Compounds for Treating MDS-Associated Anemia and Other Conditions

Abstract

Provided herein is the use of certain pyruvate kinase activators, or pharma- ceutically acceptable salts or compositions thereof, for treating MDS-associated anemia and other conditions.

Description

[Technical field]

[0001] Related Applications This application claims priority to U.S. Provisional Application No. 63 / 280,069, filed November 16, 2021, and U.S. Provisional Application No. 63 / 357,240, filed June 30, 2022, each of which is incorporated herein in its entirety. [Background technology]

[0002] Myelodysplastic syndromes (MDS) are a heterogeneous group of rare hematologic malignancies characterized by impaired hematopoiesis, progressive cytopenias, and an increased risk of progression to acute myeloid leukemia (AML). MDS occurs when blood-forming cells in the bone marrow become abnormal (dysplastic) and have trouble making new blood cells. Many of the blood cells formed by bone marrow cells often die or are destroyed by the body, thus leaving the individual with not enough normal blood cells. Although different cell types are affected by this phenomenon, the most common finding in MDS is a shortage of red blood cells (anemia). However, hemolytic anemia, although rare, has also been found to occur in patients suffering from MDS. See, for example, Leukemia Research Reports, Vol. 5, 2016, pp 23-26.

[0003] MDS patients, whose average age at diagnosis is 71 years, can be severely affected by chronically low levels of hemoglobin. This can lead to fatigue, impaired cardiopulmonary function, increased falls, and significant cognitive decline. Thus, treatment of anemia is essential for overall health and quality of life. Currently, transfusion of packed red blood cells (PRBCs) is the standard treatment for MDS-associated anemia. However, the problem for transfusion-dependent MDS-associated anemia patients is that these patients are at high risk for iron overload and transfusion reactions, and report reduced quality of life. Therefore, alternative approaches to treat MDS-associated anemia are needed. [Prior art documents] [Non-patent literature]

[0004] [Non-Patent Document 1] Leukemia Research Reports,Vol.5,2016,pp23-26 Summary of the Invention [Means for solving the problem]

[0005] It has now been discovered that certain activators of pyruvate kinase may also be effective in treating anemia associated with MDS, particularly anemia associated with very low-risk, low-risk and moderate-risk MDS. These activators include those having the following structural formula: [ka] and pharmaceutically acceptable salts thereof. In one aspect, pyruvate kinase activators of the present disclosure have been found to increase hemoglobin levels, decrease reticulocyte counts, and / or improve red blood cell (RBC) function in animal models of MDS. See, e.g., Figures 1 and 2.

[0006] Accordingly, provided herein are methods of treating MDS-associated anemia and other conditions in a subject, comprising administering to the subject one or more of the disclosed pyruvate kinase activators, or pharma- ceutically acceptable salts or compositions thereof.

[0007] Also provided is a method for increasing hemoglobin levels in a subject suffering from MDS, comprising administering to the subject one or more of the disclosed pyruvate kinase activators, or pharma- ceutically acceptable salts or compositions thereof.

[0008] Also provided is a method of treating acquired PK deficiency (PKD) in a subject suffering from MDS, comprising administering to the subject one or more of the disclosed pyruvate kinase activators, or pharma- ceutically acceptable salts or compositions thereof.

[0009] Also provided is a method of treating anemia associated with acquired PKD in a subject suffering from MDS, comprising administering to the subject one or more of the disclosed pyruvate kinase activators, or pharma- ceutically acceptable salts or compositions thereof.

[0010] Also provided is a method of treating cytopenia in a subject suffering from MDS, comprising administering to the subject one or more of the disclosed pyruvate kinase activators, or pharma- ceutically acceptable salts or compositions thereof.

[0011] Further provided is a method of treating hemolytic anemia in a subject suffering from MDS, comprising administering to the subject one or more of the disclosed pyruvate kinase activators, or pharma- ceutically acceptable salts or compositions thereof.

[0012] Further provided is a method of treating ineffective erythropoiesis in a subject suffering from MDS, comprising administering to the subject one or more of the disclosed pyruvate kinase activators, or pharma- ceutically acceptable salts or compositions thereof.

[0013] Also provided are methods of reducing disease progression in a subject suffering from MDS. In some embodiments, administering to a subject one or more of the disclosed pyruvate kinase activators, or pharma- ceutically acceptable salts or compositions thereof, reduces inflammation in the subject's bone marrow, thereby reducing disease progression in the subject.

[0014] Still further provided is a method of treating a disease associated with mitochondrial dysfunction in a subject in need thereof, comprising administering to the subject one or more of the disclosed pyruvate kinase activators, or pharma- ceutically acceptable salts or compositions thereof. [Brief description of the drawings]

[0015] [Figure 1A-B] The effects of mitapivat administration in chow for 6 weeks (Figure 1A), 8 weeks (Figure 1B), 12 weeks (Figure 1C), and 18 weeks (Figure 1D) on hemoglobin levels in a mouse model of MDS-associated anemia are shown. [Figure 1C-D] Same as above. [Figure 2A-B] The hematological parameter results of red blood cell count (RBC) (Figure 2A), reticulocyte percentage (Figure 2B), reticulocyte concentration (Figure 2C), and hematocrit percentage (Figure 2D) from an MDS-associated anemia mouse model after administration of mitapivat are shown. [Figure 2C-D] Same as above. [Diagram 3] 1 shows a multiphase clinical trial design for the treatment of MDS-associated anemia. [Figure 4] 1 shows an overview of a Phase 2a study design with Compound 1 in subjects with anemia due to lower-risk myelodysplastic syndrome (LR-MDS). [Diagram 5] 1 shows an overview of the Phase 2b study design with Compound 1 in subjects with anemia due to lower-risk myelodysplastic syndrome (LR-MDS). [Figure 6A-B] 1 shows flow cytometry data of bone marrow aspirates from PolgD257A mice following treatment with Compound 1. [Figure 6C-D] Same as above. [Figure 6E] Same as above. [Figure 7A-B] 1 shows flow cytometry data of bone marrow aspirates from NHD13 mice following treatment with mitapivat or Compound 1. [Figure 7C-D] Same as above. [Figure 8]The hematological parameter results of hemoglobin (Figure 8A), RBC (Figure 8B), and reticulocyte concentrations (Figure 8C) from NHD13 mice after treatment with mitapivat or Compound 1 are shown. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0016] In a first embodiment, provided herein is a method for treating anemia associated with myelodysplastic syndrome (MDS) in a subject suffering from MDS, comprising administering to the subject a therapeutically effective amount of: [ka] or a pharma- ceutically acceptable salt of any of the foregoing.

[0017] In a second embodiment, provided herein is a method for treating hemolytic anemia associated with myelodysplastic syndrome (MDS) in a subject suffering from MDS, comprising administering to the subject a therapeutically effective amount of: [ka] or a pharma- ceutically acceptable salt of any of the foregoing.

[0018] In a third embodiment, provided herein is a method of increasing hemoglobin levels in a subject suffering from myelodysplastic syndrome (MDS), comprising administering to the subject a therapeutically effective amount of: [ka] or a pharma- ceutically acceptable salt of any of the foregoing.

[0019] In a fourth embodiment, provided herein is a method of treating acquired PK deficiency (PKD) in a subject suffering from myelodysplastic syndrome (MDS), comprising administering to the subject a therapeutically effective amount of: [ka] or a pharma- ceutically acceptable salt of any of the foregoing.

[0020] In a fifth embodiment, provided herein is a method of treating anemia associated with acquired PK deficiency (PKD) in a subject suffering from myelodysplastic syndrome (MDS), comprising administering to the subject a therapeutically effective amount of: [ka] or a pharma- ceutically acceptable salt of any of the foregoing.

[0021] In a sixth embodiment, provided herein is a method of treating cytopenia in a subject suffering from myelodysplastic syndrome (MDS), comprising administering to the subject a therapeutically effective amount of: [ka] or a pharma- ceutically acceptable salt of any of the foregoing.

[0022] In a seventh embodiment, provided herein is a method of treating a disease associated with mitochondrial dysfunction in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of: [ka] or a pharma- ceutically acceptable salt of any of the foregoing.

[0023] In an eighth embodiment, provided herein is a method of treating ineffective erythropoiesis in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of: [ka] or a pharma- ceutically acceptable salt of any of the foregoing.

[0024] As used herein, the term "hemolytic anemia" refers to a subtype of anemia in which a subject's low red blood cell count is caused by the destruction of red blood cells rather than their underproduction. Unless otherwise stated, as used herein, the term "anemia" refers to a low red blood cell count caused by underproduction of red blood cells, including ineffective erythropoiesis.

[0025] The terms "MDS-related anemia," "anemia associated with MDS," and "anemia due to MDS" are synonymous and refer to anemia that develops or is acquired in a subject as a result of having or being affected by MDS.

[0026] "Anemia associated with acquired PK deficiency (PKD)" and "PKD-associated anemia" are synonymous and refer to anemia that develops in a subject as a result of acquired PKD resulting from having or suffering from MDS. In some embodiments, the anemia associated with acquired PK deficiency in a subject suffering from MDS is hemolytic anemia.

[0027] The terms "subject" and "patient" are synonymous and refer to a mammal in need of treatment, such as companion animals (e.g., dogs, cats, etc.), farm animals (e.g., cows, pigs, horses, sheep, goats, etc.), and laboratory animals (e.g., rats, mice, guinea pigs, etc.). Unless stated to the contrary, a subject is a human in need of treatment. In some embodiments, a subject is an adult (e.g., 18 years of age or older). In other embodiments, a subject is a human child (e.g., under 18 years of age). In yet other embodiments, a subject is a human female (adult or child). In yet other embodiments, a subject is a human male (adult or child).

[0028] The terms "administer," "administering," or "administration" refer to providing, implanting, absorbing, ingesting, injecting, inhaling, or otherwise introducing a compound described herein, or a pharma- ceutically acceptable salt or composition thereof, to, into, or onto a subject.

[0029] The terms "treatment", "treat" and "treating" refer to reversing, alleviating, delaying the onset, or inhibiting the progression of one or more diseases described herein. In some embodiments, treatment may be performed after one or more signs or symptoms of a disease have occurred (i.e., therapeutic treatment). In other embodiments, treatment may be performed in the absence of signs or symptoms of a disease. For example, treatment may be performed in a susceptible subject before symptoms develop (e.g., in light of a history of symptoms and / or exposure to a pathogen) (i.e., prophylactic treatment). Treatment may also be continued after symptoms have resolved, e.g., to delay or prevent their recurrence. In certain embodiments, treatment includes delaying the onset of at least one symptom of a disorder for a period of time.

[0030] The term "effective amount" or "therapeutically effective amount" of a compound or a pharma- ceutically acceptable salt thereof described herein refers to an amount of the compound or a pharma- ceutically acceptable salt thereof sufficient to provide a therapeutic benefit in the treatment of a condition described herein. In one embodiment, an effective amount is about 0.01 to about 100 mg / kg body weight / day of a provided compound or a pharma- ceutically acceptable salt, e.g., about 0.1 to about 100 mg / kg body weight / day. In another embodiment, an effective amount is about 0.01 to about 2000 mg of a provided compound or a pharma- ceutically acceptable salt thereof, which may be administered once or twice per day.

