between μg / L, between approximately 800 μg / L and approximately 840 μg / L, between approximately 800 μg / L and approximately 850 μg / L, between approximately 810 μg / L and approximately 820 μg / L, between approximately 810 μg / L and approximately 830 μg / L, between approximately 810 μg / L and approximately 840 μg / L, between approximately 810 μg / L and approximately 850 μg / L, between approximately 820 μg / L and approximately 830 μg / L, between approximately 820 μg / L and approximately 840 μg / L, between approximately 820 μg / L and approximately 850 μg / L, between approximately 830 μg / L and approximately 840 μg / L, between approximately 830 μg / L and approximately 850 μg / L, between approximately 840 μg / L and approximately 850 μg / L),Higher than 850 μg / L (for example, between approximately 850 μg / L and 860 μg / L, between approximately 850 μg / L and 870 μg / L, between approximately 850 μg / L and 880 μg / L, between approximately 850 μg / L and 890 μg / L, between approximately 850 μg / L and 900 μg / L, between approximately 860 μg / L and 870 μg / L, approximately 860 μg / L, Between L and approximately 880 μg / L, between approximately 860 μg / L and approximately 890 μg / L, between approximately 860 μg / L and approximately 800 μg / L, between approximately 870 μg / L and approximately 880 μg / L, between approximately 870 μg / L and approximately 890 μg / L, between approximately 870 μg / L and approximately 900 μg / L, between approximately 880 μg / L and approximately 890 μg / L, between approximately 880 μg / L and approximately 900 μg / L, between approximately 890 μg / L and approximately 900 μg / L, higher than 900 μg / L (for example, between approximately 900 μg / L and approximately 910 μg / L, between approximately 900 μg / L and approximately 920 μg / L, between approximately 900 μg / L and approximately 930 μg / L) Between approximately 900 μg / L and 940 μg / L, between approximately 900 μg / L and 950 μg / L, between approximately 910 μg / L and 920 μg / L, between approximately 910 μg / L and 930 μg / L, between approximately 910 μg / L and 940 μg / L, between approximately 910 μg / L and 950 μg / L, between approximately 920 μg / L and 930 μg / L, between approximately 920 μg / L and 940 μg / L, between approximately 920 μg / L and 950 μg / L, between approximately 930 μg / L and 940 μg / L, between approximately 930 μg / L and 950 μg / L, between approximately 940 μg / L and 950 μg / L), 95 Higher than 0 μg / L (for example, between approximately 950 μg / L and 960 μg / L, between approximately 950 μg / L and 970 μg / L, between approximately 950 μg / L and 380 μg / L, between approximately 950 μg / L and 990 μg / L, between approximately 950 μg / L and 1000 μg / L, between approximately 960 μg / L and 970 μg / L, between approximately 960 μg / L and 980 μg / L, between approximately 960 μg / L and 990 μg / L, between approximately 960 μg / L and 1000 μg / L, between approximately 970 μg / L and 980 μg / L, between approximately 970 μg / L and 990 μg / L, between approximately 970 μg / L and 1 Between 000 μg / L, between approximately 980 μg / L and 990 μg / L, between approximately 980 μg / L and 1000 μg / L, between approximately 990 μg / L and 1000 μg / L), or higher than 1000 μg / L (for example, between approximately 1000 μg / L and 1500 μg / L, between approximately 1000 μg / L and 2000 μg / L, between approximately 1000 μg / L and 2500 μg / L, between approximately 1000 μg / L and 3000 μg / L, between approximately 1000 μg / L and 4000 μg / L, between approximately 1000 μg / L and 5000 μg / L, between approximately 2000 μg / L and 3000 μg / L,The serum ferritin level is between approximately 2000 μg / L and 4000 μg / L, between approximately 2000 μg / L and 5000 μg / L, between approximately 3000 μg / L and 4000 μg / L, and between approximately 4000 μg / L and 5000 μg / L. In some embodiments, the subject has serum ferritin levels between approximately 100 μg / L and approximately 1000 μg / L, between approximately 100 μg / L and approximately 500 μg / L, between approximately 100 μg / L and approximately 500 μg / L, between approximately 200 μg / L and approximately 1000 μg / L, between approximately 200 μg / L and approximately 500 μg / L, between approximately 300 μg / L and approximately 1000 μg / L, between approximately 300 μg / L and approximately 500 μg / L, between approximately 400 μg / L and approximately 1000 μg / L, and between approximately 400 μg / L and approximately 500 μg / L. However, naturally, other suitable markers (e.g., TSAT%, serum iron level, total iron-binding capacity (TIBC), hemoglobin level, liver iron content, reticulocyte hemoglobin content, hepcidin level, IL-6 level, creatinine level, etc.) may be evaluated to determine whether the subject is suitable for the treatment methods described herein.

[0174] In some embodiments, myelofibrosis-associated anemia is characterized based on reticulocyte hemoglobin content (RET-He or CHr). Reticulocyte hemoglobin content measures the amount of hemoglobin in reticulocytes. The normal range for CHr is approximately 28–36 pg / cell. In some embodiments, subjects have a CHr lower than the normal range. In some embodiments, the target range is lower than 36 pg / ml, lower than 35 pg / ml, lower than 34 pg / ml, lower than 33 pg / ml, lower than 32 pg / ml, lower than 31 pg / ml, lower than 30 pg / ml, lower than 29 pg / ml, lower than 28 pg / ml, lower than 27 pg / ml, lower than 26 pg / ml, lower than 25 pg / ml, lower than 24 pg / ml, lower than 23 pg / ml, lower than 21 pg / ml, lower than 20 pg / ml, lower than 19 pg / ml, and 18 pg / ml. It has a CHr level lower than l, lower than 17 pg / ml, lower than 16 pg / ml, lower than 15 pg / ml, lower than 14 pg / ml, lower than 13 pg / ml, lower than 12 pg / ml, lower than 11 pg / ml, lower than 10 pg / ml, lower than 9 pg / ml, lower than 8 pg / ml, lower than 7 pg / ml, lower than 6 pg / ml, lower than 5 pg / ml, lower than 4 pg / ml, lower than 3 pg / ml, lower than 2 pg / ml, or lower than 1 pg / ml.In some embodiments, the target range is approximately 1 pg / ml to 36 pg / ml, 1 pg / ml to 32 pg / ml, 1 pg / ml to 30 pg / ml, 1 pg / ml to 28 pg / ml, 1 pg / ml to 25 pg / ml, 1 pg / ml to 20 pg / ml, 1 pg / ml to 15 pg / ml, 1 pg / ml to 12 pg / ml, 1 pg / ml to 10 pg / ml, 1 pg / ml to 8 pg / ml, 1 pg / ml to 6 pg / ml, and 1 pg / ml to 4 pg / ml. , between approximately 5 pg / ml and approximately 36 pg / ml, between approximately 5 pg / ml and approximately 32 pg / ml, between approximately 5 pg / ml and approximately 30 pg / ml, between approximately 5 pg / ml and approximately 28 pg / ml, between approximately 5 pg / ml and approximately 25 pg / ml, between approximately 5 pg / ml and approximately 20 pg / ml, between approximately 5 pg / ml and approximately 15 pg / ml, between approximately 5 pg / ml and approximately 12 pg / ml, between approximately 5 pg / ml and approximately 10 pg / ml, between approximately 5 pg / ml and approximately 8 pg / ml, between approximately 5 pg / ml and approximately 6 pg / ml, between approximately 10 pg / ml and approximately 36 pg / ml, between approximately 10 pg / ml and approximately Between 32 pg / ml, between approximately 10 pg / ml and 30 pg / ml, between approximately 1 pg / ml and 28 pg / ml, between approximately 10 pg / ml and 25 pg / ml, between approximately 10 pg / ml and 20 pg / ml, between approximately 10 pg / ml and 15 pg / ml, between approximately 10 pg / ml and 12 pg / ml, between approximately 15 pg / ml and 36 pg / ml, between approximately 15 pg / ml and 32 pg / ml, between approximately 15 pg / ml and 30 pg / ml, between approximately 15 pg / ml and 28 pg / ml, between approximately 15 pg / ml and 25 pg / ml, and between approximately 15 pg / ml and It contains CHr in the following ranges: approximately 20 pg / ml, approximately 20 pg / ml to approximately 36 pg / ml, approximately 20 pg / ml to approximately 32 pg / ml, approximately 20 pg / ml to approximately 30 pg / ml, approximately 20 pg / ml to approximately 28 pg / ml, approximately 20 pg / ml to approximately 25 pg / ml, approximately 25 pg / ml to approximately 36 pg / ml, approximately 25 pg / ml to approximately 32 pg / ml, approximately 25 pg / ml to approximately 30 pg / ml, approximately 25 pg / ml to approximately 28 pg / ml, approximately 30 pg / ml to approximately 36 pg / ml, and approximately 30 pg / ml to approximately 32 pg / ml.However, naturally, other suitable markers (e.g., TSAT%, serum iron level, total iron-binding capacity (TIBC), ferritin level, hemoglobin level, liver iron content, hepcidin level, IL-6 level, creatinine level, etc.) may be evaluated to determine whether the subject is suitable for the treatment methods described herein.