[0031] As used herein, the recitation of ranges of values ​​is intended to serve as a shorthand method of referring individually to each separate value falling within the range, as well as the maximum and minimum values ​​defining the range, and unless expressly stated to the contrary, each value is incorporated herein as if it were individually recited herein. For example, a range of values ​​from X to Y includes both X and Y, and all values ​​between X and Y.

[0032] The use of any and all examples or exemplary language (e.g., "such as") provided herein is intended to better illustrate the disclosure and does not limit the scope of the disclosure unless otherwise stated. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the disclosure.

[0033] The term "pharmaceutically acceptable salt" refers to a salt that is suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic reaction, etc., within the scope of sound medical judgment, and is commensurate with a reasonable benefit / risk ratio. Pharmaceutically acceptable salts are well known, for example, Berge et al. describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66, 1-19. Pharmaceutically acceptable salts of the compounds disclosed herein include those derived from suitable inorganic and organic acids. Examples of pharmaceutically acceptable acid addition salts are salts of amino groups, formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, and perchloric acid, or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid, or malonic acid, or by using other methods used in the art, such as ion exchange. Other pharma- ceutically acceptable salts include adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, besylate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecyl sulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, gentisate, hemisulfate, heptanoate, hexanoate, hydroiodide, and the like. Examples of suitable salts include salts of 2-hydroxyethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, tosylate, p-toluenesulfonate, undecanoate, and valerate.

[0034] In one aspect, as part of the ninth embodiment, the compound administered to the subject (e.g., as in any one of the first through eighth embodiments) has the structural formula: [ka] or a pharma- ceutically acceptable salt thereof, which is also referred to herein as the chemical name 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one, or simply Compound 1.

[0035] In one aspect, as part of the tenth embodiment, the compound administered to the subject (e.g., as in any one of the first through eighth embodiments) has the structural formula: [ka] or a pharma- ceutically acceptable salt thereof, which is also referred to herein by its chemical name 2-((1H-pyrazol-3-yl)methyl)-4-methyl-6-((1-methyl-1H-pyrazol-3-yl)methyl)-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one, or simply Compound 2.

[0036] In one aspect, as part of the eleventh embodiment, the compound administered to the subject (e.g., as in any one of the first through eighth embodiments) has the structural formula: [ka] or a pharmaceutically acceptable salt thereof. This compound is also known as "Mitapibat", "AG-348", or by its chemical name "N-(4-(4-(cyclopropylmethyl)piperazine-1-carbonyl)phenyl)quinoline-8-sulfonamide". Alternatively, as part of the eleventh embodiment, mitapivat or a pharmaceutically acceptable salt thereof is administered to the subject in crystalline form (e.g., as in any one of the first through eighth embodiments). In another alternative, as part of the eleventh embodiment, mitapivat or a pharmaceutically acceptable salt thereof is administered to the subject in amorphous form (e.g., as in any one of the first through eighth embodiments). In a further alternative embodiment, as part of the eleventh embodiment, mitapivat or a pharma- ceutically acceptable salt thereof is administered to a subject as a mixture of solid forms (e.g., a mixture of one or more crystalline forms or a mixture of one or more crystalline forms and an amorphous form) (e.g., as in any one of the first through eighth embodiments).

[0037] In one aspect, as part of the twelfth embodiment, the compound administered to the subject (e.g., as in any one of the first through eighth embodiments) is a pharma- ceutically acceptable salt of mitapivat. Alternatively, as part of the twelfth embodiment, the compound administered to the subject (e.g., as in any one of the first through eighth embodiments) is a sulfate salt of mitapivat. In another alternative, as part of the twelfth embodiment, the compound administered to the subject (e.g., as in any one of the first through eighth embodiments) is a hemisulfate salt of mitapivat. In another alternative, as part of the twelfth embodiment, the compound administered to the subject (e.g., as in any one of the first through eighth embodiments) is a hydrated hemisulfate salt of mitapivat. In another alternative, as part of the twelfth embodiment, the compound administered to the subject (e.g., as in any one of the first through eighth embodiments) is mitapivat sulfate or the hemisulfate sesquihydrate salt of mitapivat, also known as 1-(cyclopropylmethyl)-4-(4-(quinoline-8-sulfonamido)benzoyl)piperazin-1-ium hemisulfate sesquihydrate, having formula A: [ka] Formula A. In another alternative, as part of the twelfth embodiment, the compound administered to the subject (e.g., as in any one of the first through eighth embodiments) is a sulfate trihydrate salt also called (and equivalents) mitapivat trihydrate or 1-(cyclopropylmethyl)-4-(4-(quinoline-8-sulfonamido)benzoyl)piperazin-1-ium sulfate trihydrate: [ka] Formula B. The hemisulfate sesquihydrate salt of mitapivat (i.e., mitapivat sulfate) can be crystalline, for example, Form A, as disclosed in U.S. Publication No. 2020 / 0277279. Form A is a crystalline form of Cu Characterized by one or more of the following X-ray powder diffraction patterns at 2θ angles (±0.2°) using Kα radiation: 9.9°, 15.8°, and 22.6°; 15.0°, 17.1°, 21.3°, and 21.9°; 9.9°, 15.0°, 15.8°, 17.1°, 21.3°, 21.9°, and 22.6°; 9.9°, 11.4°, 15.0°, 15.3°, 15.8°, 17.1°, 17.7°, 21.3°, 21.9°, 22.6°, and 23.5°; or 4.9°, 9.9°, 11.0°, 11.4°, 11.7°, 12.3°, 12.8°, 13.6°, 13.9°, 14.2°, 15.0°, 15.3°, 15.8°, 17.1°, 17.4°, 17.7°, 18.8°, 19.1°, 19.8°, 21.3°, 21.9°, 22.6°, 23.0°, 23.2°, 23.5°, 23.8°, 24.1°, 24.5°, 25.3°, 25.6°, 26.1°, 27.1°, 28.1°, and 29.8°. In some embodiments, Form A is characterized by X-ray powder diffraction peaks at 2θ angles (±0.2°) 9.9°, 15.8°, and 22.6°. In certain embodiments, Form A is characterized by X-ray powder diffraction peaks at 2θ angles (±0.2°) 9.9°, 15.8°, and 22.6°, and at least one additional X-ray powder diffraction peak at a 2θ angle (±0.2°) selected from 15.0°, 17.1°, 21.3°, and 21.9°. In certain embodiments, Form A is characterized by X-ray powder diffraction peaks at 2θ angles (±0.2°) 9.9°, 15.8°, and 22.6°, and at least two additional X-ray powder diffraction peaks at 2θ angles (±0.2°) selected from 15.0°, 17.1°, 21.3°, and 21.9°. In yet another alternative, Form A is characterized by X-ray powder diffraction peaks at 2θ angles (±0.2°) of 9.9°, 15.8°, and 22.6°, and at least three additional X-ray powder diffraction peaks at 2θ angles (±0.2°) selected from 15.0°, 17.1°, 21.3°, and 21.9°.In certain embodiments, Form A is characterized by X-ray powder diffraction peaks at 2θ angles (±0.2°) 9.9°, 15.0°, 15.8°, 17.1°, 21.3°, 21.9°, and 22.6°. In certain embodiments, Form A is characterized by X-ray powder diffraction peaks at 2θ angles (±0.2°) 9.9°, 11.4°, 15.0°, 15.3°, 15.8°, 17.1°, 17.7°, 21.3°, 21.9°, 22.6°, and 23.5°. In certain embodiments, Form A is characterized by X-ray powder diffraction peaks at 2θ angles (±0.2°) 4.9°, 9.9°, 11.0°, 11.4°, 11.7°, 12.3°, 12.8°, 13.6°, 13.9°, 14.2°, 15.0°, 15.3°, 15.8°, 17.1°, 17.4°, 17.7°, 18.8°, 19.1°, 19.8°, 21.3°, 21.9°, 22.6°, 23.0°, 23.2°, 23.5°, 23.8°, 24.1°, 24.5°, 25.3°, 25.6°, 26.1°, 27.1°, 28.1°, and 29.8°. In yet another alternative, Form A is characterized by a differential scanning calorimetry (DSC) thermogram comprising endothermic peaks at about 159° C.±5° C. and 199° C.±5° C. In yet another alternative, crystalline Form A is characterized by a thermogravimetric analysis (TGA) thermogram comprising a weight loss of about 4.5±0.5%, up to 180° C.±2° C. In some embodiments, the hemisulfate sesquihydrate salt of mitapivat is 1-(cyclopropylmethyl)-4-(4-(quinoline-8-sulfonamido)benzoyl)piperazin-1-ium hemisulfate sesquihydrate Form A.

[0038] The term "amorphous" refers to a solid that exists in a non-crystalline state or form. Amorphous solids are disordered arrangements of molecules and therefore have no distinguishable crystal lattice or unit cell and therefore no definable long-range order. The solid-state order of a solid can be determined by standard techniques known in the art, for example, by X-ray powder diffraction (XRPD) or differential scanning calorimetry (DSC). Amorphous solids can also be distinguished from crystalline solids, for example, by birefringence using a polarized light microscope.

[0039] The 2-theta values ​​of X-ray powder diffraction patterns for the crystalline forms described herein may vary slightly from instrument to instrument and may be dependent on sample preparation variations and batch-to-batch variations due to factors such as temperature variations, sample shifts, and the presence or absence of an internal standard. Thus, unless otherwise defined, the XRPD patterns / assignments listed herein should not be taken as absolute and may vary by ±0.2 degrees. This variation is well known to those skilled in the art to account for the above factors without precluding unambiguous identification of the crystalline forms. Unless otherwise stated, the 2-theta values ​​provided herein were obtained using Cu Kα1 radiation.

[0040] The compounds or pharma- ceutically acceptable salts described herein, for example, the compounds or pharma- ceutically acceptable salts described in any one of the first to twelfth embodiments, may be formulated and administered as pharmaceutical compositions. The pharmaceutical compositions may be prepared by methods known in the art of pharmacology. In one aspect, the pharmaceutical compositions are orally administered in orally acceptable dosage forms, including, but not limited to, granules or mini-tablets, capsules, tablets, emulsions, and aqueous suspensions, dispersions, and solutions.

[0041] In one aspect, as part of the thirteenth embodiment, a subject treated with a disclosed compound, pharma- ceutically acceptable salt, or composition (including a subject and compound described in any one of the first, second, and fourth through twelfth embodiments) experiences a hemoglobin response.

[0042] As used herein, "hemoglobin response" refers to an increase from a subject's baseline Hb level (i.e., Hb concentration), where the subject's hemoglobin response is measured during treatment with a compound disclosed herein, or a pharma- ceutically acceptable salt or composition thereof, or over a period of time after administration of a compound disclosed herein, or a pharma- ceutically acceptable salt or composition thereof. Unless specified to the contrary, the terms "during treatment" or "after administration," when used in connection with a disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, refer to ongoing treatment or administration (i.e., the subject continues to be treated or administered with a disclosed compound, a pharma- ceutically acceptable salt, or composition thereof). The terms hemoglobin (Hb) level and hemoglobin concentration are used interchangeably herein. As used herein, the term "baseline" refers to a level or concentration measured or established prior to treatment or during treatment with a compound disclosed herein, or a pharma- ceutically acceptable salt or composition thereof. For example, as used herein, the term "baseline hemoglobin level" refers to a subject's hemoglobin (Hb) level measured or established before treatment or during treatment with a compound or a pharma- ceutically acceptable salt or composition disclosed herein. In one embodiment, "hemoglobin response" refers to an increase from a subject's baseline Hb level (i.e., Hb concentration), where the subject's hemoglobin response is measured over a period of time during treatment. In another embodiment, "hemoglobin response" refers to an increase from a subject's baseline Hb level (i.e., Hb concentration), where the subject's hemoglobin response is measured over a period of time after administration, for example, 1 week of treatment, 2 weeks of treatment, 3 weeks of treatment, 4 weeks of treatment, 3 months of treatment, 6 months of treatment, or 1 year or more of treatment.