[0175] In some embodiments, myelofibrosis-associated anemia is characterized by liver iron levels. In some embodiments, the normal range for liver iron levels is 200–2400 μg / g dry weight in men and 400–1600 μg / g dry weight in women. In some embodiments, the subject has liver iron levels higher than normal. In some embodiments, the patient may have a blood sugar level higher than 200 μg / g dry weight (for example, between approximately 200 μg / g and approximately 250 μg / g dry weight, between approximately 200 μg / g and approximately 250 μg / g dry weight, between approximately 200 μg / g and approximately 300 μg / g dry weight, between approximately 220 μg / g and approximately 250 μg / g dry weight, between approximately 220 μg / g and approximately 300 μg / g dry weight, between approximately 250 μg / g and approximately 300 μg / g dry weight, between approximately 260 μg / g and approximately 300 μg / g dry weight, or between approximately 280 μg / g (between g and approximately 300 μg / g dry weight), higher than 300 μg / g dry weight (for example, between approximately 300 μg / g and 320 μg / g dry weight, between approximately 300 μg / g and 350 μg / g dry weight, between approximately 300 μg / g and 400 μg / g dry weight, between approximately 320 μg / g and 350 μg / g dry weight, between approximately 320 μg / g and 400 μg / g dry weight, between approximately 350 μg / g and 400 μg / g dry weight, between approximately 360 μg / g and 400 μg / g dry weight, or between approximately 380 μg / g and 400 μg / g dry weight) (between μg / g dry weight), higher than 400 μg / g dry weight (for example, between approximately 400 μg / g and 420 μg / g dry weight, between approximately 400 μg / g and 450 μg / g dry weight, between approximately 400 μg / g and 500 μg / g dry weight, between approximately 420 μg / g and 450 μg / g dry weight, between approximately 420 μg / g and 500 μg / g dry weight, between approximately 450 μg / g and 500 μg / g dry weight, between approximately 460 μg / g and 500 μg / g dry weight, or between approximately 480 μg / g and 500 μg / g dry weight) (Between weights), higher than 500 μg / g dry weight (for example, between approximately 500 μg / g and 520 μg / g dry weight, between approximately 500 μg / g and 550 μg / g dry weight, between approximately 500 μg / g and 600 μg / g dry weight, between approximately 520 μg / g and 550 μg / g dry weight, between approximately 520 μg / g and 600 μg / g dry weight, between approximately 550 μg / g and 600 μg / g dry weight, between approximately 560 μg / g and 600 μg / g dry weight, or between approximately 580 μg / g and 600 μg / g dry weight),Higher than 600 μg / g dry weight (for example, between approximately 600 μg / g and 620 μg / g dry weight, between approximately 600 μg / g and 650 μg / g dry weight, between approximately 600 μg / g and 700 μg / g dry weight, between approximately 620 μg / g and 650 μg / g dry weight, between approximately 620 μg / g and 700 μg / g dry weight, between approximately 650 μg / g and 700 μg / g dry weight, between approximately 660 μg / g and 700 μg / g dry weight, or between approximately 680 μg / g and 700 μg / g dry weight), higher than 700 μg / g dry weight (for example, between approximately 700 μg / g and 720 μg / g dry weight) Between weight, between approximately 700 μg / g and 750 μg / g dry weight, between approximately 700 μg / g and 800 μg / g dry weight, between approximately 720 μg / g and 750 μg / g dry weight, between approximately 720 μg / g and 800 μg / g dry weight, between approximately 750 μg / g and 800 μg / g dry weight, between approximately 760 μg / g and 800 μg / g dry weight, or between approximately 780 μg / g and 800 μg / g dry weight), higher than 800 μg / g dry weight (for example, between approximately 800 μg / g and 820 μg / g dry weight, between approximately 800 μg / g and 850 μg / g dry weight, approximately 800 μg / g and Between 900 μg / g dry weight, between approximately 820 μg / g and 850 μg / g dry weight, between approximately 820 μg / g and 900 μg / g dry weight, between approximately 850 μg / g and 900 μg / g dry weight, between approximately 860 μg / g and 900 μg / g dry weight, or between approximately 880 μg / g and 900 μg / g dry weight), higher than 900 μg / g dry weight (for example, between approximately 900 μg / g and 920 μg / g dry weight, between approximately 900 μg / g and 950 μg / g dry weight, between approximately 900 μg / g and 1000 μg / g dry weight, between approximately 920 μg / g and 950 μg / g dry weight) , between approximately 920 μg / g and 1000 μg / g dry weight, between approximately 950 μg / g and 1000 μg / g dry weight, between approximately 960 μg / g and 1000 μg / g dry weight, or between approximately 980 μg / g and 1000 μg / g dry weight), higher than 1000 μg / g dry weight (for example, between approximately 1000 μg / g and 1200 μg / g dry weight, between approximately 1000 μg / g and 1500 μg / g dry weight, or between approximately 1200 μg / g and 1500 μg / g dry weight), higher than 1500 μg / g dry weight (for example, between approximately 1500 μg / g and 1800 μg / g dry weight,Between approximately 1500 μg / g and 2000 μg / g dry weight, or between approximately 1800 μg / g and 2000 μg / g dry weight, higher than 2000 μg / g dry weight (for example, between approximately 2000 μg / g and 2200 μg / g dry weight, between approximately 2000 μg / g and 2500 μg / g dry weight, or between approximately 2200 μg / g and 2500 μg / g dry weight), higher than 2500 μg / g dry weight (for example, between approximately 2500 μg / g and 2800 μg / g dry weight, between approximately 2500 μg / g and 3000 μg / g dry weight, or between approximately 2800 μg / g and 3000 μg / g dry weight) (between amounts), higher than 3000 μg / g dry weight (e.g., between approximately 3000 μg / g and 3200 μg / g dry weight, between approximately 3000 μg / g and 3500 μg / g dry weight, or between approximately 3200 μg / g and 3500 μg / g dry weight), higher than 3500 μg / g dry weight (e.g., between approximately 3500 μg / g and 3800 μg / g dry weight, between approximately 3500 μg / g and 4000 μg / g dry weight, or between approximately 3800 μg / g and 4000 μg / g dry weight), higher than 4000 μg / g dry weight (e.g., between approximately 4000 μg / g and 4200 μg / g dry weight, Between approximately 4000 μg / g and 4500 μg / g dry weight, or between approximately 4200 μg / g and 4500 μg / g dry weight, higher than 4500 μg / g dry weight (for example, between approximately 4500 μg / g and 4800 μg / g dry weight, between approximately 4500 μg / g and 5000 μg / g dry weight, or between approximately 4800 μg / g and 5000 μg / g dry weight), higher than 5000 μg / g dry weight (for example, between approximately 5000 μg / g and 5200 μg / g dry weight, between approximately 5000 μg / g and 5500 μg / g dry weight, or between approximately 5200 μg / g and 5500 μg / g dry weight) (between amounts), higher than 5500 μg / g dry weight (e.g., between approximately 5500 μg / g and 5800 μg / g dry weight, between approximately 5500 μg / g and 6000 μg / g dry weight, or between approximately 5800 μg / g and 6000 μg / g dry weight), higher than 6000 μg / g dry weight (e.g., between approximately 6000 μg / g and 6200 μg / g dry weight, between approximately 6000 μg / g and 6500 μg / g dry weight, or between approximately 6200 μg / g and 6500 μg / g dry weight), higher than 6500 μg / g dry weight (e.g., between approximately 6500 μg / g and 6800 μg / g dry weight,Between approximately 6500 μg / g and 7000 μg / g dry weight, or between approximately 6800 μg / g and 7000 μg / g dry weight, higher than 7000 μg / g dry weight (for example, between approximately 7000 μg / g and 7200 μg / g dry weight, between approximately 7000 μg / g and 7500 μg / g dry weight, or between approximately 7200 μg / g and 7500 μg / g dry weight), higher than 7500 μg / g dry weight (for example, between approximately 7500 μg / g and 7800 μg / g dry weight) Between dry weights, between approximately 7500 μg / g and 8000 μg / g dry weight, or between approximately 7800 μg / g and 8000 μg / g dry weight, higher than 8000 μg / g dry weight (for example, between approximately 8000 μg / g and 8200 μg / g dry weight, between approximately 8000 μg / g and 8500 μg / g dry weight, or between approximately 8200 μg / g and 8500 μg / g dry weight), higher than 8500 μg / g dry weight (for example, between approximately 8500 μg / g and 880 Between 0 μg / g dry weight, between approximately 8500 μg / g and 9000 μg / g dry weight, or between approximately 8800 μg / g and 9000 μg / g dry weight, higher than 9000 μg / g dry weight (e.g., between approximately 9000 μg / g and 9200 μg / g dry weight, between approximately 9000 μg / g and 9500 μg / g dry weight, or between approximately 9200 μg / g and 9500 μg / g dry weight), higher than 9500 μg / g dry weight (e.g., approximately 9500 μg Having liver iron levels between 1 / g and 9800 μg / g dry weight, between approximately 9000 μg / g and 10000 μg / g dry weight, or between approximately 9800 μg / g and 10000 μg / g dry weight, or higher than 10000 μg / g dry weight (e.g., between approximately 10000 μg / g and 15000 μg / g dry weight, between approximately 10000 μg / g and 20000 μg / g dry weight, or between approximately 10000 μg / g and 15000 μg / g dry weight). In some embodiments, the patient is given a dose between approximately 200 μg / g and approximately 500 μg / g dry weight, between approximately 200 μg / g and approximately 1000 μg / g dry weight, between approximately 200 μg / g and approximately 2000 μg / g dry weight, between approximately 200 μg / g and approximately 5000 μg / g dry weight, between approximately 200 μg / g and approximately 8000 μg / g dry weight, between approximately 200 μg / g and approximately 10000 μg / g dry weight, between approximately 500 μg / g and approximately 1000 μg / g dry weight, between approximately 500 μg / g and approximately 5000 μg / g dry weight,Liver iron levels are found to be between approximately 500 μg / g and approximately 10,000 μg / g dry weight, between approximately 1,000 μg / g and approximately 2,000 μg / g dry weight, between approximately 1,000 μg / g and approximately 5,000 μg / g dry weight, between approximately 1,000 μg / g and approximately 10,000 μg / g dry weight, between approximately 5,000 μg / g and approximately 8,000 μg / g dry weight, or between approximately 5,000 μg / g and approximately 10,000 μg / g dry weight. However, naturally, other suitable markers (e.g., TSAT%, serum iron level, total iron-binding capacity (TIBC), ferritin level, hemoglobin level, reticulocyte hemoglobin content, hepcidin level, IL-6 level, creatinine level, etc.) may be evaluated to determine whether the subject is suitable for the treatment methods described herein.

[0176] In some embodiments, myelofibrosis-associated anemia is also characterized by low serum iron levels. In some embodiments, the normal range for serum iron levels is 50–150 μg / dL dry weight for men and 35–145 μg / dL dry weight for women. In some embodiments, the subject has serum iron levels lower than normal. In some embodiments, the subject has a serum iron level lower than 150 μg / dL, lower than 140 μg / L, lower than 130 μg / L, lower than 120 μg / dL, lower than 110 μg / dL, lower than 100 μg / dL, lower than 90 μg / dL, lower than 80 μg / dL, lower than 70 μg / dL, lower than 60 μg / dL, lower than 50 μg / dL, lower than 45 μg / dL, lower than 40 μg / dL, lower than 35 μg / dL, lower than 30 μg / dL, lower than 25 μg / dL, lower than 20 μg / dL, lower than 15 μg / L, lower than 10 μg / L, or lower than 5 μg / L.In some embodiments, the target ranges are between approximately 1 μg / dL and approximately 150 μg / dL, between approximately 5 μg / dL and approximately 150 μg / dL, between approximately 10 μg / dL and approximately 150 μg / dL, between approximately 20 μg / dL and approximately 150 μg / dL, between approximately 50 μg / dL and approximately 150 μg / dL, between approximately 80 μg / dL and approximately 150 μg / dL, between approximately 100 μg / dL and approximately 150 μg / dL, between approximately 120 μg / dL and approximately 150 μg / dL, between approximately 1 μg / dL and approximately 120 μg / dL, between approximately 5 μg / dL and approximately 120 μg / dL, and between approximately 10 μg / dL and approximately 120 μg / dL. Between approximately 20 μg / dL and approximately 120 μg / dL, between approximately 50 μg / dL and approximately 120 μg / dL, between approximately 80 μg / dL and approximately 120 μg / dL, between approximately 120 μg / dL and approximately 120 μg / dL, between approximately 1 μg / dL and approximately 100 μg / dL, between approximately 5 μg / dL and approximately 100 μg / dL, between approximately 10 μg / dL and approximately 100 μg / dL, between approximately 20 μg / dL and approximately 100 μg / dL, between approximately 50 μg / dL and approximately 100 μg / dL, between approximately 80 μg / dL and approximately 100 μg / dL, between approximately 1 μg / dL and approximately 80 μg / dL, between approximately 5 μg / dL and approximately 80 μg Between / dL, between approximately 10μg / dL and approximately 80μg / dL, between approximately 20μg / dL and approximately 80μg / dL, between approximately 50μg / dL and approximately 80μg / dL, between approximately 1μg / dL and approximately 50μg / dL, between approximately 5μg / dL and approximately 50μg / dL, between approximately 10μg / dL and approximately 50μg / dL, between approximately 20μg / dL and approximately 50μg / dL, between approximately 25μg / dL and approximately 50μg / dL, between approximately 30μg / dL and approximately 50μg / dL, between approximately 1μg / dL and approximately 25μg / dL, between approximately 5μg / dL and approximately 25μg / dL, between approximately 10μg / dL and approximately 25μg / dL, The serum iron level is between approximately 10 μg / dL and 20 μg / dL, between approximately 10 μg / dL and 15 μg / dL, between approximately 1 μg / dL and 20 μg / dL, between approximately 5 μg / dL and 18 μg / dL, between approximately 10 μg / dL and 16 μg / dL, between approximately 1 μg / dL and 15 μg / dL, between approximately 5 μg / dL and 12 μg / dL, between approximately 10 μg / dL and 12 μg / dL, between approximately 1 μg / dL and 10 μg / dL, between approximately 1 μg / dL and 8 μg / dL, between approximately 1 μg / dL and 5 μg / dL, or between approximately 1 μg / dL and 3 μg / dL.However, naturally, other suitable markers (e.g., TSAT%, total iron-binding capacity (TIBC), ferritin levels, hemoglobin levels, liver iron content, reticulocyte hemoglobin content, hepcidin levels, IL-6 levels, creatinine levels, etc.) may be evaluated to determine whether the subject is suitable for the treatment methods described herein.

[0177] In some embodiments, myelofibrosis-associated anemia is characterized by a low total iron-binding capacity (TIBC). In some embodiments, the normal range for TIBC is 250–400 μg / dL. In some embodiments, subjects have a TIBC lower than normal. In some embodiments, subjects have a TIBC lower than 400 μg / dL, lower than 350 μg / dL, lower than 300 μg / dL, lower than 250 μg / dL, lower than 200 μg / dL, lower than 150 μg / dL, lower than 100 μg / dL, lower than 90 μg / dL, lower than 80 μg / dL, lower than 70 μg / dL, lower than 60 μg / dL, lower than 50 μg / dL, lower than 40 μg / dL, lower than 30 μg / dL, lower than 20 μg / dL, or lower than 10 μg / dL.In some embodiments, the target ranges are between approximately 1 μg / dL and approximately 400 μg / dL, between approximately 1 μg / dL and approximately 300 μg / dL, between approximately 1 μg / dL and approximately 200 μg / dL, between approximately 1 μg / dL and approximately 100 μg / dL, between approximately 1 μg / dL and approximately 50 μg / dL, between approximately 1 μg / dL and approximately 25 μg / dL, between approximately 1 μg / dL and approximately 10 μg / dL, between approximately 1 μg / dL and approximately 5 μg / dL, between approximately 5 μg / dL and approximately 400 μg / dL, between approximately 5 μg / dL and approximately 300 μg / dL, and between approximately 5 μg / dL and approximately 200 μg Between 5 μg / dL and 100 μg / dL, between 5 μg / dL and 50 μg / dL, between 5 μg / dL and 25 μg / dL, between 5 μg / dL and 10 μg / dL, between 10 μg / dL and 400 μg / dL, between 10 μg / dL and 300 μg / dL, between 10 μg / dL and 200 μg / dL, between 10 μg / dL and 100 μg / dL, between 10 μg / dL and 50 μg / dL, between 10 μg / dL and 25 μg / dL, and between 25 μg / dL and 400 μg / dL Between L, between approximately 25 μg / dL and approximately 300 μg / dL, between approximately 25 μg / dL and approximately 200 μg / dL, between approximately 25 μg / dL and approximately 100 μg / dL, between approximately 25 μg / dL and approximately 50 μg / dL, between approximately 50 μg / dL and approximately 400 μg / dL, between approximately 50 μg / dL and approximately 300 μg / dL, between approximately 50 μg / dL and approximately 200 μg / dL, between approximately 50 μg / dL and approximately 100 μg / dL, between approximately 100 μg / dL and approximately 400 μg / dL, between approximately 100 μg / dL and approximately 300 μg / dL, approximately 100 μg / d The TIBC is between L and approximately 200 μg / dL, between approximately 100 μg / dL and approximately 150 μg / dL, between approximately 100 μg / dL and approximately 250 μg / dL, between approximately 100 μg / dL and approximately 350 μg / dL, between approximately 200 μg / dL and approximately 250 μg / dL, between approximately 200 μg / dL and approximately 300 μg / dL, between approximately 200 μg / dL and approximately 350 μg / dL, between approximately 200 μg / dL and approximately 400 μg / dL, between approximately 300 μg / dL and approximately 350 μg / dL, or between approximately 350 μg / dL and approximately 450 μg / dL.However, naturally, other suitable markers (e.g., TSAT%, serum iron levels, ferritin levels, hemoglobin levels, liver iron content, reticulocyte hemoglobin content, hepcidin levels, IL-6 levels, creatinine levels, etc.) may be evaluated to determine whether the subject is suitable for the treatment methods described herein.