[0043] In one aspect, as part of the fourteenth embodiment, during treatment with a disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, the hemoglobin level of the treated subject (including any one of the first through thirteenth embodiments) is increased from baseline over a period of at least 1 week, at least 2 weeks, at least 3 weeks, at least 4 weeks, at least 6 weeks, at least 8 weeks, at least 10 weeks, at least 12 weeks, at least 14 weeks, at least 16 weeks, at least 18 weeks, at least 20 weeks, at least 30 weeks, at least 40 weeks, or at least 50 weeks. In one aspect, as part of the fourteenth embodiment, during treatment with the disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, the hemoglobin level of the treated subject (including any one of the first to thirteenth embodiments) increases from baseline for a period of at least 1 week, at least 2 weeks, at least 3 weeks, at least 4 weeks, at least 6 weeks, at least 8 weeks, at least 10 weeks, at least 12 weeks, at least 14 weeks, at least 16 weeks, at least 18 weeks, at least 20 weeks, at least 30 weeks, at least 40 weeks, or at least 50 weeks. Alternatively, as part of the fourteenth embodiment, during treatment with the disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, the hemoglobin level of the treated subject (including any one of the first to thirteenth embodiments) increases from baseline for a period of at least 4 weeks, at least 6 weeks, at least 8 weeks, at least 10 weeks, at least 12 weeks, at least 14 weeks, at least 16 weeks, at least 18 weeks, or at least 20 weeks.In another alternative, as part of the fourteenth embodiment, during treatment with a disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, the hemoglobin level of a subject being treated (including any one of the first through thirteenth embodiments) increases from baseline by 1 week to 20 weeks, by 1 week to 18 weeks, by 1 week to 16 weeks, by 4 weeks to 20 weeks, by 4 weeks to 18 weeks, by 4 weeks to 16 weeks, by 6 weeks to 20 weeks, by 6 weeks to 18 weeks, by 6 weeks to 16 weeks, by 8 weeks to 20 weeks, by 8 weeks to 18 weeks, by 8 weeks to 16 weeks, by 10 weeks to 20 weeks, by 10 weeks to 18 weeks, or by 10 weeks to 16 weeks. In another alternative, as part of the fourteenth embodiment, during treatment with a disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, the hemoglobin level of a treated subject (including any one of the first through thirteenth embodiments) is increased from baseline from 8 weeks to 16 weeks. In another alternative, as part of the fourteenth embodiment, during treatment with a disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, the hemoglobin level of a subject being treated (including any one of the first through thirteenth embodiments) is increased from baseline at two or more, three or more, four or more, five or more, or six or more consecutive time points from 1 week to 20 weeks, from 1 week to 18 weeks, from 1 week to 16 weeks, from 4 weeks to 20 weeks, from 4 weeks to 18 weeks, from 4 weeks to 16 weeks, from 6 weeks to 20 weeks, from 6 weeks to 18 weeks, from 6 weeks to 16 weeks, from 8 weeks to 20 weeks, from 8 weeks to 18 weeks, from 8 weeks to 16 weeks, from 10 weeks to 20 weeks, from 10 weeks to 18 weeks, or from 10 weeks to 16 weeks. In another alternative, as part of the fourteenth embodiment, during treatment with a disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, the hemoglobin level of a treated subject (including any one of the first through thirteenth embodiments) is increased from baseline at two or more consecutive time points from 8 weeks to 16 weeks.In another alternative, as part of the fourteenth embodiment, during treatment with a disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, the hemoglobin level of the treated subject (including any one of the first to thirteenth embodiments) increases from baseline for one or more consecutive weeks, two or more consecutive weeks, three or more consecutive weeks, four or more consecutive weeks, five or more consecutive weeks, six or more consecutive weeks, seven or more consecutive weeks, eight or more consecutive weeks, nine or more consecutive weeks, or ten or more consecutive weeks. In another alternative, during treatment with a disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, as part of the fourteenth embodiment, the hemoglobin level of the treated subject (including any one of the first to thirteenth embodiments) increases from baseline for eight or more consecutive weeks.

[0044] In one aspect, as part of the fifteenth embodiment, during treatment with the disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, the hemoglobin level of the subject being treated (including any one of the first to fourteenth embodiments) is improved by at least 1.0 g / dL (e.g., increases by 1.0 g / dL or more from baseline). Alternatively, during treatment with the disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, as part of the fifteenth embodiment, the hemoglobin level of the subject being treated (including any one of the first to fourteenth embodiments) is improved by at least 1.5 g / dL (e.g., increases by 1.5 g / dL or more from baseline). In another alternative, during treatment with the disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, as part of the fifteenth embodiment, the hemoglobin level of the subject being treated (including any one of the first to fourteenth embodiments) is improved by at least 2.0 g / dL (e.g., increases by 2.0 g / dL or more from baseline). In another alternative, as part of the fifteenth embodiment, during treatment with a disclosed compound, or a pharma- ceutically acceptable salt or composition thereof, the hemoglobin level of a treated subject (including any one of the first through fourteenth embodiments) improves by more than 2.0 g / dL (e.g., increases from baseline by more than 2.0 g / dL).

[0045] In one aspect, as part of the sixteenth embodiment, a subject treated with a disclosed compound, or a pharma- ceutically acceptable salt or composition thereof (including any one of the first through fifteenth embodiments), is classified as a subject that is non-transfusion dependent (NTD) prior to treatment.

[0046] As used herein, the terms non-transfusion dependent (NTD) and "transfusion free" refer to a subject who does not require regular transfusions prior to treatment with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof. In one aspect, an NTD refers to a subject who has less than 3 red blood cell (RBC) units in the 16 weeks prior to administration of a first dose of a disclosed compound or a pharma- ceutically acceptable salt or composition thereof, and has no transfusions in the 8 weeks prior to administration of a first dose of a disclosed compound or a pharma- ceutically acceptable salt or composition thereof.

[0047] As used herein, the term transfusion dependent (TD) refers to a subject who requires regular blood transfusions.

[0048] In one aspect, as part of the seventeenth embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first through fifteenth embodiments) becomes transfusion independent during treatment. In some aspects, as part of the seventeenth embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first through fifteenth embodiments) becomes transfusion independent for a period of time during treatment.

[0049] As used herein, the terms "transfusion independent" or "transfusion independence" are used interchangeably herein and refer to a subject who has not received (i.e., has not had) red blood cell transfusions for a period of time (also referred to herein as "transfusion free"). In some embodiments, the terms "transfusion independent" or "transfusion independent" refer to a subject who has not received red blood cell transfusions for a period of 16 consecutive weeks. In some embodiments, a subject who has not received red blood cell transfusions for a period of 16 consecutive weeks is referred to as transfusion free. In other embodiments, the terms "transfusion independent" or "transfusion independent" refer to a subject who has not received red blood cell transfusions for a period of 8 consecutive weeks or more. In some embodiments, a subject who has not received red blood cell transfusions for a period of 8 consecutive weeks or more is referred to as transfusion free.

[0050] In one aspect, as part of the 18th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the 1st to 15th or 17th embodiments) is classified as having high transfusion burden (HTB). Alternatively, as part of the 18th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the 1st to 15th or 17th embodiments) is classified as having high transfusion burden (HTB) prior to treatment. In another alternative, as part of the 18th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the 1st to 15th or 17th embodiments) is classified as having high transfusion burden (HTB) characterized by receiving at least 8 red blood cell (RBC) units over a 16-week period and having 4 or more transfusion episodes over the course of 8 weeks. In another alternative, as part of the 18th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the 1st to 15th embodiments or the 17th embodiment) is classified as having a high transfusion burden (HTB) characterized by the subject receiving at least 8 RBC units over a 16 week period and having 4 or more transfusion episodes over the course of 8 weeks prior to treatment. In another alternative, as part of the 18th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the 1st to 15th embodiments or the 17th embodiment) is classified as having a high transfusion burden (HTB) characterized by the subject receiving at least 8 RBC units over a 16 week period and having at least 4 RBC units over the course of 8 weeks prior to treatment.

[0051] In one aspect, as part of the 19th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the 1st to 15th embodiments or the 17th embodiment) is classified as having a low transfusion burden (LTB). Alternatively, as part of the 19th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the 1st to 15th embodiments or the 17th embodiment) is classified as having a low transfusion burden (LTB) prior to treatment. In another alternative, as part of the 19th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the 1st to 15th embodiments or the 17th embodiment) is classified as having a low transfusion burden (LTB) characterized by the subject receiving 3 to 7 RBC units over a 16-week period in at least 2 transfusion episodes with up to 3 transfusion episodes over the course of 8 weeks prior to treatment. In another alternative, as part of the nineteenth embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first through fifteenth or seventeenth embodiments) is classified as having a low transfusion burden (LTB), characterized in that the subject, at some point during treatment, receives 3-7 RBC units over a 16-week period in at least two transfusion episodes, with up to three transfusion episodes over the course of eight weeks.

[0052] In one aspect, as part of the twentieth embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first through twenty-first embodiments) becomes transfusion independent for one or more consecutive weeks, two or more consecutive weeks, three or more consecutive weeks, four or more consecutive weeks, five or more consecutive weeks, six or more consecutive weeks, seven or more consecutive weeks, eight or more consecutive weeks, nine or more consecutive weeks, or ten or more consecutive weeks during treatment. Alternatively, as part of the twentieth embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first through twenty-first embodiments) becomes transfusion independent for eight or more consecutive weeks during treatment. In another alternative, as part of the twentieth embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first through twenty-first embodiments) becomes transfusion independent for eight or more consecutive weeks during treatment, and the subject is classified as having a low transfusion burden (LTB) prior to treatment.

[0053] In one aspect, as part of the twenty-first embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first through twentieth embodiments) experiences a change from baseline in total transfused red blood cell (RBC) units during treatment. In one aspect, as part of the twenty-first embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first through twentieth embodiments) experiences a decrease in total transfused red blood cell (RBC) units during treatment compared to the subject's baseline total transfused red blood cell (RBC) units.