[0178] In some embodiments, myelofibrosis-associated anemia is characterized based on transferrin saturation levels (TSAT%). In some embodiments, the normal range for TSAT is approximately 20% to 50%. In some embodiments, transferrin saturation below 20% indicates iron deficiency, while in some embodiments, transferrin saturation above 50% indicates iron overload. In some embodiments, subjects have TSAT% below 100%, below 90%, below 80%, below 70%, below 60%, below 50%, below 40%, below 30%, below 20%, or below 10%. In some cases, if the subject's TSAT% is 70% or higher, 75% or higher, 80% or higher, 85% or higher, 90% or higher, or 95% or higher, ongoing treatment with a hepcidin antagonist is stopped or temporarily stopped to prevent, for example, iron overload. In some embodiments, the target is between 5% and 10%, between 5% and 20%, between 5% and 30%, between 5% and 40%, between 5% and 50%, between 5% and 60%, between 5% and 70%, between 8% and 10%, between 8% and 20%, between 8% and 30%, between 8% and 40%, between 8% and 50%, between 8% and 60%, between 8% and 70%, and between 10% and 15%. Between 10% and 20%, between 10% and 30%, between 10% and 40%, between 10% and 50%, between 10% and 60%, between 10% and 70%, between 15% and 20%, between 15% and 25%, between 15% and 30%, between 15% and 40%, between 15% and 50%, between 15% and 60%, between 15% and 70%, between 20% and 25%, between 20% and 30% Between %, between 20% and 33%, between 20% and 40%, between 20% and 50%, between 20% and 60%, between 20% and 70%, between 25% and 30%, between 25% and 35%, between 25% and 40%, between 25% and 50%, between 25% and 60%, between 25% and 70%, between 30% and 40%, between 30% and 50%, between 30% and 55%, 30% and The TSAT% has a percentage between 60%, 30% and 70%, 35% and 40%, 35% and 50%, 35% and 55%, 35% and 60%, 35% and 70%, 40% and 50%, 40% and 55%, 40% and 60%, 40% and 70%, 50% and 55%, 50% and 60%, or 50% and 70%.In other embodiments, administration of a hepcidin antagonist may be performed when the subject's TSAT% is 95% or less, 90% or less, 80% or less, 70% or less, 65% or less, 60% or less, 55% or less, 50% or less, 45% or less, 40% or less, 35% or less, or 30% or less. Thus, in some embodiments, while the patient is receiving medical treatment or care from a physician, for example, for anemia, the subject's TSAT% can be monitored, for example, continuously or periodically, to assess whether iron overload can be prevented or whether further treatment is appropriate. However, naturally, other suitable markers (e.g., serum iron levels, total iron-binding capacity (TIBC), ferritin levels, hemoglobin levels, liver iron content, reticulocyte hemoglobin content, hepcidin levels, IL-6 levels, creatinine levels, etc.) may be evaluated to determine whether the subject is suitable for the treatment methods described herein.

[0179] In some embodiments, myelofibrosis-associated anemia is also characterized by high serum hepcidin levels. In some embodiments, the normal range for hepcidin is 1–55 ng / ml. In some embodiments, subjects have serum hepcidin levels higher than normal. In some embodiments, subjects have serum hepcidin levels higher than 55 ng / ml, higher than 55 ng / ml, higher than 60 ng / ml, higher than 65 ng / ml, higher than 70 ng / ml, higher than 75 ng / ml, higher than 80 ng / ml, higher than 85 ng / ml, higher than 90 ng / ml, higher than 95 ng / ml, higher than 100 ng / ml, higher than 150 ng / ml, higher than 200 ng / ml, higher than 250 ng / ml, higher than 300 ng / ml, higher than 350 ng / ml, higher than 400 ng / ml, higher than 450 ng / ml, or higher than 500 ng / ml. In some embodiments, the target range is between approximately 55 ng / ml and approximately 1000 ng / ml, between approximately 55 ng / ml and approximately 800 ng / ml, between approximately 55 ng / ml and approximately 600 ng / ml, between approximately 55 ng / ml and approximately 500 ng / ml, between approximately 55 ng / ml and approximately 400 ng / ml, between approximately 55 ng / ml and approximately 300 ng / ml, between approximately 55 ng / ml and approximately 250 ng / ml, between approximately 55 ng / ml and approximately 300 ng / ml, between approximately 55 ng / ml and approximately 200 ng / ml, between approximately 55 ng / ml and approximately 250 ng / ml, between approximately 55 ng / ml and approximately 200 ng / ml, between approximately 55 ng / ml and approximately 150 ng / ml, between approximately 55 ng / ml and approximately 100 ng / ml, and between approximately 55 ng / ml and approximately 80 ng / ml. Between approximately 55 ng / ml and approximately 75 ng / ml, between approximately 100 ng / ml and approximately 1000 ng / ml, between approximately 100 ng / ml and approximately 800 ng / ml, between approximately 100 ng / ml and approximately 600 ng / ml, between approximately 100 ng / ml and approximately 500 ng / ml, between approximately 100 ng / ml and approximately 400 ng / ml, between approximately 100 ng / ml and approximately 300 ng / ml, between approximately 100 ng / ml and approximately 250 ng / ml, between approximately 100 ng / ml and approximately 200 ng / ml, between approximately 100 ng / ml and approximately 250 ng / ml, between approximately 100 ng / ml and approximately 200 ng / ml, between approximately 100 ng / ml and approximately 150 ng / ml, between approximately 100 ng / ml and approximately 125 ng / ml,Between approximately 200 ng / ml and approximately 1000 ng / ml, between approximately 200 ng / ml and approximately 800 ng / ml, between approximately 200 ng / ml and approximately 600 ng / ml, between approximately 200 ng / ml and approximately 500 ng / ml, between approximately 200 ng / ml and approximately 400 ng / ml, between approximately 200 ng / ml and approximately 300 ng / ml, between approximately 200 ng / ml and approximately 250 ng / ml, between approximately 300 ng / ml and approximately 1000 ng / ml, between approximately 300 ng / ml and approximately 800 ng / ml, between approximately 300 ng / ml and approximately 600 ng / ml, between approximately 300 ng / ml and approximately 500 ng / ml, and between approximately 300 ng / ml and approximately 400 ng / ml The serum hepcidin levels are between approximately 300 ng / ml and 350 ng / ml, between approximately 400 ng / ml and 1000 ng / ml, between approximately 400 ng / ml and 800 ng / ml, between approximately 400 ng / ml and 600 ng / ml, between approximately 400 ng / ml and 500 ng / ml, between approximately 400 ng / ml and 450 ng / ml, between approximately 800 ng / ml and 1000 ng / ml, between approximately 800 ng / ml and 900 ng / ml, between approximately 800 ng / ml and 850 ng / ml, between approximately 900 ng / ml and 1000 ng / ml, or between approximately 900 ng / ml and 950 ng / ml. However, naturally, other suitable markers (e.g., serum iron levels, total iron-binding capacity (TIBC), ferritin levels, hemoglobin levels, liver iron content, reticulocyte hemoglobin content, IL-6 levels, creatinine levels, etc.) may be evaluated to determine whether the subject is suitable for the treatment methods described herein.

[0180] In some embodiments, myelofibrosis-associated anemia is characterized by high serum creatinine levels. In some embodiments, the normal range for serum creatinine is approximately 0.84–1.21 mg / dL. In some embodiments, the subject has serum creatinine levels higher than normal. In some embodiments, the subject has a serum creatinine level greater than 1 mg / dL, greater than 1.5 mg / dL, greater than 2 mg / dL, greater than 2.5 mg / dL, greater than 3 mg / dL, greater than 3.5 mg / dL, greater than 4 mg / dL, greater than 4.5 mg / dL, greater than 5 mg / dL, greater than 5.5 mg / dL, greater than 6 mg / dL, greater than 6.5 mg / dL, greater than 7 mg / dL, greater than 7.5 mg / dL, greater than 8 mg / dL, greater than 8.5 mg / dL, greater than 9 mg / dL, greater than 9.5 mg / dL, greater than 10 mg / dL, greater than 15 mg / dL, greater than 20 mg / dL, greater than 30 mg / dL, greater than 40 mg / dL, greater than 50 mg / dL, greater than 60 mg / dL, greater than 70 mg / dL, greater than 80 mg / dL, greater than 90 mg / dL, or greater than 100 mg / dL.In some devices, the target range is between approximately 1 mg / dl and approximately 200 mg / dL, between 1 mg / dl and approximately 175 mg / dL, between 1 mg / dl and approximately 150 mg / dL, between 1 mg / dl and approximately 100 mg / dL, between 1 mg / dl and approximately 50 mg / dL, between 1 mg / dl and approximately 25 mg / dL, between 1 mg / dl and approximately 10 mg / dL, between 1 mg / dl and approximately 5 mg / dL, between 1 mg / dl and approximately 2 mg / dL, and between approximately 5 mg / dl and approximately 200 mg / dL. Between 5 mg / dl and approximately 175 mg / dL, between 5 mg / dl and approximately 150 mg / dL, between 5 mg / dl and approximately 100 mg / dL, between 5 mg / dl and approximately 50 mg / dL, between 5 mg / dl and approximately 25 mg / dL, between 5 mg / dl and approximately 10 mg / dL, between approximately 10 mg / dl and approximately 200 mg / dL, between 10 mg / dl and approximately 175 mg / dL, between 10 mg / dl and approximately 150 mg / dL, between 10 mg / dl and approximately 100 mg / dL, 10 mg Between 10 mg / dl and approximately 50 mg / dL, between 10 mg / dl and approximately 25 mg / dL, between 10 mg / dl and approximately 20 mg / dL, between 10 mg / dl and approximately 25 mg / dL, between approximately 20 mg / dl and approximately 200 mg / dL, between 20 mg / dl and approximately 175 mg / dL, between 20 mg / dl and approximately 150 mg / dL, between 20 mg / dl and approximately 100 mg / dL, between 20 mg / dl and approximately 50 mg / dL, between 20 mg / dl and approximately 25 mg / dL, approximately 50 mg / The serum creatinine levels are between dl and approximately 200 mg / dL, between 50 mg / dl and approximately 175 mg / dL, between 50 mg / dl and approximately 150 mg / dL, between 50 mg / dl and approximately 100 mg / dL, between 50 mg / dl and approximately 75 mg / dL, between approximately 100 mg / dl and approximately 200 mg / dL, between 100 mg / dl and approximately 175 mg / dL, between 100 mg / dl and approximately 150 mg / dL, or between 100 mg / dl and approximately 125 mg / dL. However, naturally, other suitable markers (e.g., serum iron level, total iron-binding capacity (TIBC), ferritin level, hemoglobin level, liver iron content, reticulocyte hemoglobin content, etc.) may be evaluated to determine whether the subject is suitable for the treatment methods described herein.

[0181] In some embodiments, myelofibrosis-associated anemia is also characterized by high serum IL-6 levels. The normal range for IL-6 is ≤ 1.8 pg / ml. In some embodiments, subjects have serum IL-6 levels higher than normal. In some embodiments, subjects have levels higher than 0.5 pg / ml, higher than 0.6 pg / ml, higher than 0.7 pg / ml, higher than 0.8 pg / ml, higher than 0.9 pg / ml, higher than 1 pg / ml, higher than 1.1 pg / ml, higher than 1.2 pg / ml, higher than 1.3 pg / ml, higher than 1.4 pg / ml, higher than 1.5 pg / ml, higher than 1.6 pg / ml, higher than 1.7 pg / ml, higher than 1.8 pg / ml, higher than 2 pg / ml, higher than 3 pg / ml, higher than 4 pg / ml, higher than 5 pg / ml, higher than 6 pg / ml, higher than 7 pg / ml, higher than 8 pg / ml Having a high serum IL-6 level, higher than 9 pg / ml, higher than 10 pg / ml, higher than 20 pg / ml, higher than 30 pg / ml, higher than 40 pg / ml, higher than 50 pg / ml, higher than 60 pg / ml, higher than 70 pg / ml, higher than 80 pg / ml, higher than 90 pg / ml, higher than 100 pg / ml, higher than 200 pg / ml, higher than 300 pg / ml, higher than 400 pg / ml, higher than 500 pg / ml, higher than 600 pg / ml, higher than 700 pg / ml, higher than 800 pg / ml, higher than 900 pg / ml, or higher than 1000 pg / ml. In some embodiments, the target range is between approximately 0.5 pg / ml and approximately 1500 pg / ml, between approximately 0.5 pg / ml and approximately 1000 pg / ml, between approximately 0.5 pg / ml and approximately 800 pg / ml, between approximately 0.5 pg / ml and approximately 750 pg / ml, between approximately 0.5 pg / ml and approximately 500 pg / ml, between approximately 0.5 pg / ml and approximately 250 pg / ml, and between approximately 0.5 pg / ml and approximately 200 pg / ml. Between approximately 0.5 pg / ml and approximately 150 pg / ml, between approximately 0.5 pg / ml and approximately 100 pg / ml, between approximately 0.5 pg / ml and approximately 50 pg / ml, between approximately 0.5 pg / ml and approximately 25 pg / ml, between approximately 0.5 pg / ml and approximately 10 pg / ml, between approximately 0.5 pg / ml and approximately 5 pg / ml, between approximately 0.5 pg / ml and approximately 2.5 pg / ml, between approximately 0.5 pg / ml and approximately 1 pg / ml,Between approximately 1 pg / ml and approximately 1500 pg / ml, between approximately 1 pg / ml and approximately 1000 pg / ml, between approximately 1 pg / ml and approximately 800 pg / ml, between approximately 1 pg / ml and approximately 750 pg / ml, between approximately 1 pg / ml and approximately 500 pg / ml, between approximately 1 pg / ml and approximately 250 pg / ml, between approximately 1 pg / ml and approximately 200 pg / ml, between approximately 1 pg / ml and approximately 150 pg / ml, between approximately 1 pg / ml and approximately 100 pg / ml, between approximately 1 pg / ml and approximately 50 pg / ml, between approximately 1 pg / ml and approximately 25 pg / ml, between approximately 1 pg / ml and approximately 10 pg / ml, and Between approximately 5 pg / ml, between approximately 1 pg / ml and approximately 2.5 pg / ml, between approximately 1 pg / ml and approximately 2 pg / ml, between approximately 1.2 pg / ml and approximately 2 pg / ml, between approximately 1.5 pg / ml and approximately 2 pg / ml, between approximately 1.2 pg / ml and approximately 1.8 pg / ml, between approximately 2 pg / ml and approximately 1500 pg / ml, between approximately 2 pg / ml and approximately 1000 pg / ml, between approximately 2 pg / ml and approximately 800 pg / ml, between approximately 2 pg / ml and approximately 750 pg / ml, between approximately 2 pg / ml and approximately 500 pg / ml, between approximately 2 pg / ml and approximately 250 pg / ml, between approximately 2 pg / ml and approximately 200 pg / Between ml, between approximately 2 pg / ml and approximately 150 pg / ml, between approximately 2 pg / ml and approximately 100 pg / ml, between approximately 2 pg / ml and approximately 50 pg / ml, between approximately 2 pg / ml and approximately 25 pg / ml, between approximately 2 pg / ml and approximately 10 pg / ml, between approximately 2 pg / ml and approximately 5 pg / ml, between approximately 2 pg / ml and approximately 3 pg / ml, between approximately 2 pg / ml and approximately 4 pg / ml, between approximately 2 pg / ml and approximately 2.5 pg / ml, between approximately 5 pg / ml and approximately 1500 pg / ml, between approximately 5 pg / ml and approximately 1000 pg / ml, between approximately 5 pg / ml and approximately 800 pg / ml, between approximately 5 pg / ml and approximately Between 750 pg / ml, between approximately 5 pg / ml and approximately 500 pg / ml, between approximately 5 pg / ml and approximately 250 pg / ml, between approximately 5 pg / ml and approximately 200 pg / ml, between approximately 5 pg / ml and approximately 150 pg / ml, between approximately 5 pg / ml and approximately 100 pg / ml, between approximately 5 pg / ml and approximately 50 pg / ml, between approximately 5 pg / ml and approximately 25 pg / ml, between approximately 5 pg / ml and approximately 10 pg / ml, between approximately 5 pg / ml and approximately 7.5 pg / ml, between approximately 5 pg / ml and approximately 15 pg / ml, between approximately 5 pg / ml and approximately 20 pg / ml, between approximately 10 pg / ml and approximately 1500 pg / ml,Between approximately 10 pg / ml and approximately 1000 pg / ml, between approximately 10 pg / ml and approximately 800 pg / ml, between approximately 10 pg / ml and approximately 750 pg / ml, between approximately 10 pg / ml and approximately 500 pg / ml, between approximately 10 pg / ml and approximately 250 pg / ml, between approximately 10 pg / ml and approximately 200 pg / ml, between approximately 10 pg / ml and approximately 150 pg / ml, between approximately 10 pg / ml and approximately 100 pg / ml, between approximately 10 pg / ml and approximately 50 pg / ml, between approximately 10 pg / ml and approximately 25 pg / ml, between approximately 10 pg / ml and approximately 15 pg / ml, between approximately 20 pg / ml and approximately 1500 pg / ml Between approximately 1, between approximately 20 pg / ml and 1000 pg / ml, between approximately 20 pg / ml and 800 pg / ml, between approximately 20 pg / ml and 750 pg / ml, between approximately 20 pg / ml and 500 pg / ml, between approximately 20 pg / ml and 250 pg / ml, between approximately 20 pg / ml and 200 pg / ml, between approximately 20 pg / ml and 150 pg / ml, between approximately 20 pg / ml and 100 pg / ml, between approximately 20 pg / ml and 50 pg / ml, between approximately 20 pg / ml and 25 pg / ml, between approximately 30 pg / ml and 1500 pg / ml, between approximately 30 pg / ml and 100 Between 0 pg / ml, between approximately 30 pg / ml and approximately 800 pg / ml, between approximately 30 pg / ml and approximately 750 pg / ml, between approximately 30 pg / ml and approximately 500 pg / ml, between approximately 30 pg / ml and approximately 250 pg / ml, between approximately 30 pg / ml and approximately 200 pg / ml, between approximately 30 pg / ml and approximately 150 pg / ml, between approximately 30 pg / ml and approximately 100 pg / ml, between approximately 30 pg / ml and approximately 50 pg / ml, between approximately 30 pg / ml and approximately 45 pg / ml, between approximately 30 pg / ml and approximately 45 pg / ml, between approximately 50 pg / ml and approximately 1500 pg / ml, and approximately 50 pg / ml Between approximately 1000 pg / ml, between approximately 50 pg / ml and approximately 800 pg / ml, between approximately 50 pg / ml and approximately 750 pg / ml, between approximately 50 pg / ml and approximately 500 pg / ml, between approximately 50 pg / ml and approximately 250 pg / ml, between approximately 50 pg / ml and approximately 200 pg / ml, between approximately 50 pg / ml and approximately 150 pg / ml, between approximately 50 pg / ml and approximately 100 pg / ml, between approximately 50 pg / ml and approximately 75 pg / ml, between approximately 100 pg / ml and approximately 1500 pg / ml, between approximately 100 pg / ml and approximately 1000 pg / ml, between approximately 100 pg / ml and approximately 800 pg / ml,The patient has serum IL-6 levels between approximately 100 pg / ml and 750 pg / ml, between approximately 100 pg / ml and 500 pg / ml, between approximately 100 pg / ml and 250 pg / ml, between approximately 100 pg / ml and 200 pg / ml, between approximately 100 pg / ml and 150 pg / ml, between approximately 100 pg / ml and 125 pg / ml, between approximately 500 pg / ml and 1500 pg / ml, between approximately 500 pg / ml and 1000 pg / ml, between approximately 500 pg / ml and 800 pg / ml, between approximately 500 pg / ml and 750 pg / ml, between approximately 1000 pg / ml and 1500 pg / ml, or between approximately 1000 pg / ml and 1250 pg / ml. However, naturally, other suitable markers (e.g., serum iron levels, total iron-binding capacity (TIBC), ferritin levels, hemoglobin levels, liver iron content, reticulocyte hemoglobin content, creatinine levels, etc.) may be evaluated to determine whether the subject is suitable for the treatment methods described herein.

[0182] In some embodiments, subjects requiring treatment according to this disclosure have not received any other treatment for a blood disorder. In some embodiments, subjects requiring treatment are treated for MF-associated anemia with any hepcidin antagonist described herein. In some embodiments, the hepcidin antagonist is a hemoduverin-induced BMP signaling antagonist. In some embodiments, the hemoduverin-induced BMP signaling antagonist is a BMP antagonist. In some embodiments, the BMP antagonist is a BMP2, BMP4, BMP5, or BMP6 antagonist. In some embodiments, the BMP antagonist is a BMP6 antagonist. In some embodiments, the hemoduverin-induced BMP signaling antagonist is a BMP6 neutralizing antibody. In some embodiments, the BMP6 neutralizing antibody is LY311359, CSJ137, or KY1070.

[0183] In some embodiments, the subject requiring treatment in accordance with this disclosure is treated with a modified heparin selected from SST0001, RO-82, RO-68, NAc-91, and NacRO-00.

[0184] In some embodiments, the subject requiring treatment in accordance with this disclosure is treated with a hemoduvelin (HJV) antagonist. In some embodiments, the HJV antagonist is an anti-HJV antibody (e.g., any anti-HJV antibody listed in Table 1 or Table 2). In some embodiments, the HJV antagonist is HJV-35202. In some embodiments, the HJV antagonist is soluble HJV. In some embodiments, soluble HJV is a soluble hemoduvelin-Fc fusion protein. In some embodiments, the soluble HJV-Fc fusion protein is FMX8. In some embodiments, the HJV antagonist is any other HJV antagonist described herein.

[0185] In some embodiments, the subject requiring treatment according to this disclosure is treated with a BMP receptor antagonist. In some embodiments, the BMP receptor antagonist is a BMP type I receptor antagonist. In some embodiments, the BMP type I receptor antagonist is an ALK2 antagonist. In some embodiments, the ALK2 antagonist is KER-047 or BLU-782. In some embodiments, the ALK2 inhibitor selectively inhibits its target molecule (e.g., ALK2) compared to a reference molecule (e.g., JAK1 / 2). In some embodiments, the reference molecule is JAK2. In some embodiments, the ALK2 inhibitor is not momerotinib. In some embodiments, the ALK2 antagonist is any ALK2 antagonist described herein. In some embodiments, the ALK2 inhibitor is not a selective ALK2 inhibitor. In some embodiments, the ALK2 inhibitor also inhibits other target molecules (e.g., JAK1 / 2). In some embodiments, the ALK2 inhibitor is momerotinib. In some embodiments, the BMP receptor antagonist is a BMP type II receptor antagonist. In some embodiments, the BMP type II receptor antagonist is an ActRIIA or ActRIIB antagonist. In some embodiments, the ActRIIA or ActRIIB antagonist is a GDF ligand trap. In some embodiments, the GDF ligand trap is a sotatercept or raspatercept. In some embodiments, the BMP receptor antagonist is any BMP receptor antagonist described herein.

[0186] In some embodiments, subjects requiring treatment in accordance with this disclosure are treated with recombinant SMAD6 or SMAD7. In some embodiments, subjects requiring treatment in accordance with this disclosure are treated with an antagonist that targets SMAD1, SMAD4, SMAD5 and / or SMAD8 (e.g., an intracellular antibody or inhibitory nucleic acid that targets SMAD1, SMAD4, SMAD5 and / or SMAD8).

[0187] In some embodiments, the subject requiring treatment in accordance with this disclosure is treated with a hepcidine neutralizer. In some embodiments, the hepcidine neutralizer is NOX-94, a PEGylated L-stereoisomerized RNA aptamer that binds to and neutralizes hepcidine. In some embodiments, the hepcidine neutralizer is PRS-080, an anticharin against hepcidine. In some embodiments, the hepcidine neutralizer is LY2787106, a monoclonal antibody that targets hepcidine. In some embodiments, the hepcidine neutralizer is any hepcidine neutralizer described herein.

[0188] In some embodiments, subjects requiring treatment in accordance with this disclosure continue to receive therapeutic treatment for blood disorders. Accordingly, this disclosure provides compositions and methods for treating myelofibrosis and / or one or more conditions resulting from myelofibrosis by administration to subjects requiring it in some embodiments. In some embodiments, subjects are administered the hepcidin antagonists described herein in combination with one or more additional therapeutic agents (e.g., JAK-STAT inhibitors, GDF traps, BET inhibitors, or immunomodulators / erythropoietin stimulants). In some embodiments, the hepcidin antagonist used in combination therapy with one or more additional therapeutic agents (e.g., JAK-STAT inhibitors, GDF traps, BET inhibitors, or immunomodulators / erythropoietin stimulants) is an HJV-induced BMP signaling pathway antagonist, e.g., a BMP antagonist (e.g., a BMP6 antagonist or modified heparin), a BMP receptor antagonist (e.g., an ALK2 antagonist), an HJV antagonist (e.g., an anti-HJV antibody or soluble HJV such as a soluble HJV.Fc fusion protein), or a hepcidin neutralizer (e.g., an anti-hepcidin antibody, antikalin targeting hepcidin, or inhibitory nucleic acid targeting hepcidin). In some embodiments, administration of the hepcidin antagonists described herein results in an increase in the level of bioavailable iron for erythrocyte production.