[0054] In one aspect, as part of the twenty-second embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first through twenty-first embodiments) experiences a change from baseline in total transfused red blood cell (RBC) units characterized by a decrease of 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, or 80% or more in total transfused RBC units during treatment. Alternatively, as part of a twenty-second embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first through twenty-first embodiments) experiences a change from baseline in total transfused red blood cell (RBC) units characterized by a decrease of 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, or 80% or more in total transfused red blood cell (RBC) units over a period of 1 or more consecutive weeks, 2 or more consecutive weeks, 3 or more consecutive weeks, 4 or more consecutive weeks, 5 or more consecutive weeks, 6 or more consecutive weeks, 7 or more consecutive weeks, 8 or more consecutive weeks, 9 or more consecutive weeks, or 10 or more consecutive weeks. In another alternative, as part of the 22nd embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the 1st to 21st embodiments) experiences a change from baseline in total transfused red blood cell (RBC) units characterized by a 50% or greater decrease in total transfused red blood cell (RBC) units during treatment. In another alternative, as part of the 22nd embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the 1st to 21st embodiments) experiences a change from baseline in total transfused red blood cell (RBC) units characterized by a 50% or greater decrease in RBC units over a period of 1 or more consecutive weeks, 2 or more consecutive weeks, 3 or more consecutive weeks, 4 or more consecutive weeks, 5 or more consecutive weeks, 6 or more consecutive weeks, 7 or more consecutive weeks, 8 or more consecutive weeks, 9 or more consecutive weeks, or 10 or more consecutive weeks during treatment.In another alternative, as part of the 22nd embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the 1st to 21st embodiments) experiences a change from baseline in total transfused red blood cell (RBC) units characterized by a 50% or greater decrease in total transfused red blood cell (RBC) units over a period of 8 or more consecutive weeks during treatment. In another alternative, as part of the 22nd embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the 1st to 21st embodiments) experiences a change from baseline in total transfused red blood cell (RBC) units characterized by a 50% or greater decrease in total transfused red blood cell (RBC) units over a period of 8 or more consecutive weeks during treatment, and the subject is classified as having a high transfusion burden (HTB) prior to treatment.

[0055] In one aspect, as part of the twenty-third embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first through twenty-second embodiments) experiences a decrease in 2,3-diphosphoglycerate (2,3-DPG) levels during treatment, as compared to the subject's baseline 2,3-DPG levels.

[0056] In one aspect, as part of the 24th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first to 23rd embodiments) experiences an increase in adenosine triphosphate (ATP) concentration during treatment (compared to the subject's baseline ATP concentration). Alternatively, as part of the 24th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first to 23rd embodiments) experiences an increase in adenosine triphosphate (ATP) concentration during treatment (compared to the subject's baseline ATP concentration) of more than 50%, more than 55%, more than 60%, more than 65%, more than 70%, and more than 75%. In another alternative, as part of the 24th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first to 23rd embodiments) experiences an increase in adenosine triphosphate (ATP) concentration of 68%, 71%, or 74% during treatment (compared to the subject's baseline ATP concentration). In another alternative, as part of the 24th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first to 23rd embodiments) experiences an increase in adenosine triphosphate (ATP) concentration of greater than 50%, greater than 55%, greater than 60%, greater than 65%, greater than 70%, and greater than 75% (relative to the subject's baseline ATP concentration) after 10, 20, 30, 40, 50, 60, or 65 days of daily administration. In another alternative, as part of the 24th embodiment, a subject treated with a disclosed compound or a pharma- ceutically acceptable salt or composition thereof (including any one of the first to 23rd embodiments) experiences an increase in adenosine triphosphate (ATP) concentration of greater than 68% after 56 days of daily administration, greater than 71% after 56 days of daily administration, or greater than 74% after 56 days of daily administration.

[0057] In one aspect, as part of the 25th embodiment, the MDS described herein (including any one of the 1st to 6th embodiments and the 10th to 24th embodiments) is low-risk MDS as characterized by the Revised International Prognostic Scoring System for MDS (IPSS-R). See, for example, Greenberg PL, Tuechler H, Schanz J, et al. Revised International Prognostic Scoring System for myelodysplastic syndromes. Blood. 2012; 120: 2454-2465. Low-risk MDS includes, for example, an IPSS-R score of 1.5 to 3 or more. Alternatively, as part of the 25th embodiment, the MDS described herein (including any one of the 1st to 6th embodiments and the 10th to 24th embodiments) is low-risk MDS as characterized by the Revised International Prognostic Scoring System for MDS (IPSS-R). Very low-risk MDS, for example, includes an IPSS-R score of 1.5 or less. In another alternative, as part of the 25th embodiment, the MDS described herein (including any one of the 1st to 6th embodiments and the 10th to 24th embodiments) is a moderate-risk MDS as characterized by the International Prognostic Scoring System for MDS Revised (IPSS-R). Moderate-risk MDS, for example, includes an IPSS-R score of more than 3 to 4.5. In another alternative, as part of the 25th embodiment, the MDS described herein (including any one of the 1st to 6th embodiments and the 10th to 24th embodiments), the term "low-risk MDS" encompasses very low-risk MDS and low-risk MDS as described above.

[0058] In one aspect, as part of the 26th embodiment, the subject described herein (including any one of the 1st to 25th embodiments) is a male. Alternatively, as part of the 26th embodiment, the subject described herein (including any one of the 1st to 25th embodiments) is a female. Alternatively, as part of the 26th embodiment, the subject described herein (including any one of the 1st to 25th embodiments) is an adult male. Alternatively, as part of the 26th embodiment, the subject described herein (including any one of the 1st to 25th embodiments) is an adult female. In another alternative, as part of the 26th embodiment, the subject described herein (including any one of the 1st to 25th embodiments) is a male or female child. Alternatively, as part of the 26th embodiment, the subject described herein (including any one of the 1st to 25th embodiments) is an adult (i.e., 18 years of age or older). In yet another alternative, as part of the 26th embodiment, the subject described herein (including any one of the 1st to 25th embodiments) is a child (i.e., under the age of 18).

[0059] In one aspect, as part of the twenty-seventh embodiment, a subject described herein (including any one of the first through twenty-sixth embodiments) is administered a therapeutically effective amount of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one, or a pharma- ceutically acceptable salt thereof. Alternatively, as part of the 27th embodiment, a subject described herein (including any one of the 1st to 26th embodiments) is administered a composition comprising a therapeutically effective amount of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one, or a pharma- ceutically acceptable salt thereof, and a pharma- ceutically acceptable carrier.

[0060] In one aspect, as part of the twenty-eighth embodiment, the therapeutically effective amount of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one disclosed herein (including any one of the first through twenty-seventh embodiments) is 2 mg per day, 3 mg per day, or 5 mg per day. Alternatively, as part of the twenty-eighth embodiment, a therapeutically effective amount of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one disclosed herein (including any one of the first through twenty-seventh embodiments) is 2 mg administered once per day (QD), 3 mg administered QD, or 5 mg administered QD. In another alternative, as part of the twenty-eighth embodiment, a therapeutically effective amount of a pharma- ceutically acceptable salt of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one) disclosed herein (including any one of the first through twenty-seventh embodiments) is an amount equivalent to 2 mg per day, 3 mg per day, or 5 mg per day of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one as the free base.In another alternative, as part of the twenty-eighth embodiment, a therapeutically effective amount of a pharma- ceutically acceptable salt of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one disclosed herein (including any one of the first to twenty-seventh embodiments) is an amount equivalent to 2 mg administered once per day (QD), 3 mg administered QD, or 5 mg administered QD of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one as the free base. In another alternative, as part of the twenty-eighth embodiment, a therapeutically effective amount of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one disclosed herein (including any one of the first through twenty-seventh embodiments) ranges from 0.5 mg to 10 mg QD. In another alternative, as part of the twenty-eighth embodiment, a therapeutically effective amount of a pharma- ceutically acceptable salt of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one disclosed herein (including any one of the first through twenty-seventh embodiments) is an amount equivalent to an amount in the range of 0.5 mg to 10 mg QD of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one as the free base.In another alternative, as part of the twenty-eighth embodiment, a therapeutically effective amount of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one disclosed herein (including any one of the first through twenty-seventh embodiments) ranges from 0.5 mg to 10 mg twice per day (BID). In another alternative, as part of the twenty-eighth embodiment, a therapeutically effective amount of a pharma- ceutically acceptable salt of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one disclosed herein (including any one of the first through twenty-seventh embodiments) is an amount equivalent to a range of 0.5 mg to 10 mg BID of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one as the free base. In another alternative, as part of the 28th embodiment, a therapeutically effective amount of compound 1 disclosed herein (including any one of the first to 27th embodiments) is in the range of 0.25 mg to 15 mg per day. In another alternative, as part of the 28th embodiment, a therapeutically effective amount of a pharma- ceutically acceptable salt of compound 1 disclosed herein (including any one of the first to 27th embodiments) is in the range of 0.25 mg to 15 mg per day as the free base. In another alternative, as part of the 28th embodiment, a therapeutically effective amount of compound 1 disclosed herein (including any one of the first to 27th embodiments) is in the range of 0.25 mg to 2 mg QD or BID, or 1.5 mg to 5.5 mg QD or BID, or 4 mg to 6 mg QD or BID.In another alternative as part of the 28th embodiment, a therapeutically effective amount of a pharma- ceutically acceptable salt of compound 1 disclosed herein (including any one of the first to 27th embodiments) is an amount equivalent to 0.25 mg to 2 mg of QD or BID, or 1.5 mg to 5.5 mg of QD or BID, or 4 mg to 6 mg of QD or BID, as a free base. In another alternative, as part of the 28th embodiment, a therapeutically effective amount of compound 1 disclosed herein (including any one of the first to 27th embodiments) is 1 mg or 5 mg of QD or BID. In another alternative as part of the 28th embodiment, a therapeutically effective amount of a pharma- ceutically acceptable salt of compound 1 disclosed herein (including any one of the first to 27th embodiments) is an amount equivalent to 1 mg or 5 mg of QD or BID, as a free base.

[0061] In one aspect, as part of the 29th embodiment, a subject described herein (including any one of the 1st to 28th embodiments) is treated (i.e., administered a compound, pharma- ceutically acceptable salt, or composition described herein) for a period of at least 12 weeks, at least 14 weeks, at least 16 weeks, at least 24 weeks, at least 30 weeks, or at least 6 weeks. Alternatively, as part of the 29th embodiment, a subject described herein (including any one of the 1st to 28th embodiments) is treated (i.e., administered a compound, pharma- ceutically acceptable salt, or composition described herein) for a period of up to 12 weeks, up to 14 weeks, up to 16 weeks, up to 24 weeks, up to 30 weeks, up to 36 weeks, up to 50 weeks, up to 100 weeks, or up to 160 weeks. In another alternative, as part of the 29th embodiment, a subject described herein (including any one of the 1st through 28th embodiments) is treated (i.e., administered with a compound, pharma- ceutically acceptable salt, or composition described herein) for a period of 16 weeks, 24 weeks, or 156 weeks. In another alternative, as part of the 29th embodiment, a subject described herein (including any one of the 1st through 28th embodiments) is treated (i.e., administered with a compound, pharma- ceutically acceptable salt, or composition described herein) indefinitely or for the life of the subject.

[0062] In one aspect, there is provided a use of one or more of the disclosed pyruvate kinase activators, or pharma- ceutically acceptable salts or compositions thereof, for the manufacture of a medicament for treating a disclosed condition (e.g., as in any one of the above disclosed embodiments). In another aspect, there is also provided a use of one or more of the disclosed pyruvate kinase activators, or pharma- ceutically acceptable salts or compositions thereof, for treating a disclosed condition (e.g., as in any one of the above disclosed embodiments).

[0063] Further details are provided in the Exemplification section below, which is included as part of the present invention. EXAMPLES

[0064] Preparation of compounds Example 1: Preparation of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one (compound 1). 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one can be prepared according to the procedure outlined for compound E8-4 in Example 8A of U.S. Pat. No. 11,040,036, the entire contents of which are incorporated herein by reference. In view of the disclosure of U.S. Pat. No. 11,040,036, pharma- ceutically acceptable salts of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one can also be prepared.

[0065] 2-((1H-pyrazol-3-yl)methyl)-4-methyl-6-((1-methyl-1H-pyrazol-3-yl)methyl)-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one can be prepared according to the procedure outlined for compound E8-41 in U.S. Pat. No. 11,040,036, the entire contents of which are incorporated herein by reference. In view of the disclosure of U.S. Pat. No. 11,040,036, 2-((1H-pyrazol-3-yl)methyl)-4-methyl-6-((1-methyl-1H-pyrazol-3-yl)methyl)-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one pharma- ceutically acceptable salts can also be prepared.

[0066] Mitapivat can be prepared according to the procedures outlined for compound VIII-8 in WO2011 / 002817, the entire contents of which are incorporated herein by reference. Additional methods, salts, and crystalline forms can also be found in WO2016 / 201227, and salts and crystalline forms can be found in WO2019 / 104134 and WO2020 / 237047, each of which is incorporated herein by reference in its entirety.

[0067] MDS-associated anemia mouse model In vivo mouse studies were performed to determine the impact of Mitapivat and Compound 1 on the maturation of erythroblast populations under conditions of ineffective erythropoiesis comparable to MDS patients. To accomplish this, two established mouse models that have been demonstrated to have ineffective erythropoiesis were used. To determine the impact of Mitapivat and Compound 1 under these conditions, bone marrow aspirates were collected from diseased mice and flow cytometry was used to assess the population of erythroblasts at various stages of maturation. Hemoglobin (Hb), red blood cell (RBC) counts, and reticulocyte fractions were also monitored in whole blood on a Sysmex XN-1000™ hematology analyzer every 4 weeks to determine whether the disease burden and impact on erythroblast maturation could be detected peripherally. Pharmacodynamic and pharmacokinetic analyses were also performed to confirm adequate exposure and target engagement of the compounds (data not shown).

[0068] Example 2: Polg D257A Studying Mitapivat in Mouse Models Mitochondrial disruption is associated with the development of anemia in MDS patients (see, e.g., Chen et al. Blood. 2009; 114(19): 4045-53). Mitochondria play a key role in regulating apoptosis, a mechanism that has been highlighted in the inactive hematopoiesis of MDS (Green et al., Science 1998; 281(5381): 1309-1312; Kerbauy et al., Exp Hematol. 2007; 35(11): 1739-1746; Ahlqvist et al. Nature Communications. 2015; 6: 6494). B6.129S7(Cg)-Polg tm1Prol / J or Polg D257A is a mouse model with a mutation in the N-terminus of the DNA polymerase gamma gene that results in impaired mitochondrial proofreading activity and increased mtDNA mutagenesis (Jackson Laboratory #017341). See Chen et al. Blood. 2009; 114(19): 4045-53 and Ahlqvist et al. Nature Communications. 2015; 6: 6494. The accumulation of these mutations leads to deregulated hematopoiesis and elevated apoptosis, comparable to the disease mechanisms seen in MDS patients (Chen et al. Blood. 2009; 114(19): 4045-53). Furthermore, several lines of evidence indicate acquired pyruvate deficiency in MDS and suggest that PKR may be a potential therapeutic target in this indication (Bovin P et al. British Journal of hematology. 1975;18(3):175-87; Valentine WN et al. Blood. 1973;41(6):857-75; Arnold H et al. Clinica Chimica Acta. 1974;57:187-9; Lin G et al. Chin J Hematol. 1997;18(7):350-3). Here, we present the results of a study of PKR-related pyruvate deficiency in MDS and its role in the treatment of MDS. D257AA mouse model was used to evaluate the therapeutic benefit of mitapivat, a PK activator, in this mechanism of anemia.

[0069] Once the anemia phenotype was established by hematological analysis, the Polg D257A Mice were administered 200 mg / kg / day of mitapivat in chow ad libitum for 18 weeks starting at 7.5 months of age. Whole blood was collected into EDTA tubes via the tail vein and analyzed using a Sysmex XN-2000™ hematology system according to the manufacturer's protocol. Hemoglobin, red blood cells (RBCs), and reticulocytes were monitored every 2-4 weeks to track disease progression and severity. After 18 weeks of mitapivat treatment, RBC counts were elevated by 45% and hemoglobin levels increased by 2.4 g / dL compared to untreated mutants. Concomitantly, the frequency of reticulocytes was reduced by 30% in treated mice, suggesting improved erythropoiesis over untreated mice.

[0070] See Figures 1A-D and 2A-D (C57BL / 6J mice were used as controls). Interestingly, male mutants showed signs of therapeutic benefit beginning 6 weeks after treatment with a 1-2 g / dL increase in hemoglobin, whereas female mutants responded after 18 weeks. Commensurate with the increase in hemoglobin was a marked decrease (approximately 30%) in reticulocyte counts. See Figure 2C.

[0071] Example 3: Polg D257A Study of mitapivat and compound 1 in mouse models Polg from Example 2 D257AA second experiment was performed using the model (RRID: IMSR_JAX:017341). In this experiment, 20 mice (10 males and 10 females per group) were administered either mitapivat or compound 1 in chow as described in Example 2. In the mitapivat group, the compound was administered at 200 mg / kg / day, and in the compound 1 group, compound 1 was administered at 10 mg / kg / day. Treatment began when the mice were 4 months old and continued for 8 months (rolling enrollment). After 8 months of treatment (at 12 months of age), bone marrow aspirates were evaluated by flow cytometry using established erythropoiesis markers according to a modified procedure outlined in Suragani et al., Nat Med 2014 Apr;20(4):408-14. Bone marrow aspirates were treated with ACK lysis buffer for 3 minutes to remove red blood cells from the samples. The remaining cells were washed and maintained in PBS+5% FBS throughout the remainder of the procedure. The flow cytometry panel included live / dead (Indo1), B220 (BV421), CD5 (BV421), CD71 (AF700), and Ter119 (PE) antibodies. The gating strategy used for the analysis was as follows: live / B220-CD5- / Ter119+. Cells were then gated into populations using CD71 and FSC as outlined in the literature (Suragani et al., Nat Med 2014 Apr;20(4):408-14; Figure 6E). As shown in the bone marrow flow cytometry data shown in Figures 6A-D, treatment with Compound 1 resulted in a decrease in basophilic erythroblasts (EryA) coupled with an increase in late basophilic and polychromatic erythroblasts (EryB), and normochromatic and reticulocytes (EryC), indicating that treatment with Compound 1 increased Polg D257A This evidence suggests that Polg improved erythroblast maturation in the bone marrow of mice. D257A This suggests that treatment with Compound 1 improves erythropoiesis in the model. D257A An increase in the ProE (protoerythroid) population in mice was also observed, whereas the Polg D257AThere was no additional effect on the ProE population in Compound 1-treated mice (compared to mice with mitapivat-treated Polg D257A Mouse flow cytometry data are not available.

[0072] Analysis of Hb, RBC, and reticulocytes in whole blood showed that vehicle control Polg D257A Mice had a mean Hb concentration of 7.9 g / dL, demonstrating that a significant disease burden had been established in this model. Data not shown. After 24 weeks (six months so far) of treatment, mitapivat reduced the number of Polg D257A In mice, compound 1 increased Hb and RBC counts by 7% without changes in reticulocytes. Data not shown. After 32 weeks of treatment with compound 1, no changes in Hb, RBC count, or reticulocyte fraction were observed. Data not shown.

[0073] Polg D257A The model also supports the use of the disclosed PK activators in the treatment of diseases or disorders associated with mitochondrial dysfunction. Red blood cells lack mitochondria, but are retained late in the process of hematopoiesis. Thus, the effect of the PK activators on increasing erythropoiesis and hemoglobin production is the result of PK activation in blood cell progenitor stem cells. Polg D257AThe model has a proofreading-deficient mitochondrial DNA polymerase and has an extended effect on hematopoietic stem cell (HSC) differentiation (see Cell Stem Cell Volume 8, Issue 5, 6 May 2011, Pages 499-510). The result of rapidly accumulating mitochondrial DNA mutations causes progenitor stem cell differentiation and loss of downstream progenitors. These defects are likely due to reduced ATP generation from dysfunctional mitochondria. HSCs express both PKR and PKM2 during differentiation, and the disclosed PK activators enhance the function of both isozymes, so increased ATP generation from the glycolytic pathway compensates for the loss of ATP generation in cells with mitochondrial dysfunction. The improvement in HSC multilineage stem cell differentiation shown by PK activators points to a potential opportunity for PK activators to support somatic stem cell differentiation in patients where mitochondrial function may be compromised.

[0074] Example 4: NHD13 Mouse Model NUP98-HOXD13 (NHD13) transgenic MDS-associated mouse models are also used to evaluate the therapeutic benefits of the disclosed compounds. See, for example, Lin et al., Blood 2005 Jul 1;106(1):287-95 and Suragani et al., Nat Med 2014 Apr;20(4):408-14. Male and female NHD13 mice aged 4 or 10 months are administered the disclosed compounds daily in chow diet, as appropriate (e.g., mitapivat or compound 1). Whole blood panel and erythrocyte precursor analysis are performed after 2 months of treatment, and the results are analyzed.

[0075] Evidence regarding the effect of the disclosed PKR activators on ineffective erythropoiesis that may be expected to translate to MDS (e.g., very low-risk, low-risk, and moderate-risk MDS) includes that the characteristics of ineffective erythropoiesis are similar between thalassemia and MDS, that PKR activators can improve survival and differentiation of erythroid cells in the bone marrow, and that PKR activators improve RBC function via increasing energy ATP, nucleotide biosynthesis, and antioxidant stress response via activating glycolysis. The proposed human clinical study is shown in Figure 3.

[0076] Example 5: Study of Mitapivat or Compound 1 in the NHD13 Mouse Model (4 months old) Using Example 4, 20 NHD13 mice (10 males and 10 females per group; RRID:IMSR_JAX:010505) aged 4 months were randomized into treatment groups using a matched distribution of hemoglobin levels. Mice were administered either mitapivat or compound 1 for 20 weeks in a manner similar to Example 3 (chow diet, as appropriate). After 20 weeks of treatment with either mitapivat or compound 1, bone marrow aspirates of NHD13 mice were collected as described above and analyzed by flow cytometry. Results from the assessment of the erythroblast population in the bone marrow showed little difference between the erythroblast population from wild-type and NHD13 vehicle control mice. In addition, no significant reduction in survival of vehicle control NHD13 mice was observed, as previously reported (Lin et al.,Neoplasia 2005 July;106(1)). Taken together, the data suggest that the NHD13 model produces a milder form of the disease than described in the literature, complicating the evaluation of treatment effects. As a result, it was difficult to detect a therapeutic benefit from administration of either mitapivat or compound 1 in the flow cytometry data (Figure 7A-D). This mild disease state was also supported in whole blood analysis. At 9 months of age, NHD13 mice on standard chow (vehicle control) had a mean hemoglobin level of 10.4 g / dL, only 4 g / dL lower than wild-type controls. When the effect of treatment in NHD13 mice was evaluated, both mitapivat and compound 1 showed a significant reduction (20%) in the reticulocyte fraction. (Figure 8C) Treatment with mitapivat or compound 1 also showed a significant reduction in reticulocyte counts (15% for mitapivat and 25% for compound 1). Data not shown. These data demonstrate improved erythropoiesis in NHD13 mice with treatment; however, minimal responses in Hb or RBC counts were observed compared to the progressive anemia with vehicle controls (FIGS. 8A-B).