[0189] In some embodiments, subjects are administered an immunomodulator / erythropoietin stimulant (e.g., danazol, prednisone, thalidomide, lenalidomide, pomalidomide) or an HJV-inducing BMP signaling pathway antagonist in combination with EPO [e.g., a BMP antagonist (e.g., a BMP6 antagonist or modified heparin), a BMP receptor antagonist (e.g., an ALK2 antagonist), an HJV antagonist (e.g., an anti-HJV antibody or soluble HJV such as soluble HJV.Fc fusion protein), or a hepcidin neutralizer (e.g., an anti-hepcidin antibody, anticharin targeting hepcidin, or inhibitory nucleic acid targeting hepcidin)]. In some embodiments, an immunomodulator / erythropoietin stimulant (e.g., danazol, prednisone, thalidomide, lenalidomide, pomalidomide) or EPO is administered in combination with an HJV-inducing BMP signaling antagonist. In some embodiments, an immunomodulator / erythropoietin stimulant (e.g., danazol, prednisone, thalidomide, lenalidomide, pomalidomide) or EPO is combined with a BMP antagonist (e.g., a BMP6 antagonist as described herein). In some embodiments, an immunomodulator / erythropoietin stimulant (e.g., danazol, prednisone, thalidomide, lenalidomide, pomalidomide) or EPO is combined with an HJV antagonist (e.g., an HJV antagonist such as an anti-HJV antibody or a soluble HJV.Fc fusion protein). In some embodiments, an immunomodulator / erythropoietin stimulant (e.g., danazol, prednisone, thalidomide, lenalidomide, pomalidomide) or EPO is combined with an anti-HJV antibody as described herein (e.g., any anti-HJV antibody listed in Table 1 or Table 2). In some embodiments, the HJV-Fc fusion protein is FMX8. In some embodiments, an immunomodulator / erythropoietin stimulant (e.g., danazol, prednisone, thalidomide, lenalidomide, pomalidomide) or EPO is combined with a BMP receptor antagonist (e.g., an ALK2 inhibitor or GDF ligand trap such as INCB000928, KER-047, or BLU-782 as described herein).In some embodiments, immunomodulators / erythropoietin stimulants (e.g., danazol, prednisone, thalidomide, lenalidomide, pomalidomide) or EPOs are combined with hepcidin antagonists (e.g., hepcidin antagonists described herein). However, in some embodiments, immunomodulators / erythropoietin stimulants (e.g., danazol, prednisone, thalidomide, lenalidomide, pomalidomide) or EPOs are administered in combination with JAK-STAT antagonists and / or any hepcidin antagonists described herein.

[0190] In some embodiments, subjects are administered a hepcidin antagonist in combination with a JAK / STAT pathway inhibitor [e.g., a BMP antagonist (e.g., a BMP6 antagonist or modified heparin), a BMP receptor antagonist (e.g., an ALK2 antagonist), an HJV antagonist (e.g., a soluble HJV such as an anti-HJV antibody or a soluble HJV.Fc fusion protein), or a hepcidin neutralizer (e.g., an anti-hepcidin antibody, anticharin targeting hepcidin, or an inhibitory nucleic acid targeting hepcidin)] which is an HJV-inducing BMP signaling pathway antagonist. Any hepcidin antagonist described herein may be combined with a JAK-STAT inhibitor. In some embodiments, the JAK-STAT pathway inhibitor is either a JAK inhibitor or a STAT inhibitor. In some embodiments, the JAK inhibitor is selective for one or both of subtypes JAK1 and JAK2 (e.g., ruxolitinib). In some embodiments, the JAK inhibitor is selective for JAK2 (e.g., fedratinib). In some embodiments, the STAT inhibitor is a STAT3 inhibitor. In some embodiments, the JAK inhibitor is not a selective JAK inhibitor. In some embodiments, the JAK inhibitor is an inhibitor of both JAK1 / 2 and ALK2 (e.g., momerotinib). In some embodiments, the JAK1 / 2 or STAT3 inhibitor is selected from the group consisting of ruxolitinib, momerotinib, pacritinib, federatinib, baricitinib, tofacitinib, oclacitinib, INCB039110, NSC13626, AG490, and PpYLKTK. In some embodiments, the JAK1 / 2 or STAT3 inhibitor is an IL6 antagonist or an IL6R antagonist (e.g., an IL6 or IL-6R antibody). In some embodiments, subjects are administered an HJV antagonist (e.g., an anti-HJV antibody as described herein, or a soluble HJV such as a soluble HJV.Fc fusion protein) in combination with a JAK-STAT inhibitor (e.g., ruxolitinib, federatinib, momerotinib, or an IL6 / IL6R antagonist).In some embodiments, subjects are administered a BMP6 antagonist (e.g., an anti-BMP6 antibody) as described herein in combination with a JAK-STAT inhibitor (e.g., ruxolitinib, federatinib, momerotinib, or an IL6 / IL6R antagonist). In some embodiments, subjects are administered an ALK2 antagonist (e.g., an anti-ALK2 antibody or an ALK2 inhibitor such as INCB000928, KER-047, or BLU-782) as described herein in combination with a JAK-STAT inhibitor (e.g., ruxolitinib, federatinib, momerotinib, or an IL6 / IL6R antagonist). In some embodiments, subjects are administered a hepcidin antagonist described herein (e.g., an anti-hepcidin antibody) in combination with a JAK-STAT inhibitor (e.g., ruxolitinib, federatinib, momerotinib, or an IL6 / IL6R antagonist).

[0191] In some embodiments, HJV-inducing BMP signaling pathway antagonists such as hepcidin antagonists (e.g., BMP6 antagonists or modified heparin), BMP receptor antagonists (e.g., ALK2 antagonists), HJV antagonists (e.g., anti-HJV antibodies or soluble HJV such as soluble HJV.Fc fusion proteins), or hepcidin neutralizers (e.g., anti-hepcidin antibodies, anticharin targeting hepcidin, or inhibitory nucleic acids targeting hepcidin) reduce the degree of anemia in subjects exhibiting an anemic response to JAK-STAT pathway inhibitors. For example, in some embodiments, subjects treated with a JAK-STAT pathway inhibitor as monotherapy may be characterized by a deficiency in the ability of the blood to transport oxygen, a deficiency in red blood cells, a deficiency in hemoglobin, and / or a deficiency in total blood volume compared to the subject's pre-treatment state. Therefore, in some embodiments, hepcidin antagonists [e.g., BMP antagonists (e.g., BMP6 antagonists or modified heparin), BMP receptor antagonists (e.g., ALK2 antagonists), HJV antagonists (e.g., anti-HJV antibodies or soluble HJV such as soluble HJV.Fc fusion proteins), or hepcidin neutralizers (e.g., anti-hepcidin antibodies, hepcidin-targeting agents] may be used. HJV-inducing BMP signaling pathway antagonists (such as inhibitory nucleic acids that target antikalin or hepcidin) reduce the degree to which subjects exhibit an anemic response to JAK-STAT pathway inhibitors selected from the group consisting of ruxolitinib, pacritinib, federatinib, baricitinib, tofacitinib, oclacitinib, INCB039110, NSC13626, AG490, and PpYLKTK.In some embodiments, HJV-inducing BMP signaling pathway antagonists such as hepcidin antagonists (e.g., BMP6 antagonists or modified heparin), BMP receptor antagonists (e.g., ALK2 antagonists), HJV antagonists (e.g., anti-HJV antibodies or soluble HJV such as soluble HJV.Fc fusion proteins), or hepcidin neutralizers (e.g., anti-hepcidin antibodies, antikalin that targets hepcidin, or inhibitory nucleic acids that target hepcidin) reduce the degree to which subjects exhibit an anemic response to JAK-STAT inhibitor (e.g., ruxolitinib) administration.

[0192] In some embodiments, subjects are administered an HJV-inducing BMP signaling pathway antagonist in combination with a growth factor ligand trap. This antagonist may include a BMP antagonist (e.g., a BMP6 antagonist or modified heparin), a BMP receptor antagonist (e.g., an ALK2 antagonist), an HJV antagonist (e.g., a soluble HJV such as an anti-HJV antibody or a soluble HJV.Fc fusion protein), or a hepcidin neutralizer (e.g., an anti-hepcidin antibody, anticarin that targets hepcidin, or an inhibitory nucleic acid that targets hepcidin). In some embodiments, the growth factor ligand trap is a transformed growth factor beta (TGF-β) ligand trap. In some embodiments, the TGF-β ligand trap is sotatercept or raspatercept. In some embodiments, subjects are administered a hemoduveline antagonist in combination with an antifibrotic agent. In some embodiments, the antifibrotic agent is PRM-151. In some embodiments, the growth factor ligand trap is administered in combination with an HJV-induced BMP signaling pathway antagonist. In some embodiments, the growth factor ligand trap is combined with a BMP antagonist (e.g., a BMP6 antagonist as described herein). In some embodiments, the growth factor ligand trap is combined with an HJV antagonist (e.g., an HJV antagonist such as an anti-HJV antibody or a soluble HJV.Fc fusion protein). In some embodiments, the growth factor ligand trap is combined with an anti-HJV antibody as described herein (e.g., any anti-HJV antibody listed in Table 1 or Table 2). In some embodiments, the HJV-Fc fusion protein is FMX8. In some embodiments, the growth factor ligand trap is combined with a BMP antagonist (e.g., a BMP6 antagonist as described herein). In some embodiments, the growth factor ligand trap is combined with a BMP receptor antagonist (e.g., an ALK2 inhibitor or GDF ligand trap such as INCB000928, KER-047, or BLU-782 as described herein).In some embodiments, the growth factor ligand trap is combined with a hepcidin neutralizer (e.g., a hepcidin neutralizer as described herein). However, in some embodiments, the growth factor ligand trap is administered in combination with a JAK-STAT antagonist and / or any hepcidin antagonist as described herein.

[0193] In some embodiments, the Disclosure provides a method for treating anemia in subjects with myelofibrosis using a combination of a hepcidin antagonist [e.g., a BMP antagonist (e.g., a BMP6 antagonist or modified heparin), a BMP receptor antagonist (e.g., an ALK2 antagonist), an HJV antagonist (e.g., a soluble HJV such as an anti-HJV antibody or a soluble HJV.Fc fusion protein), or a hepcidin neutralizer (e.g., an anti-hepcidin antibody, anticharin targeting hepcidin, or an inhibitory nucleic acid targeting hepcidin)] and a BET inhibitor (e.g., CPI-0610). In some embodiments, the BET inhibitor (e.g., CPI-0610) is administered in combination with the HJV-induced BMP signaling antagonist. In some embodiments, the BET inhibitor (e.g., CPI-0610) is combined with a BMP antagonist (e.g., a BMP6 antagonist as described herein). In some embodiments, the BET inhibitor (e.g., CPI-0610) is combined with an HJV antagonist (e.g., an anti-HJV antibody, or an HJV antagonist such as a soluble HJV.Fc fusion protein). In some embodiments, the BET inhibitor (e.g., CPI-0610) is combined with an anti-HJV antibody as described herein (e.g., any anti-HJV antibody listed in Table 1 or Table 2).

[0194] In some embodiments, the HJV-Fc fusion protein is FMX8. In some embodiments, the BET inhibitor (e.g., CPI-0610) is combined with a BMP antagonist (e.g., a BMP6 antagonist as described herein). In some embodiments, the BET inhibitor (e.g., CPI-0610) is combined with a BMP receptor antagonist (e.g., an ALK2 inhibitor such as INCB000928, KER-047, or BLU-782 as described herein, or a GDF ligand trap). In some embodiments, the BET inhibitor (e.g., CPI-0610) is combined with a hepcidin antagonist (e.g., a hepcidin antagonist as described herein). However, in some embodiments, the BET inhibitor (e.g., CPI-0610) is administered in combination with a JAK-STAT antagonist and / or any hepcidin antagonist as described herein.

[0195] The therapeutic success of a subject following this disclosure may be determined by methods known in the art or by a skilled physician. In some embodiments, hepcidin antagonist treatment is evaluated based on the subject's serum hepcidin level. For example, in some embodiments, the subject's baseline serum hepcidin level is determined (e.g., before treatment with a hepcidin antagonist or at the time of determination without hepcidin antagonist treatment) and compared to the subject's serum hepcidin level after treatment. In some embodiments, the hepcidin antagonist reduces the subject's serum hepcidin level between approximately 1 ng / mL and approximately 300 ng / mL, and the subject is considered to have undergone therapeutic success. In some embodiments, the hepcidin antagonist reduces the target serum hepcidin level between approximately 1 ng / mL and approximately 200 ng / mL, between approximately 1 ng / mL and approximately 100 ng / mL, between approximately 1 ng / mL and approximately 50 ng / mL, between approximately 1 ng / mL and approximately 10 ng / mL, between approximately 10 ng / mL and approximately 100 ng / mL, or between approximately 10 ng / mL and approximately 50 ng / mL.

[0196] In some embodiments, hepcidin antagonist treatment is evaluated based on the subject's serum ferritin level. For example, in some embodiments, the subject's baseline serum ferritin level is determined (e.g., before or at the time of determination without hepcidin antagonist treatment) and compared to the subject's serum ferritin level after treatment. In some embodiments, the hepcidin antagonist reduces the subject's serum ferritin level between approximately 1 ng / mL and approximately 200 ng / mL, and the subject is considered to have received treatment successfully. In some embodiments, the hepcidin antagonist reduces the subject's serum ferritin level between approximately 1 ng / mL and approximately 100 ng / mL, between approximately 1 ng / mL and approximately 50 ng / mL, between approximately 1 ng / mL and approximately 25 ng / mL, between approximately 1 ng / mL and approximately 10 ng / mL, between approximately 10 ng / mL and approximately 100 ng / mL, or between approximately 10 ng / mL and approximately 50 ng / mL.

[0197] In some embodiments, hepcidin antagonist treatment is evaluated based on the subject's serum hemoglobin level. For example, in some embodiments, the subject's baseline serum hemoglobin level is determined (e.g., before or at the time of determination without hepcidin antagonist treatment) and compared to the subject's serum hemoglobin level after treatment. In some embodiments, the hepcidin antagonist increases the subject's serum hemoglobin level between approximately 0.01 g / dL and approximately 5 g / dL, and the subject is considered to have received successful treatment. In some embodiments, the hepcidin antagonist reduces the target serum ferritin level between approximately 0.01 g / dL and approximately 1 g / dL, between approximately 0.1 g / dL and approximately 5 g / dL, between approximately 1 g / dL and approximately 5 g / dL, between approximately 0.01 g / dL and approximately 0.1 g / dL, between approximately 0.5 g / dL and approximately 2.5 g / dL, or between approximately 0.1 g / dL and approximately 1 g / dL.

[0198] The determination of whether the amount of hepcidin antagonist achieved a therapeutic effect is obvious to those skilled in the art based on the teachings provided herein. The effective dose will vary, as recognized by those skilled in the art, depending on the specific condition being treated, the severity of the condition, individual patient parameters including age, health status, size, sex, and weight, the duration of treatment, the nature of any concomitant therapy, the specific route of administration, and factors within the knowledge and expertise of the healthcare professional. These factors are well known to those skilled in the art and can be addressed simply by routine experimentation. The specific administration regimens used in the methods described herein, i.e., dose, timing, and frequency, will depend on the specific subject and their medical history, as described herein.

[0199] Empirical considerations, such as the time to maximum effect, half-life, and / or time to exceed a certain concentration, will usually contribute to determining the dosage.

[0200] In some embodiments, the dose of the hepcidin antagonist described herein may be empirically determined in an individual given one or more doses of the antibody. The individual is given an increasing dose of the antagonist. Disease / impairment indicators may be followed to evaluate the efficacy of the antagonist.

[0201] The frequency of administration may vary according to the claimed method. In some embodiments, the composition will be administered once. In some embodiments, the treatment will be administered on multiple occasions. In some embodiments, the frequency of administration may be weekly, every two weeks, every three weeks, every four weeks, every five weeks, every six weeks, every seven weeks, every eight weeks, every nine weeks, or every ten weeks; or once a month, every two months, or every three months, or more. In some embodiments, the composition may be administered daily, twice a week, once a week, twice a month, once a month, or at any time interval that provides a suitable (e.g., maximum) efficacy but minimizes the risk to the subject's safety. In general, efficacy and the risks of treatment and safety may be monitored throughout the course of treatment.

[0202] In some embodiments, administration of a hepcidin antagonist results in a decrease in serum hepcidin-25 concentration and / or an increase in serum TSAT%, and in some embodiments, these effects persist for a period of time (e.g., one month or longer). Therefore, in some embodiments, the timing and frequency of hepcidin antagonist administration may be determined by monitoring one or more biomarkers, e.g., criteria for evaluating iron utilization or flaggable iron overload. For example, in some embodiments, the hepcidin antagonist is administered intermittently or according to the level of a specific biomarker, such as serum hepcidin-25 levels or transferrin saturation percentage (TSAT%). In some embodiments, the biomarker levels described herein may be used to determine whether a subject is a candidate for treatment. However, in some embodiments, the biomarker may be used, for example, by the hepcidin antagonist to determine whether to continue, restart, or discontinue treatment.

[0203] For example, in some embodiments, if the subject's TSAT% is 70% or higher, 75% or higher, 80% or higher, 85% or higher, 90% or higher, or 95% or higher, the subject may not be considered a candidate for treatment. In some cases, if the subject's TSAT% is 70% or higher, 75% or higher, 80% or higher, 85% or higher, 90% or higher, or 95% or higher, ongoing treatment with a hepcidin antagonist may be stopped or temporarily stopped, for example, to prevent iron overload. In other embodiments, administration of an anti-HJV antibody may be carried out if the subject's TSAT% is 95% or lower, 90% or lower, 80% or lower, 70% or lower, 65% or lower, 60% or lower, 55% or lower, 50% or lower, 45% or lower, 40% or lower, 35% or lower, or 30% or lower. Therefore, in some embodiments, the target TSAT% may be monitored, for example, continuously or periodically, while the patient may, for example, receive or be treated for anemia and be evaluated to prevent iron overload or to determine if further treatment is appropriate. However, naturally, other suitable markers (including, for example, ferritin levels, serum iron levels, creatinine levels, etc.) may be monitored to determine the dosage and frequency of administration according to the methods provided herein.

[0204] In some embodiments, a subject may be administered a composition provided herein (e.g., a hepcidin antagonist) at intervals of one or more for a certain period of time. In some cases, the period during which the subject is administered the composition at intervals of one or more may be divided by the period during which the subject is not administered the composition. In some embodiments, the relative duration of each period may depend on the subject's response to the treatment or the severity of the disease or both, and / or be determined based on the judgment of the treating physician. For example, in some embodiments, over the course of a year, the subject may be administered the composition weekly, bi-weekly, or monthly for two months, followed by a 10-month hiatus. In some embodiments, over the course of a year, the subject may be administered the composition weekly, bi-weekly, or monthly for three months, followed by a 9-month hiatus. In some embodiments, over the course of a year, the subject may be administered the composition weekly, bi-weekly, or monthly for four months, followed by an 8-month hiatus. In some embodiments, over the course of a year, the subject may be administered the composition weekly, bi-weekly, or monthly for five months, followed by a 7-month hiatus. In some embodiments, over the course of a year, the subject may be administered the composition weekly, bi-weekly, or monthly for 6 months, followed by a 6-month suspension of administration. In some embodiments, over the course of a year, the subject may be administered the composition weekly, bi-weekly, or monthly for 7 months, followed by a 5-month suspension of administration. In some embodiments, over the course of a year, the subject may be administered the composition weekly, bi-weekly, or monthly for 8 months, followed by a 4-month suspension of administration. In some embodiments, over the course of a year, the subject may be administered the composition weekly, bi-weekly, or monthly for 9 months, followed by a 3-month suspension of administration. In some embodiments, over the course of a year, the subject may be administered the composition weekly, bi-weekly, or monthly for 10 months, followed by a 2-month suspension of administration. In some embodiments, over the course of a year, the subject may be administered the composition weekly, bi-weekly, or monthly for 2 months, followed by 2 months of no administration; or administered for 3 months, followed by 3 months of no administration; or administered for 4 months, followed by 4 months of no administration.

[0205] In some embodiments, hepcidin antagonists may be administered parenterally. For example, parenterally administered compositions may be administered subcutaneously, intradermally, intravenously, intraperitoneally, intratumorally, intramuscularly, intraarterally, intraarterially, or by infusion techniques. Furthermore, it may be administered to a subject via an injectable accumulation route of administration, such as using injectable or biodegradable substances and methods with a 1, 3, or 6 month accumulation period. In some embodiments, hepcidin antagonists are administered subcutaneously. In some embodiments, hepcidin antagonists are administered intravenously. [Examples]

[0206] [Example 1] Treatment of myelofibrosis-associated anemia with hepcidin antagonists Iron-restricted erythropoiesis occurs in both absolute and functional iron deficiency (FID). FID represents a state of iron-restricted erythropoiesis characterized by an imbalance between the iron required for effective erythropoiesis and readily available serum iron. In FID, iron is sequestrated and unavailable for erythropoiesis even when the body has sufficient or increased systemic iron stores (Figure 3A). FID has been shown to be caused by elevated hepcidin levels compared to iron storage levels. Elevated hepcidin is observed in diseases associated with FID, such as inflammation (e.g., myelofibrosis, chronic kidney disease in hemodialysis (CKD-HD), autoimmune diseases), iron overload (e.g., myelofibrosis, CKD), genetic disorders (e.g., iron-refractory iron deficiency anemia (IRIDA)), uremic toxins (e.g., CKD), decreased clearance (e.g., chronic kidney disease in peritoneal dialysis (CKD-PD)), and cancer.

[0207] FID is a common feature of inflammatory and chronic disease anemia (AI / ACD), regardless of the etiology of the disease. The contribution of FID to anemia varies among patients with the same disease (AI / ACD has different etiologies) (Figure 3B). A common feature across different etiologies of FID is increased hepcidin levels, which are sufficient to increase iron storage (Figure 3C).

[0208] This disclosure suggests that, at least in part, hepcidin antagonists may be used to reduce hepcidin levels and restore normal erythrocyte production in patients with FID (e.g., patients with myelofibrosis).

[0209] The HAMP gene encodes a hepcidin precursor protein, which is primarily expressed by hepatocytes in the liver and present at low levels in other cells of extrahepatic tissues. The precursor protein is subsequently cleaved to produce bioactive hepcidin. Transcriptional regulators of the HAMP gene include BMP signaling and JAK-STAT3 signaling. Hemomoduvelin (HJV) is a crucial co-receptor in the induction of hepcidin expression via BMP signaling. Inhibition of the HJV-induced BMP signaling pathway and / or the JAK-STAT3 signaling pathway by targeting any component of these pathways can result in reduced hepcidin expression. To reduce hepcidin levels by targeting the HJV-induced BMP signaling pathway, BMP antagonists (e.g., BMP6 antagonists), HJV antagonists (e.g., anti-HJV antibodies, HJV-Fc), BMP receptor antagonists, SMAD1 / 5 / 8 antagonists, or hepcidin neutralizers may be used (Figure 3G).

[0210] For example, anti-HJV antibodies have been shown to reduce hepcidin synthesis and decrease the severity of anemia (Kovacs et al., Anti-hemojuvelin antibody corrects anemia caused by inappropriately high hepcidin levels, Haematologica. 2016 May; 101(5): e173-e176). Figure 3E. Furthermore, HJV is regulated by matryptase-2. Matryptase-2, encoded by the TMPRSS6 gene, is a member of the type II transmembrane serine protease family. Matryptase-2 has been proven essential for iron homeostasis. TMPRSS6 is primarily expressed in the liver and negatively regulates hepcidin production by cleaving membrane-bound hemomodeverin (e.g., Du X., et al. (2008). The serine protease TMPRSS6 is required to sense iron deficiency. Science 320 1088-1092) (Figure 3F). Therefore, increasing matriptase-2 expression in hepatocytes may be another way to negatively regulate hepcidin expression.

[0211] Furthermore, activin B has been shown to stimulate SMAD1 / 5 / 8 signaling and hepcidin expression in hepatocytes to a level comparable to standard SMAD2 / 3 signaling, and with similar or moderately reduced potency compared to BMP6. Activin B stimulates hepcidin via classical activin type II receptors ACVR2A and ACVR2B, non-standard BMP type I receptors, activin receptor-like kinases 2 and 3, and SMAD5. The coreceptor hemoduberin binds to actin B and promotes activin B-SMAD1 / 5 / 8 signaling. (Canali et al., Activin B Induces Noncanonical SMAD1 / 5 / 8 Signaling via BMP Type I Receptors in Hepatocytes: Evidence for a Role in Hepcidin Induction by Inflammation in Male Mice, Endocrinology. 2016 Mar; 157(3): 1146-1162). Figure 4 shows that activin B mediates hepcidin regulation in hepatocytes.

[0212] Myelofibrosis (MF) is a myeloproliferative disorder characterized by abnormal proliferation of hematopoietic stem cells that lead to fibrosis of the bone marrow. The production of healthy blood cells (megakaryocytes, which are responsible for platelet production, and erythrocytes) is impaired. Several genes are associated with the pathogenesis, and most patients have JAK2 mutations (Kralovics R, 2005), followed by CALR and MPL, which result in constitutively active JAK / STAT signaling and dysfunctional hematopoiesis. In some embodiments, molecular loci associated with myelofibrosis include JAK2, CALR, MPL, ASXL1, SRSF2, IDH1 / 2, TET2, EXH2, U2AF1, and CBL.