[0077] Example 6: Study of Mitapivat or Compound 1 in the NHD13 Mouse Model (10 Months Old) A second experiment with 10-month-old NHD13 mice can also be performed in the hope that the disease burden will be more advanced and detectable by flow cytometry in the bone marrow, as described in Example 5. For this study, NHD13 mice can be randomized into 20 mice (10 males and 10 females) per treatment group, with a matched distribution of hemoglobin levels at 10 months of age. As described in Example 5, either mitapivat and / or compound 1 can be administered for 8 weeks on chow fed ad libitum. Hemoglobin, red blood cell count, and reticulocyte fraction can be monitored every 4 weeks. Terminal collection, including bone marrow flow cytometry analysis, can occur at 12 months of age.

[0078] A Phase 2a / 2b Study of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one in Subjects With Anemia Due to Lower-Risk Myelodysplastic Syndromes A Phase 2a / 2b multicenter study is being conducted to evaluate the efficacy and safety of 2-((1H-pyrazol-3-yl)methyl)-6-((6-aminopyridin-2-yl)methyl)-4-methyl-4H-thiazolo[5',4':4,5]pyrrolo[2,3-d]pyridazin-5(6H)-one (referred to herein as Compound 1) in subjects with anemia due to lower-risk myelodysplastic syndromes (LR-MDS).

[0079] research design The Phase 2a portion of the study is a single-arm evaluation of one dose level of Compound 1 (5 mg once per day [QD]) to establish proof-of-concept for Compound 1 in LR-MDS. Eligible subjects will receive Compound 1 for oral administration for a 16-week core period. Subjects who complete the 16-week core period will be eligible to continue receiving the same dose of Compound 1 for up to 156 weeks during an extension period. An overview of the Phase 2a study design is shown in Figure 4.

[0080] The Phase 2b portion of the study is a double-blind, randomized, placebo-controlled evaluation of the efficacy and safety of Compound 1 (2 mg QD, 3 mg QD, and 5 mg QD) compared to placebo. Initiation of the Phase 2b portion of the study will be based on pre-specified GO / NO-GO criteria. Eligible subjects will be randomized in a 1:1:1:1 ratio to receive Compound 1 at 2 mg QD (dose level 1), Compound 1 at 3 mg QD (dose level 2), Compound 1 at 5 mg QD (dose level 3), or matched placebo for QD oral administration. Subjects who complete the 24-week double-blind period will be eligible to receive Compound 1 for up to 156 weeks in the extension period. Randomization will be stratified by baseline transfusion burden (low transfusion burden [subjects who are NTD and subjects with LTB], high transfusion burden (HTB)). An overview of the Phase 2b study design is shown in Figure 5.

[0081] During the Phase 2b extension period, all subjects will receive Compound 1. Subjects who received placebo during the double-blind period will receive 2 mg QD Compound 1 (dose level 1), 3 mg QD Compound 1 (dose level 2), or 5 mg QD Compound 1 (dose level 3) and will be randomized 1:1:1 to allow for evaluation of the long-term safety and efficacy of these multiple doses of Compound 1. Subjects who received Compound 1 during the double-blind period will be eligible to continue receiving the same dose of Compound 1.

[0082] Inclusion criteria Subjects are eligible for inclusion in the Phase 2a portion of the study if all of the following criteria apply: 1. At least 18 years of age at the time of providing informed consent. 2. Documented diagnosis of MDS according to the World Health Organization (WHO) classification, fulfilling the IPSS-R classification of low-risk disease (risk score: ≤ 3.5) and < 5% blasts, as determined by the participant's bone marrow biopsy / aspirate during the screening period. 3. Non-transfused or LTB based on transfusion history in the participant's medical record according to the revised IWG 2018 criteria: a. NTD: < 3 RBC units in the 16 week period prior to administration of the first dose of study drug and no transfusion in the 8 week period prior to administration of the first dose of study drug, or b. LTB: 3-7 RBC units in the 16 week period prior to administration of the first dose of study drug and < 4 RBC units in the 8 week period prior to administration of the first dose of study drug. 4. Hb concentration <11.0 g / dL during the 4-week screening period. Eastern Cooperative Oncology Group (ECOG) performance status score of 5.0, 1, or 2. 6. If receiving iron chelation therapy, the dose of iron chelation therapy must be stable and have been initiated ≥ 56 days prior to administration of the first dose of study medication. 7. Women of childbearing potential (WOCBP) and men with partners who are WOCBP must either abstain from sexual activities that could lead to pregnancy as part of their usual lifestyle or agree to use two forms of contraception, one of which must be considered highly effective from the time they provide informed consent, throughout the study, for 28 days after the last dose of study drug for women, and for 90 days after the last dose of study drug for men. The second form of contraception may be an acceptable barrier method. 8. Written informed consent from the participant is given before any study-related procedures are performed and the participant is willing to comply with all study procedures for the duration of the study.

[0083] Subjects are eligible for inclusion in the Phase 2b portion of the study if all of the following criteria apply: 1. At least 18 years of age at the time of providing informed consent. 2. Documented diagnosis of MDS according to the WHO classification, fulfilling the IPSS-R classification of low-risk disease (risk score: ≤ 3.5) and < 5% blasts, as determined by the participant's bone marrow biopsy / aspirate during the screening period. 3. Non-transfused, with LTB, or with HTB based on transfusion history in the participant's medical record according to the revised IWG 2018 criteria: a. NTD: < 3 RBC units in the 16 weeks prior to randomization and no transfusions in the 8 weeks prior to randomization, or b. LTB: 3-7 RBC units in the 16 weeks prior to randomization and < 4 RBC units in the 8 weeks prior to randomization, or c. HTB: 8 or more RBC units in the 16 weeks prior to randomization and 4 or more RBC units in the 8 weeks prior to randomization. 4. Hb concentration <11.0 g / dL during the 4-week screening period. 5. Up to two prior therapies including erythropoiesis-stimulating agents (ESAs) (e.g., erythropoietin [EPO], EPO plus granulocyte-colony stimulating factor [G-CSF]) and / or luspatercept. 6.ECOG performance status score of 0, 1, or 2. 7. If receiving iron chelation therapy, the dose of iron chelation therapy must be stable and have been initiated ≥56 days prior to randomization. 8. WOCBP and men with partners who are WOCBP must either abstain from sexual activities that could lead to pregnancy as part of their usual lifestyle or agree to use two forms of contraception, one of which must be considered highly effective from the time they provide informed consent, throughout the study, for 28 days after the last dose of study drug for women, and for 90 days after the last dose of study drug for men. The second form of contraception may be an acceptable barrier method. 9. Participant's written informed consent was given before any study-related procedures were performed and he / she is willing to comply with all study procedures for the duration of the study.

[0084] Exclusion criteria Subjects will be excluded from the Phase 2a portion of the study if any of the following criteria apply: 1. Known history of acute myeloid leukemia (AML). 2. Secondary MDS, defined as MDS known to have arisen as a result of chemical insult or treatment with chemotherapy and / or radiation therapy for other diseases. 3. Prior exposure to pyruvate kinase activators, treatments administered for high-risk MDS (hypomethylating agents [HMA], isocitrate dehydrogenase [IDH] inhibitors, or allogeneic or autologous stem cell transplant), and / or disease-modifying agents (e.g., immunomodulatory agents such as lenalidomide). Participants may not be excluded, at the investigator's discretion, if they received 1 week or less of treatment with a disease-modifying agent 8 weeks or more prior to administration of the first dose of study drug. 4. Currently receiving treatment with luspatercept, EPO, or G-CSF. Treatment with EPO or G-CSF must have been discontinued ≥28 days prior to the first dose of study drug, and treatment with luspatercept must have been discontinued ≥65 days prior to the first dose of study drug. 5. History of active and / or uncontrolled cardiac or pulmonary disease within 6 months prior to providing informed consent, including but not limited to: New York Heart Association Class III or IV heart failure or clinically significant arrhythmia b. Myocardial infarction, unstable angina, or unstable hypertension, high-risk thrombosis, hemorrhagic, embolic, or thrombotic stroke, deep vein thrombosis, or pulmonary or arterial embolism c. Heart rate-corrected QT interval using Fridericia's method of 470 milliseconds or more in female subjects and 450 milliseconds or more in male subjects, excluding right or left bundle branch block. Severe pulmonary fibrosis defined as severe hypoxia, evidence of right-sided heart failure, and greater than 50% radiation pulmonary fibrosis. e. Severe pulmonary hypertension defined by hypoxia, right-sided heart failure, and severe oxygen-related symptoms. 6. History of hepatic and biliary disorders defined by: Serum AST > 2.5 x upper limit of normal (ULN) (except in cases due to hemolysis and / or hepatic iron deposition) and ALT > 2.5 x ULN (except in cases due to hepatic iron deposition) b. Serum bilirubin > ULN when elevation is associated with clinically significant choledocholithiasis, cholecystitis, biliary obstruction, or hepatocellular disease Impaired renal function, as defined by an estimated glomerular filtration rate (eGFR) less than 7.45 mL / min. 8. Active infection requiring systemic antimicrobial therapy at the time of providing informed consent. If antimicrobial therapy is required during the screening period, screening procedures should not be performed while antimicrobial therapy is being administered, and the final dose of antimicrobial therapy should be administered ≥ 7 days prior to administration of the first dose of study drug. 9. Major surgery within 12 weeks prior to administration of the first dose of study drug. Subjects must be fully recovered from any previous surgery prior to administration of the first dose of study drug. 10. History of any malignancy, except for in situ nonmelanoma skin cancer, in situ cervical cancer, or in situ breast cancer. Subjects must have no active disease or have received anticancer treatment for 5 years or less prior to providing informed consent. 11. A positive test for Hepatitis C virus (HCV) antibody (Ab) with evidence of active HCV infection or a positive test for Hepatitis B surface antigen (HBsAg). 12. Positive test for HIV-1 Ab or HIV-2 Ab. 13. Absolute neutrophil count (ANC) < 500 / μL (0.5 × 109 / L). 14. Platelet count ≤ 75,000 / μL (75 × 109 / L) assessed without platelet transfusion within 28 days prior to screening. 15. Nonfasting triglyceride concentration >500 mg / dL. 16. Receiving an inhibitor of P-glycoprotein (P-gp) that has not been discontinued for a time frame equivalent to 5 days or 5 half-lives (whichever is longer) prior to administration of the first dose of study drug. 17. Current enrollment or past participation in any other clinical study involving an investigational treatment or device (within 4 weeks or within a time frame equivalent to 5 half-lives of the investigational study drug prior to administration of the first dose of the study drug, whichever is longer). 18. Known allergy to Compound 1 or its excipients (silicified microcrystalline cellulose, croscarmellose sodium, sodium stearyl fumarate, and Opadry® II blue film coat [polyvinyl alcohol, titanium dioxide, macrogol / polyethylene glycol, talc, FD&C blue #2 / indigo carmine aluminum lake / E132]). 19. Pregnant or breastfeeding. 20. Any medical, hematological, psychological, or behavioral condition, or previous or current treatment, that, in the opinion of the Investigator, may confer an unacceptable risk to participation in the clinical study and / or may confound the interpretation of the clinical study data.