[0213] MF is one of three Philadelphia chromosome-negative myeloproliferative neoplasms (MPNs) and belongs to a class that also includes essential thrombocythemia (ET) and polycythemia vera (PV). MF can be classified into primary MF (PMF) and secondary MF (SMF). PMF and SMF have similar clinical profiles, including common main symptoms such as anemia, fatigue, and splenomegaly.

[0214] PMF is the most common result of a driver mutation within a single hematopoietic stem cell. Approximately 95% of PMF patients have a mutation in one of three genes: JAK2 (63%), CALR (25%), and MPL (7%) (Klampf T, 2013; Nangalia J, 2013; Cazzola M, 2014; Tefferi A, 2014c). Some of these mutations are mutually exclusive (Cazzola M, 2014; Tapper W, 2015). In fewer than 10% of patients, the disease is not driven by a (known) mutation (Tefferi A, 2014c; Tefferi A, 2016). Somatic JAK2V617F is a gain-of-function mutation, and only JAK2 mutations are associated with MF. In a study of 244 MPN patients, 57% of patients with PMF had the JAK2V617F mutation (Kralovics R, 2005). Another common mutation was identified in the MPL gene of MF patients (MPL W515L / K mutation; Guglielmelli P, 2007). Unlike JAK2 and MPL, CALR has significantly more mutagenic variations; approximately 140 CALR mutations were identified in 19 variants, with exon 9 mutations being the most frequently found in MF patients (Nangalia J, 2013). Further loci are associated with PMF, including TET2, ASXL1, SRSF2, IDH1 / 2, U2AF1, and CBL. More than 80% of MF patients have at least one of these further mutations (Tefferi A, 2016). The presence of one of these mutations may negatively impact disease progression and prognosis (Lasho TL, 2012; Vannucchi AM, 2013).

[0215] SMF shares many similarities with PMF in its pathogenesis, and common genetic mechanisms, namely JAK2, CALR, and MPL driver mutations, are found in both PV and ET. Similar mutation rates are found in ET patients (JAK2 [58%], CALR [23%], and MPL [4%]) versus PMF patients (Elala Y, 2015).

[0216] In patients with myelofibrosis, approximately 32.2% of all MF patients or 33.3% of newly diagnosed patients exhibit functional iron deficiency (FID). Furthermore, elevated hepcidin levels are associated with decreased hemoglobin and iron overload (Pardanani et al., Associations and Prognostic Interactions Between Circulating Levels of Hepcidin, Ferritin and Inflammatory Cytokines in Primary Myelofibrosis, Am J Hematol. 2013 Apr;88(4):312-6). Such patients were predicted to have a lower survival rate. FID was associated with poor QuL scores in myelofibrosis, and IL-6 levels were elevated in anemic MF patients (Birgegard et al., Inflammatory Functional Iron Deficiency Common in Myelofibrosis, Contributes to Anemia and Impairs Quality of Life. From the Nordic MPN Study Group, Eur J Haematol. 2019 Mar;102(3):235-240). In myelofibrosis, pro-inflammatory cytokines that induce hepcidin synthesis, such as IL-6 and oncostatin-M, are typically increased and associated with iron blockade, macrophage iron loading, and myeloproliferation and macrophage activation (Figures 1 and 2).

[0217] Current standard treatment for myelofibrosis is tailored to individual patients based on their risk category (Figure 5A). The approved JAK1 / JAK2 inhibitor ruxolitinib (Jakafi) and the JAK2 inhibitor fedratinib (FED) improve splenomegaly and symptoms but worsen anemia. The experimental drug momerotinib offers a prospect for treating MF with JAK inhibitors without causing severe anemia. Momerotinib also inhibits JAK1, JAK2, and ACRV1 (HJV signaling partner). It can improve disease symptoms and alleviate anemia. Combination therapy with an HJV-induced BMP signaling antagonist and a JAK / STAT inhibitor is effective in reducing MF symptoms and simultaneously mitigating FID-induced anemia. [Example 2]

[0218] In non-human primates, anti-HJV antibodies reduce IL-6-induced hepcidin expression. As shown in Figure 1, in myelofibrosis, pro-inflammatory cytokines that induce hepcidin synthesis, such as IL-6 and oncostatin-M, are typically increased and associated with iron blockade, macrophage iron loading, and myeloproliferation and macrophage activation. To test whether IL-6 indeed increases hepcidin expression and whether anti-HJV antibodies can inhibit IL-6-induced hepcidin expression in non-human primates, sinus was loaded with IL-6 on day 1 and divided into three groups. On day 4, sinus in group 1 received a medium control, sinus in group 2 received 0.6 mg / kg of anti-HJV antibodies (CDR-H1: SEQ ID NO: 1, CDR-H2: SEQ ID NO: 2, CDR-H3: SEQ ID NO: 3, CDR-L1: SEQ ID NO: 7, CDR-L2: SEQ ID NO: 8, and CDR-L3: SEQ ID NO: 9), and sinus in group 3 received the same anti-HJV antibodies at 6.0 mg / kg. On day 11, sinuses in all three groups were again loaded with IL-6, and plasma hepcidin-25 was measured in all sinuses. As shown in Figure 6, IL-6 loading increased plasma hepcidin-25 concentrations on day 1 compared to pre-loading baseline (BL) in sinuses in all three groups. After the second IL-6 loading on day 11, sinuses in group 1 showed an increase in plasma hepcidin-25 similar to that observed on day 1. However, in sinuses in group 2 (0.6 mg / kg anti-HJV antibody) and group 3 (6 mg / kg anti-HJV antibody), the presence of anti-HJV antibody prevented the IL-6-induced increase in plasma hepcidin-25 on day 11 in a dose-dependent manner. In other words, anti-HJV antibody was effective in preventing the dose-dependent increase in inflammation-inducible (IL-6) hepcidin in sinuses. These results indicate that anti-HJV antibodies can inhibit hepcidin expression induced by the IL-6 signaling pathway.

[0219] Equivalents and range In the claims, articles such as “a,” “an,” and “the” may mean one and more unless otherwise indicated or evident from the context. Claims or descriptions containing “or” between one or more members of a group are considered satisfied if one, more than one, or all members of the group are present, used, or related to a given product or method, unless otherwise indicated or evident from the context. The present invention includes embodiments in which exactly one member of the group is present, used, or related to a given product or method. The present invention includes embodiments in which one or more, or all members of the group are present, used, or related to a given product or method.

[0220] Furthermore, the present invention encompasses all variations, combinations, and reorders in which one or more limitations, elements, items, and descriptive terms from one or more enumerated claims are introduced into another claim. For example, any claim that depends on another claim may be modified to include one or more limitations found in any other claim that depends on the same base claim. Where elements are presented as a list, for example in Markush form, each subgroup of the elements is also disclosed, and any element(s) may be excluded from the group. In general, where the present invention or an aspect of the present invention is said to include certain elements and / or features, it should be understood that certain embodiments of the present invention or aspects of the present invention consist of, or are based on, such elements and / or features. For simplicity, these embodiments are not specifically described herein in these terms.

[0221] The phrase "and / or," as used herein and in the claims, should be understood to mean "either or both" of the elements being combined, i.e., elements that exist as a combination in some cases and as uncombined in others. Multiple elements listed using "and / or" should be interpreted in the same way, i.e., "one or more" of the elements being combined. Other elements represent elements other than those specifically identified by the "and / or" clause, and may or may not be related to those specifically identified elements. Thus, as a non-restrictive example, a reference to "A and / or B," when used in conjunction with an unrestrictive word such as "comprising," may refer, in one embodiment, to A only (which may include elements other than B); in another embodiment, to B only (which may include elements other than A); in yet another embodiment, to both A and B (which may include other elements); and so on.

[0222] In this specification and in the claims, “or” as used herein should be understood to have the same meaning as “and / or” as described above. For example, when dividing items in a list, “or” or “and / or” should be interpreted as inclusive, that is, including at least one but more than one of the elements, many or a list, and may include further unlisted items. Only terms that explicitly indicate the opposite, such as “only one of” or “exactly one of,” or, as used in the claims, “consisting of,” would refer to the inclusion of exactly one element of many elements or elements of a list. In general, the term “or” as used herein should be interpreted as indicating an exclusive substitution (i.e., “one or the other, but not both”) when preceded by terms of exclusivity such as “either,” “one of,” “only one of,” or “exactly one of.” When used in patent claims, "consisting essentially of" shall have its ordinary meaning as used in the field of patent law.

[0223] In this specification and in the claims, as used herein, the phrase “at least one” should be understood to mean at least one element selected from any one or more elements of the list of elements, with reference to one or more elements of a list of elements, but not necessarily including at least one of each and all elements specifically enumerated in the list of elements, and not excluding any combination of elements of the list of elements. This definition also makes it possible that an element may represent an element other than those specifically identified in the list of elements to which the phrase “at least one” refers, whether related to or unrelated to those specifically identified elements. Therefore, as a non-restrictive example, “at least one of A and B” (or in other words, “at least one of A or B” or in other words, “at least one of A and / or B”) could mean, in one embodiment, at least one A (and other elements) which may include more than one, in which B is absent; in another embodiment, at least one B (and other elements) which may include more than one, in which A is absent; in yet another embodiment, at least one A which may include more than one, and at least one B (and other elements) which may include more than one; and so on.

[0224] Conversely, unless explicitly stated otherwise, it should be understood that in any method claimed herein that includes more than one step or action, the order of the steps or actions of the method is not necessarily limited to the order in which the steps or actions of the method are enumerated.

[0225] In the claims and in the specification above, all transitional clauses such as “comprising,” “including,” “carrying,” “having,” “containing,” “involving,” “holding,” and “composed of” are understood to be unrestrictive, meaning that they include but are not limited. Only the transitional clauses “consisting of” and “consisting essentially of” are considered restricted or partially restricted transitional clauses, as described in section 2111.03 of the U.S. Patent and Trademark Examination Manual. Embodiments described in this document using an unrestrictive transitional clause (e.g., “comprising”) should be understood to be, in alternative embodiments, equivalent to the features “consisting of” and “consisting essentially of” described by the unrestrictive transitional clauses. For example, if this application describes a "composition comprising A and B," this application also considers alternative embodiments such as a "composition consisting of A and B" and a "basic composition consisting of A and B."

[0226] Where a range is given, it includes the endpoint. Furthermore, unless otherwise indicated or evident from the context and the understanding of those skilled in the art, values ​​expressed as a range may be assumed to be any specific value or subrange within the range described in different embodiments of the invention, up to one-tenth of the lower limit of the range, unless the context explicitly indicates otherwise.

[0227] This application references various published patents, published patent applications, publications, and other papers, all of which are incorporated herein by reference. In the event of any conflict between any incorporated reference and this specification, this specification shall prevail. Furthermore, any particular embodiment of the Invention that exists in the prior art may be expressly excluded from any one or more claims. Such embodiments may be excluded even if the exclusion is not expressly stated herein, as they would be considered publicly known to those skilled in the art. Any particular embodiment of the Invention may be excluded from any claim for any reason, whether or not it is related to the existence of the prior art.

[0228] Those skilled in the art will be able to identify or verify many equivalents of the specific embodiments described herein by means of ordinary experiments. The scope of the embodiments described herein is not intended to be limited to the abov...

Claims

1. A method for treating anemia in a subject with myelofibrosis, Administer an effective dose of hepcidin antagonist to the subject. Methods that include...

2. The method according to claim 1, wherein the subject has iron utilization disorder / functional iron deficiency.

3. The method according to claim 1 or 2, wherein the hepcidin antagonist is a hemoduvelin-induced BMP signaling antagonist.

4. The method according to claim 3, wherein the hemoduvelin-induced BMP signaling antagonist is a BMP antagonist.

5. The method according to claim 4, wherein the BMP antagonist is a BMP2, BMP4, BMP5, or BMP6 antagonist.

6. The method according to claim 5, wherein the BMP antagonist is a BMP6 antagonist.

7. The method according to any one of claims 3 to 6, wherein a hemoduvelin-induced BMP signaling antagonist selectively inhibits its target molecule.

8. The method according to claim 7, wherein the target molecule is a BMP receptor.

9. The method according to claim 7 or 8, wherein the hemoduvelin-induced BMP signaling antagonist selectively inhibits its target molecule compared to a reference molecule.

10. The method according to claim 9, wherein the reference molecule is JAK2.

11. Hemoduvelin-induced BMP signaling antagonists selectively inhibit their target molecule compared to a reference molecule, resulting in a higher half-percentage inhibitory concentration (IC) of the target molecule as measured by kinase efficacy assays. ５０ ) is at least 10 times higher (for example, 10 １ ~10 ６ IC of a reference molecule (in a range twice as high) ５０ The method according to claim 10, comprising:

12. The method according to any one of claims 3 to 10, wherein the hemoduvelin-induced BMP signaling antagonist is sHJV or a soluble hemoduvelin-Fc fusion protein.

13. The method according to claim 12, wherein the soluble HJV-Fc fusion protein is FMX8.

14. The method according to any one of claims 4 to 11, wherein the hemoduvelin-induced BMP signaling antagonist is a BMP6 neutralizing antibody.

15. The method according to claim 14, wherein the BMP6 neutralizing antibody is LY311359, CSJ137, or KY1070.

16. The method according to claim 3 or 4, wherein the hemoduvelin-induced BMP signaling antagonist is a modified heparin selected from SST0001, RO-82, RO-68, NAc-91, and NAcRO-00.

17. The method according to claim 3 or 4, wherein the hemoduvelin-induced BMP signaling antagonist is recombinant SMAD6 or SMAD7.

18. The method according to claim 1 or 2, wherein the hepcidin antagonist is a hepcidin antagonist.

19. The method according to claim 18, wherein the hepcidin neutralizing agent is NOX-94, which is a PEG-modified L-stereoisomer RNA aptamer that binds to and neutralizes hepcidin.

20. The method according to claim 18, wherein the hepcidin neutralizer is PRS-080, which is anticharin for hepcidin.

21. The method according to claim 18, wherein the hepcidin neutralizing agent is LY2787106, a monoclonal antibody that targets hepcidin.

22. The method according to any one of claims 1 to 11, wherein the hemoduvelin-induced BMP signaling antagonist is an ALK2 antagonist.

23. The method according to claim 22, wherein the ALK2 antagonist is INCB000928, KER-047, or BLU-782.

24. The method according to any one of claims 1 to 4, wherein the hepcidin antagonist is a hemoduverin antagonist.

25. The method according to claim 24, wherein the hemoduvelin antagonist is an anti-hemoduvelin antibody.

26. The method according to claim 25, wherein the anti-hemoduvelin antibody preferentially binds to RGMc over RGMa and RGMb.

27. Antihemoduvelin antibodies have an equilibrium dissociation constant (K) lower than 100 nM. Ｄ The method according to claim 26, wherein the RGMc is bound to the RGMc.

28. The method according to any one of claims 25 to 27, wherein the anti-HJV antibody is HJV-35202.

29. The method according to any one of claims 25 to 27, wherein the anti-HJV antibody is the anti-HJV antibody shown in Table 1.

30. Anti-hemoduvelin antibodies: (a) Variable heavy chain regions including CDR1 containing the amino acid sequence of SEQ ID NO: 1, CDR2 containing the amino acid sequence of SEQ ID NO: 2, and CDR3 containing the amino acid sequence of SEQ ID NO: 3; and / or (b) Variable light chain region including CDR1 containing the amino acid sequence of SEQ ID NO: 4, CDR2 containing the amino acid sequence of SEQ ID NO: 5, and CDR3 containing the amino acid sequence of SEQ ID NO: 6 The method according to claim 29, including the method described in claim 29.

31. Anti-hemoduvelin antibodies: (a) Variable heavy chain regions including CDR1 containing the amino acid sequence of SEQ ID NO: 1, CDR2 containing the amino acid sequence of SEQ ID NO: 2, and CDR3 containing the amino acid sequence of SEQ ID NO: 3; and / or (b) Variable light chain region including CDR1 containing the amino acid sequence of SEQ ID NO: 7, CDR2 containing the amino acid sequence of SEQ ID NO: 8, and CDR3 containing the amino acid sequence of SEQ ID NO: 9 The method according to claim 29, including the method described in claim 29.

32. Anti-hemoduvelin antibodies: (a) Variable heavy chain regions including CDR1 containing the amino acid sequence of SEQ ID NO: 1, CDR2 containing the amino acid sequence of SEQ ID NO: 2, and CDR3 containing the amino acid sequence of SEQ ID NO: 3; and / or (b) Variable light chain region including CDR1 containing the amino acid sequence of SEQ ID NO: 10, CDR2 containing the amino acid sequence of SEQ ID NO: 11, and CDR3 containing the amino acid sequence of SEQ ID NO: 12 The method according to claim 29, including the method described in claim 29.

33. Anti-hemoduvelin antibodies: (a) Variable heavy chain regions including CDR1 containing the amino acid sequence of SEQ ID NO: 1, CDR2 containing the amino acid sequence of SEQ ID NO: 2, and CDR3 containing the amino acid sequence of SEQ ID NO: 3; and / or (b) Variable light chain region including CDR1 containing the amino acid sequence of SEQ ID NO: 13, CDR2 containing the amino acid sequence of SEQ ID NO: 14, and CDR3 containing the amino acid sequence of SEQ ID NO: 15 The method according to claim 29, including the method described in claim 29.

34. Anti-hemoduvelin antibodies: (a) Variable heavy chain regions including CDR1 containing the amino acid sequence of SEQ ID NO: 19, CDR2 containing the amino acid sequence of SEQ ID NO: 20, and CDR3 containing the amino acid sequence of SEQ ID NO: 21; and (b) Variable light chain region including CDR1 containing the amino acid sequence of SEQ ID NO: 22, CDR2 containing the amino acid sequence of SEQ ID NO: 23, and CDR3 containing the amino acid sequence of SEQ ID NO: 24 The method according to claim 29, including the method described in claim 29.

35. The method according to any one of claims 1 to 34, wherein the subject has a myelofibrosis-inducing mutation in JAK2, LNK, PPM1D, MPL, ASXL1, TET2, NFE2, SH2B3, SF3B1, or CALR.

36. The method according to any one of claims 1 to 35, wherein the subject has a mutation in a gene involved in epigenetic regulation or splicing, namely ASXL1, DNMT3A, TET2, SRSF2, U2AF1, EZH2, or SF3B1.

37. The method according to any one of claims 1 to 36, wherein the subject has a mutation in IDH1 / 2 associated with the risk of progression to MBN-BP.

38. The method according to any one of claims 35 to 37, wherein the subject contains a human JAK2 gene having an inducement mutation in exon 12 or exon 14.

39. The method according to claim 38, wherein the triggering mutation in the JAK2 gene is located in exon 14 and causes a V617F substitution.

40. The method according to any one of claims 1 to 39, wherein myelofibrosis is associated with increased levels of pro-inflammatory cytokines (e.g., IL-6, oncostatin-M) in the subject.

41. The method according to any one of claims 1 to 40, wherein the subject has or is at risk of having systemic symptoms or microvascular symptoms associated with MPN.

42. The method according to claim 41, wherein the subject has or is at risk of having thromboembolic complications or hemorrhagic complications.

43. The method according to any one of claims 1 to 42, wherein the subject has or is at risk of having MPN-acute transformation phase acute myeloid leukemia (AML).

44. The method according to any one of claims 1 to 43, wherein the subject exhibits ribosomal disease in megakaryocytes.

45. The method according to claim 44, wherein the subject exhibits a decrease in GATA1 expression, particularly in megakaryocytes.

46. The method according to claim 44 or 45, wherein the subject exhibits a defect in megakaryocyte function or maturation.

47. The method according to any one of claims 1 to 46, wherein the subject does not have nutritional iron deficiency.

48. The method according to any one of claims 1 to 47, wherein the subject has ferritin at a level exceeding 100 μg / L.

49. The method according to any one of claims 1 to 48, wherein the subject has a reticulocyte hemoglobin content less than 26 pg / cell.

50. The method according to any one of claims 1 to 49, wherein the subject has a transferrin saturation level lower than 50%.

51. The method according to any one of claims 1 to 50, wherein the subject has a liver iron level higher than 2000 μg / g dry weight.

52. The method according to any one of claims 1 to 51, wherein the subject has a serum iron level in the range of less than 50 μg / dL.

53. The method according to any one of claims 1 to 52, wherein the target has a total iron-binding capacity in the range of 400 μg / dL or less.

54. The method according to any one of claims 1 to 53, wherein the subject has a hepcidin level in the range higher than 55 ng / ml.

55. The method according to any one of claims 1 to 54, wherein the subject has an IL-6 level higher than 1.8 pg / mL.

56. The method according to any one of claims 1 to 55, wherein the subject has a serum creatinine level higher than 2 mg / dL.

57. The method according to any one of claims 1 to 56, wherein the subject is identified as having a hemoglobin level in the range of 1.5 to 2.0 g / dL or 2.0 to 4.0 g / dL or a hemoglobin level lower than normal.

58. The method according to claim 57, wherein the subject exhibits a serum hemoglobin level lower than 10 g / dL.

59. The method according to claim 58, wherein the subject exhibits a serum hemoglobin level lower than 8 g / dL.

60. The method according to any one of claims 1 to 59, wherein administration of a hepcidin antagonist increases the hemoglobin level by at least 1 g / dL from baseline.

61. The method according to any one of claims 1 to 60, wherein the subject is thrombocytopenia, anemia, and / or neutropenia.

62. The method according to any one of claims 1 to 61, wherein the subject has received one or more blood transfusions.

63. The method according to any one of claims 1 to 62, wherein the subject has transfusion-dependent anemia.

64. The method according to claim 63, wherein the subject has received multiple blood transfusions over a period of 12 weeks.

65. The method according to any one of claims 1 to 64, wherein the subject has previously received one or more doses of a JAK / STAT antagonist as treatment for Philadelphia chromosome-negative myeloproliferative neoplasm (MPN).

66. The method according to claim 65, wherein the subject has received a JAK / STAT antagonist as treatment for polycythemia vera (PV), essential thrombocythemia (ET), or pre-fibrotic / early primary myelofibrosis (pre-MF).

67. The method according to claim 66, wherein the subject has received a JAK / STAT antagonist as treatment for myelogenic fibrosis.

68. The method according to any one of claims 65 to 67, wherein the subject has been treated with a JAK / STAT antagonist for 2 to 6 weeks.

69. The method according to any one of claims 65 to 68, wherein the JAK / STAT antagonist is selective for JAK1 or JAK2.

70. The method according to any one of claims 65 to 68, wherein the JAK / STAT antagonist is not active against ACVR1 / ALK2.

71. The method according to any one of claims 65 to 70, wherein the JAK / STAT antagonist is ruxolitinib, fedratinib, pacritinib, baricitinib, tofacitinib, oclacitinib, or NSC13626.

72. The method according to any one of claims 65 to 70, wherein the JAK / STAT antagonist inhibits IL6-mediated STAT3 activation.

73. The method according to any one of claims 65 to 70, wherein the JAK / STAT antagonist is GS-0387 or CYT-387.

74. The method according to any one of claims 1 to 73, further comprising administering one or more additional therapeutic agents to a target.

75. The method according to claim 74, wherein the additional therapeutic agent is selected from a GDF trap, a bromodomain and extra-terminal domain (BET) inhibitor, a erythropoiesis stimulant, or an immunomodulator.

76. The method according to claim 75, wherein the GDF trap is sotatercept, raspatercept, or KER-050.

77. The method according to claim 75, wherein the BET inhibitor is CPI-0610.

78. The method according to claim 75, wherein the immunomodulator / erythropoietin stimulant is pomalidomide, danazol, prednisone, thalidomide, or lenalidomide.

79. The method according to claim 75, wherein the red blood cell production stimulant is erythropoietin (EPO).

80. A method for treating anemia in a subject with myelofibrosis, comprising administering an effective dose of a hepcidin antagonist and one or more additional therapeutic agents to the subject.

81. The method according to claim 80, wherein the hepcidin antagonist is an HJV-inducing BMP signaling antagonist or a hepcidin neutralizer.

82. The method according to claim 81, wherein the HJV-induced BMP signaling antagonist is a BMP antagonist, an HJV antagonist, a modified heparin that targets BMP6, or recombinant SMAD6 or SMAD7.

83. The method according to claim 82, wherein the BMP antagonist is a BMP6 neutralizing antibody selected from LY311359, CSJ137, and KY1070.

84. The method according to claim 81, wherein the HJV-induced BMP signaling antagonist is an HJV antagonist.

85. The method according to claim 84, wherein the HJV antagonist is an anti-HJV antibody.

86. The method according to any one of claims 80 to 85, wherein the additional therapeutic agent is selected from a GDF trap, a JAK / STAT inhibitor, a BET inhibitor, a erythropoiesis stimulant, or an immunomodulator / erythropoietin stimulant.

87. The method according to claim 86, wherein the additional therapeutic agent is a GDF trap.

88. The method according to claim 87, wherein the GDF trap is sotatercept, raspatercept, or KER-050.

89. The method according to claim 86, wherein the JAK / STAT inhibitor is ruxolitinib, fedratinib, pacritinib, baricitinib, tofacitinib, oclacitinib, NSC13626, or momerotinib.

90. The method according to claim 86, wherein the BET inhibitor is CPI-0610.

91. The method according to claim 86, wherein the immunomodulator / erythropoietin stimulant is pomalidomide.

92. The method according to claim 86, wherein the erythrocyte production stimulant is erythropoietin (EPO).

93. A method for treating a subject who has or is at risk of having an adverse reaction to a JAK-STAT antagonist, comprising administering an effective dose of a hemoduverin-induced BMP signaling antagonist to the subject.

Images