[0085] Subjects will be excluded from the Phase 2b portion of the study if any of the following criteria apply: 1. Known history of AML. 2. Secondary MDS, defined as MDS known to have arisen as a result of chemical insult or treatment with chemotherapy and / or radiation therapy for other diseases. 3. Exposure to Compound 1 in the Phase 2a portion of the study, treatment administered for high-risk MDS (HMA, IDH inhibitors, or allogeneic or autologous stem cell transplant), and / or prior exposure to pyruvate kinase activators, including disease-modifying agents (e.g., immunomodulatory agents such as lenalidomide). If participants received 1 week or less of treatment with a disease-modifying agent 8 weeks or more prior to randomization, they may not be excluded, at the investigator's discretion. 4. Currently receiving treatment with luspatercept, EPO, or G-CSF. Treatment with EPO or G-CSF must have been discontinued ≥28 days prior to the first dose of study drug, and treatment with luspatercept must have been discontinued ≥65 days prior to randomization. 5. History of active and / or uncontrolled cardiac or pulmonary disease within 6 months prior to providing informed consent, including but not limited to: New York Heart Association Class III or IV heart failure or clinically significant arrhythmia b. Myocardial infarction, unstable angina, or unstable hypertension, high-risk thrombosis, hemorrhagic, embolic, or thrombotic stroke, deep vein thrombosis, or pulmonary or arterial embolism c. Heart rate-corrected QT interval using Fridericia's method of 470 milliseconds or more in female subjects and 450 milliseconds or more in male subjects, excluding right or left bundle branch block. Severe pulmonary fibrosis defined as severe hypoxia, evidence of right-sided heart failure, and greater than 50% radiation pulmonary fibrosis. e. Severe pulmonary hypertension defined by hypoxia, right-sided heart failure, and severe oxygen-related symptoms. 6. History of hepatic and biliary disorders defined by: Serum AST > 2.5 x ULN (except in cases due to hemolysis and / or hepatic iron deposition) and ALT > 2.5 x ULN (except in cases due to hepatic iron deposition) b. Serum bilirubin > ULN when elevation is associated with clinically significant choledocholithiasis, cholecystitis, biliary obstruction, or hepatocellular disease Renal dysfunction, as defined by an eGFR less than 7.45 mL / min 8. Active infection requiring systemic antimicrobial therapy at the time of providing informed consent. If antimicrobial therapy is required during the screening period, screening procedures should not be performed while antimicrobial therapy is being administered, and the final dose of antimicrobial therapy should be administered ≥7 days prior to randomization. 9. Major surgery within 12 weeks prior to randomization. Subjects must have fully recovered from previous surgery prior to randomization. 10. History of any malignancy, except for in situ nonmelanoma skin cancer, in situ cervical cancer, or in situ breast cancer. Subjects must have no active disease or have received anticancer treatment for 5 years or less prior to providing informed consent. 11. A positive test for HCV Ab with evidence of active HCV infection or a positive test for HBsAg. 12. Positive test for HIV-1 Ab or HIV-2 Ab. 13. ANC<500 / μL (0.5×109 / L). 14. Platelet count <50,000 / μL (50 × 109 / L) assessed without platelet transfusion within 28 days prior to screening. 15. Nonfasting triglyceride concentration >500 mg / dL. 16. Receiving an inhibitor of P-gp that has not been discontinued for a time frame equivalent to ≥ 5 days or 5 half-lives (whichever is longer) prior to randomization. 17. Current enrollment or past participation in any other clinical study involving an investigational treatment or device (within 4 weeks or within a time frame equivalent to 5 half-lives of study drug prior to randomization, whichever is longer). 18. Known allergy to Compound 1 or its excipients, including placebo (silicified microcrystalline cellulose, microcrystalline cellulose, croscarmellose sodium, mannitol, sodium stearyl fumarate, magnesium stearate, and Opadry® II blue film coat [polyvinyl alcohol, hypromellose, titanium dioxide, lactose monohydrate, macrogol / polyethylene glycol, triacetin, talc, FD&C blue #2 / indigo carmine aluminum lake / E132]). 19. Pregnant or breastfeeding. 20. Any medical, hematological, psychological, or behavioral condition, or previous or current treatment, that, in the opinion of the Investigator, may confer an unacceptable risk to participation in the clinical study and / or may confound the interpretation of the clinical study data.

[0086] Primary endpoint Phase 2a Hemoglobin response Hemoglobin response is defined as a ≥1.5-g / dL increase from baseline in mean Hb concentration from weeks 8 through 16. The proportion of subjects who achieved an Hb response (Hb response rate) will be summarized and two-sided 95% exact CIs using the Clopper-Pearson method will be calculated. Hemoglobin concentrations assessed within 14 days after RBC transfusion will be excluded from the analysis of the primary endpoint. When this exclusion is applied, subjects without any Hb concentration assessment from weeks 8 through 16 will be considered non-responders.

[0087] Transfusion independent Transfusion independence is defined as being transfusion-free for 8 or more consecutive weeks during the core period (only for subjects with LTB). The proportion of subjects who achieved TI (TI rate) will be summarized and two-sided 95% exact CIs using the Clopper-Pearson method will be calculated.

[0088] Secondary Endpoints Phase 2a Secondary endpoints for Phase 2a include: • AEs, SAEs, discontinuations due to AEs, and laboratory abnormalities during the core period. • Hb1.0+ response, defined as an increase of ≥1.0 g / dL from baseline in mean Hb concentration from weeks 8 to 16. • Change from baseline in Hb concentration during the core period. • An increase of ≥1.5 g / dL from baseline in Hb concentration at two or more consecutive time points between weeks 8 and 16. • Change from baseline in total transfused red blood cell (RBC) units during the core period. • A 50% or greater decrease in total transfused RBC units for 8 or more consecutive weeks compared to baseline during the core period. - Plasma concentrations and pharmacokinetic parameters of compound 1 during the core period. • Whole blood concentrations of pharmacodynamic parameters including 2,3-diphosphoglycerate (2,3-DPG) and adenosine triphosphate (ATP) during the core period.

[0089] Exploratory Endpoint Phase 2a Exploratory endpoints for Phase 2a include: • Changes from baseline in exploratory biomarkers, including hepcidin, erythroferon, soluble transferrin receptor, and growth differentiation factor 11 (GDF11), during the core period. • Change from baseline in iron, serum ferritin, total iron binding capacity, and transferrin saturation during the core period. • Change from baseline in markers of erythropoiesis, including absolute and percent reticulocytes, and erythropoietin during the core period. • Change from baseline in bone marrow-derived biomarkers, including erythroid precursors, during the core period. • Change from baseline in markers of hemolysis including indirect bilirubin, lactate dehydrogenase, and haptoglobin during the core period. • Change from baseline in PKR activity during the core period. Changes from baseline during the extension period in: - Total transfused RBC units (only for subjects with LTB) -Hb concentration -Markers of iron metabolism -Markers of erythropoiesis and hemolysis -Bone marrow-derived biomarkers including erythroid precursors -Exploratory biomarkers ●AEs, SAEs, discontinuation due to AEs, and laboratory abnormalities during the extension period.

[0090] Primary endpoint Phase 2b The primary endpoint for Phase 2b is mHI-E response, defined as: 1) a ≥1.5-g / dL increase in Hb concentration from baseline for ≥8 consecutive weeks during the double-blind period (subjects with NTD), 2) transfusion independence defined as no transfusion for ≥8 consecutive weeks during the double-blind period (subjects with LTB only), and 3) a ≥50% decrease in total transfused RBC units compared to baseline for ≥8 consecutive weeks during the double-blind period (subjects with HTB only). Hemoglobin concentrations assessed within 14 days after RBC transfusion are excluded from the analysis of the primary endpoint. With this exclusion applied, subjects with NTD and subjects with LTB are considered non-responders if the subject does not have at least two Hb concentration assessments, separated by ≥8 weeks, by Week 24.

[0091] Secondary Endpoints Phase 2b Secondary endpoints for Phase 2b include: -AEs, SAEs, discontinuations due to AEs, and laboratory abnormalities during the double-blind period. -Change from baseline in Hb concentration during the double-blind period. • Change from baseline in total transfused RBC units from weeks 8 to 24. • Transfusion independence, defined as the absence of blood transfusions for at least 8 consecutive weeks during the double-blind period. • Time to first mHI-E response during the double-blind period. • The maximum duration of mHI-E response in subjects who achieved an mHI-E response during the double-blind period. - Plasma concentrations and pharmacokinetic parameters of compound 1 during the double-blind period. ●Whole blood concentrations of pharmacodynamic parameters including 2,3-DPG and ATP during the double-blind period. • Exposure-response (or pharmacokinetic / pharmacodynamic) relationships between endpoints and relevant pharmacodynamic parameters that are indicative of clinical activity and safety during the double-blind period.

[0092] Exploratory Endpoints Phase 2b Exploratory endpoints for Phase 2b include: Change from baseline in frequency of RBC transfusions during the double-blind period (subjects with LTB and HTB only) Time to first transfusion during the double-blind period (NTD only) Changes from baseline in exploratory biomarkers, including hepcidin, erythroferon, soluble transferrin receptor, and GDF11, during the double-blind period Changes from baseline in iron, serum ferritin, total iron binding capacity, and transferrin saturation during the double-blind period - Changes from baseline in markers of erythropoiesis, including absolute and percent reticulocytes, and erythropoietin, during the double-blind period Changes from baseline in bone marrow-derived biomarkers, including erythroid precursors, during the double-blind period Changes from baseline in markers of hemolysis, including indirect bilirubin, lactate dehydrogenase, and haptoglobin, during the double-blind period Change from baseline in PKR activity during the double-blind period on the anemia subscale of the Functional Assessment of Cancer Therapy (FACT-An) Changes from baseline in the Myelodysplasia Scale Quality of Life (QUALMS) and QUALMS Physical Burden (QUALMS-P) subscales during the double-blind period Patient Global Impression of Severity (PGIS) - Symptoms of anemia by at least one category during the double-blind period compared to baseline, or "no change" if symptoms of anemia were absent or mild at baseline Patient Global Impression of Change (PGIC) - improvement in symptoms of anemia during the double-blind period, or "no change" if symptoms of anemia were absent or mild at baseline based on the PGIS baseline score Change from baseline in the Patient-Reported Outcomes Measurement Information System (PROMIS) Physical Function 4a questionnaire during the double-blind period Changes from baseline during the extension period in: - Total transfused RBC units (subjects with LTB and HTB only) - Frequency of RBC transfusions (subjects with LTB and HTB only) -Hb concentration -Markers of iron metabolism -Markers of erythropoiesis and hemolysis -Bone marrow-derived biomarkers including erythroid precursors -Exploratory biomarkers AEs, SAEs, discontinuation due to AEs, and laboratory abnormalities during the extension period

[0093] Dose modification An excessive Hb response is defined as an increase in Hb concentration above the ULN (by sex) without RBC transfusion. In the case of an excessive Hb response, study drug must be discontinued without RBC transfusion for 4 weeks or more.

[0094] If an acute decrease in platelet count (e.g., ≥50% decrease from baseline) is observed, the investigator should monitor platelet counts weekly and interrupt study drug for up to 28 days if clinically indicated. In case of a second occurrence or Grade 4 platelet count decrease, discontinue study drug.

[0095] Although several embodiments have been described, the scope of the present disclosure should be defined by the appended claims, and not by the specific embodiments represented by the examples. All references cited throughout this application (including literature references, issued patents, published patent applications, and co-pending patent applications) are expressly incorporated herein in their entirety by reference. Unless otherwise defined, all technical and scientific terms used herein are accorded the meaning commonly known to those skilled in the art.

Claims

1. 1. A composition for treating anemia associated with myelodysplastic syndrome (MDS) in a subject suffering from MDS, said composition comprising a compound of the structural formula: 【Chemistry 15】 or a pharmaceutically acceptable salt thereof, wherein said composition in a therapeutically effective amount of said compound or a pharmaceutically acceptable salt thereof is administered to said subject.

2. 1. A composition for treating hemolytic anemia associated with myelodysplastic syndrome (MDS) in a subject suffering from MDS, said composition comprising a compound of the structural formula: 【Chemistry 16】 or a pharmaceutically acceptable salt thereof, wherein said composition in a therapeutically effective amount of said compound or a pharmaceutically acceptable salt thereof is administered to said subject.

3. 1. A composition for increasing hemoglobin levels in a subject suffering from myelodysplastic syndrome (MDS), the composition comprising a compound of the structural formula: 【Chemistry 17】 or a pharmaceutically acceptable salt thereof, wherein said composition in a therapeutically effective amount of said compound or a pharmaceutically acceptable salt thereof is administered to said subject.

4. 1. A composition for treating acquired PK deficiency (PKD) in a subject suffering from myelodysplastic syndrome (MDS), said composition comprising a compound of the structural formula: [Chemistry 18] or a pharmaceutically acceptable salt thereof, wherein said composition in a therapeutically effective amount of said compound or a pharmaceutically acceptable salt thereof is administered to said subject.

5. 1. A composition for treating anemia associated with acquired PK deficiency (PKD) in a subject suffering from myelodysplastic syndrome (MDS), said composition comprising a compound of the structural formula: 【Chemistry 19】 or a pharmaceutically acceptable salt thereof, wherein said composition in a therapeutically effective amount of said compound or a pharmaceutically acceptable salt thereof is administered to said subject.

6. 1. A composition for treating cytopenia in a subject suffering from myelodysplastic syndrome (MDS), said composition comprising a compound of the structural formula: 【Chemistry 20】 or a pharmaceutically acceptable salt thereof, wherein said composition in a therapeutically effective amount of said compound or a pharmaceutically acceptable salt thereof is administered to said subject.

7. 7. The composition of any one of claims 1-6, wherein the subject's hemoglobin levels improve over a period of at least 4 weeks, at least 6 weeks, at least 8 weeks, at least 10 weeks, at least 12 weeks, at least 14 weeks, at least 16 weeks, at least 18 weeks, or at least 20 weeks during treatment.

8. 7. The composition of any one of claims 1-6, wherein the subject's hemoglobin level increases from baseline from 1 to 20 weeks, from 1 to 18 weeks, from 1 to 16 weeks, from 4 to 20 weeks, from 4 to 18 weeks, from 4 to 16 weeks, from 6 to 20 weeks, from 6 to 18 weeks, from 6 to 16 weeks, from 8 to 20 weeks, from 8 to 18 weeks, from 8 to 16 weeks, from 10 to 20 weeks, from 10 to 18 weeks, or from 10 to 16 weeks during treatment.

9. 7. The composition of any one of claims 1 to 6, wherein the subject's hemoglobin level increases from baseline from 8 to 16 weeks during treatment.

10. 7. The composition of any one of claims 1-6, wherein the subject's hemoglobin level increases from baseline at two or more, three or more, four or more, five or more, or six or more consecutive time points from 1 to 20 weeks, from 1 to 18 weeks, from 1 to 16 weeks, from 4 to 20 weeks, from 4 to 18 weeks, from 4 to 16 weeks, from 6 to 20 weeks, from 6 to 18 weeks, from 6 to 16 weeks, from 8 to 20 weeks, from 8 to 18 weeks, from 8 to 16 weeks, from 10 to 20 weeks, from 10 to 18 weeks, or from 10 to 16 weeks during treatment.

11. 7. The composition of any one of claims 1-6, wherein the subject's hemoglobin level increases from baseline at two or more consecutive time points from 8 to 16 weeks during treatment.

12. 7. The composition of any one of claims 1 to 6, wherein the subject's hemoglobin level increases by 1.0 g / dL or more from baseline during treatment.

13. 7. The composition of any one of claims 1 to 6, wherein the subject's hemoglobin level increases by 1.5 g / dL or more from baseline during treatment.

14. 7. The composition of any one of claims 1 to 6, wherein the subject's hemoglobin level increases by 2.0 g / dL or more from baseline during treatment.

15. The composition of any one of claims 1 to 6, wherein the subject becomes transfusion independent during treatment.

16. The composition of any one of claims 1 to 6, wherein the subject is classified as having a low transfusion burden prior to treatment.

17. 7. The composition of any one of claims 1-6, wherein the subject becomes transfusion independent for 1 or more consecutive weeks, 2 or more consecutive weeks, 3 or more consecutive weeks, 4 or more consecutive weeks, 5 or more consecutive weeks, 6 or more consecutive weeks, 7 or more consecutive weeks, 8 or more consecutive weeks, 9 or more consecutive weeks, or 10 or more consecutive weeks during treatment.

18. The composition of any one of claims 1 to 6, wherein the subject becomes transfusion independent for 8 or more consecutive weeks during treatment.

19. 7. The composition of any one of claims 1-6, wherein the subject's total transfused red blood cell (RBC) units decrease by 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, or 80% or more from baseline during treatment.

20. 7. The composition of any one of claims 1-6, wherein the subject's total transfused red blood cell (RBC) units are reduced by 50% or more from baseline during treatment.

21. 7. The composition of any one of claims 1-6, wherein the subject's total transfused red blood cell (RBC) units are reduced by 50% or more from baseline for 1 or more consecutive weeks, 2 or more consecutive weeks, 3 or more consecutive weeks, 4 or more consecutive weeks, 5 or more consecutive weeks, 6 or more consecutive weeks, 7 or more consecutive weeks, 8 or more consecutive weeks, 9 or more consecutive weeks, or 10 or more consecutive weeks during treatment.

22. 7. The composition of any one of claims 1-6, wherein the subject's total transfused red blood cell (RBC) units are reduced by 50% or more from baseline for a consecutive period of 8 weeks or more during treatment.

23. 7. The composition of any one of claims 1 to 6, wherein the MDS is low-risk MDS (as characterized by the Revised International Prognostic Scoring System for MDS (IPSS-R)).

24. 7. The composition of any one of claims 1 to 6, wherein the MDS is very low-risk MDS (as characterized by the International Prognostic Scoring System for MDS-Revised (IPSS-R)).

25. 7. The composition of any one of claims 1 to 6, wherein the MDS is intermediate-risk MDS (as characterized by the International Prognostic Scoring System for MDS-Revised (IPSS-R)).

26. The composition according to any one of claims 1 to 6, wherein the subject is a male.

27. The composition according to any one of claims 1 to 6, wherein the subject is a woman.

28. The composition described in any one of claims 1 to 6, wherein the therapeutically effective amount of the compound is 2 mg per day, 3 mg per day, or 5 mg per day.

29. The composition of claim 1, wherein the therapeutically effective amount of the compound is 2 mg QD, 3 mg QD, or 5 mg QD.

30. A composition described in any one of claims 1 to 6, characterized in that it is administered orally.

31. A composition according to any one of claims 1 to 6 in the form of a tablet or one or more granules.

32. A composition according to any one of claims 1 to 6 in the form of one or more granules.

33. 1. A composition for use in a method for treating anemia associated with lower-risk myelodysplastic syndrome (MDS) in a subject suffering from MDS, said composition comprising a compound of the structural formula: 【Chemistry 21】 or a pharmaceutically acceptable salt thereof, wherein the method comprises orally administering to the subject the composition in an amount of 2 mg per day, 3 mg per day, or 5 mg per day of the compound, or an amount of the pharmaceutically acceptable salt thereof equivalent to 2 mg per day, 3 mg per day, or 5 mg per day of the compound, wherein the subject is classified as transfusion-free, low transfusion burden, or high transfusion burden prior to administration.

34. 34. The composition of claim 33, wherein the subject is classified as having a low transfusion burden prior to treatment.

35. The composition described in claim 34, characterized in that it is administered to the subject for a period of 16 weeks.

36. 35. The composition of claim 34, wherein the subject becomes transfusion independent for 8 or more consecutive weeks during a 16-week administration period.

37. 35. The composition of claim 34, wherein the subject's hemoglobin level increases from baseline by 1.0 g / dL or more, 1.5 g / dL or more, or 2.0 g / dL or more from baseline from week 8 to week 16 of the 16-week administration period.

38. The composition described in claim 34, characterized in that it is administered to the subject for a period of 24 weeks.

39. 39. The composition of claim 38, wherein the subject becomes transfusion independent for 8 or more consecutive weeks during a 24-week administration period.

40. 34. The composition of claim 33, wherein the subject is classified as transfusion-unnecessary prior to treatment.

41. The composition of claim 40, wherein the composition is administered to the subject for a period of 24 weeks.

42. 42. The composition of claim 41, wherein the subject's hemoglobin level increases from baseline by 1.0 g / dL or more, 1.5 g / dL or more, or 2.0 g / dL or more for 8 or more consecutive weeks during a 24-week administration period.

43. 34. The composition of claim 33, wherein the subject is classified as having a high transfusion burden prior to treatment.

44. The composition of claim 42, wherein the composition is administered to the subject for a period of 24 weeks.

45. 44. The composition of claim 43, wherein the subject's total transfused red blood cell (RBC) units are reduced by 50% or more from baseline for a period of 8 or more consecutive weeks during a 24-week administration period.

46. 1. A composition for treating a disease or disorder associated with mitochondrial dysfunction in a subject in need thereof, the composition comprising a compound of the structural formula 【Chemistry 22】 or a pharmaceutically acceptable salt thereof, wherein said composition in a therapeutically effective amount of said compound or a pharmaceutically acceptable salt thereof is administered to said subject.

47. 1. A composition for treating a disease or disorder associated with ineffective erythropoiesis in a subject in need thereof, the composition comprising a compound of the structural formula: 【Chemistry 23】 or a pharmaceutically acceptable salt thereof, wherein said composition in a therapeutically effective amount of said compound or a pharmaceutically acceptable salt thereof is administered to said subject.

48. A composition described in any one of claims 1 to 6, wherein the therapeutically effective amount of the compound is in the range of 0.25 mg to 15 mg per day.

49. A composition described in any one of claims 1 to 6, wherein the therapeutically effective amount of the compound is in the range of 0.25 mg to 2 mg QD or BID, or 1.5 mg to 5.5 mg QD or BID, or 4 mg to 6 mg QD or BID.

50. The composition of claim 5, wherein the anemia associated with acquired PK deficiency (PKD) is hemolytic anemia.

51. The composition of any one of claims 33 to 45, wherein the anemia associated with acquired PK deficiency (PKD) is hemolytic anemia.

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