Application of hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) in rare anemia

Abstract

The present application relates to the use of a hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) in rare anemia. In particular, the present application discloses the use of certain hypoxia-inducible factor-prolamide hydroxylase inhibitors (HIF-PHI) in the treatment of myelodysplastic syndrome anemia (MDS anemia), beta-thalassemia (beta thalassemia) and / or sickle cell diseases (sickle anemia, SCD anemia).

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims priority to Chinese invention patent application 202311391912.1, filed on October 24, 2023, entitled “Application of hypoxia-inducible factor-proline hydroxylase inhibitors (HIF-PHI) in rare anemias”, the entire contents of which are incorporated herein by reference. Technical Field

[0003] The present application belongs to the medical field, and in particular, the present application relates to the application of certain hypoxia-inducible factor-proline hydroxylase inhibitors (HIF-PHI) in the treatment of rare anemias. Background Art

[0004] The main functions of human red blood cells include adsorbing oxygen inhaled from the lungs onto hemoglobin in red blood cells. The adsorbed oxygen is transported to body tissues with blood circulation for cell metabolism. Then the carbon dioxide produced by body cells is adsorbed by hemoglobin in red blood cells, circulated to the lungs with blood, and discharged from the lungs, completing a cycle of inhaling oxygen, transporting oxygen, transporting carbon dioxide, and discharging carbon dioxide to the body. The normal content of human hemoglobin fluctuates with age, gender and other factors. For example, the normal hemoglobin of adult men is between 14-18g / dL, and the normal hemoglobin of adult women is between 12-16g / dL. Erythropoietin (EPO) is a key hormone that maintains the homeostasis of human red blood cells and hemoglobin. The hemoglobin content of healthy people is negatively correlated with their endogenous EPO, that is, when the body's hemoglobin decreases, the endogenous EPO increases to promote the increase of hemoglobin and the growth of red blood cells to compensate for the decrease in hemoglobin, thereby maintaining homeostasis.

[0005] If the absolute number of red blood cells decreases or the hemoglobin decreases, it will lead to anemia. Anemia not only seriously affects the patient's physical health and quality of life, but if anemia is not corrected, it may even threaten the patient's life. There are many types of anemia as a disease, among which anemia caused by chronic kidney disease (CKD) is one of the most common anemias; there are also more types of rare anemias that have long troubled patients and doctors, such as myelodysplastic syndrome anemia (MDS anemia), β-thalassemia (β-thalassemia), sickle cell disease (also known as sickle anemia, or SCD anemia), etc. The clinical manifestations and biological mechanisms of common anemias such as chronic kidney disease anemia and rare anemias such as MDS anemia, β-thalassemia, and SCD anemia are completely different; the existing drug treatments for them are also not exactly the same, which is completely reasonable.

[0006] MDS anemia is a rare, heterogeneous, clonal stem cell disease characterized by abnormal development of myeloid cells, manifested as ineffective hematopoiesis, morbid hematopoiesis, leading to unilineage or multilineage blood cell reduction and high risk of transformation to acute myeloid leukemia. The global incidence of MDS anemia is approximately 2 to 12 / 100,000. 80% of the onset age is over 60 years old, and there are more males than females. MDS anemia patients can be divided into lower-risk and higher-risk groups. In addition to red blood cell transfusions, rhEPO drugs are the main supportive treatment for anemia in MDS patients, but the response rate of rHEPO to MDS anemia is very low.

[0007] β-thalassemia (abbreviated as β-thalassemia) is a common clinical hereditary chronic blood disease. The pathogenesis is that the mutation or deletion of the β-globin gene in hemoglobin (Hb) leads to an imbalance in the number of globin α and β chains, thereby causing erythropoiesis defects or ineffective erythropoiesis, leading to chronic anemia and complications in patients. The highest incidence rates are in the Mediterranean region, Southeast Asia, and the Middle East; the β-thalassemia patients in my country are mainly distributed in southern regions such as Guangdong, Guangxi, Yunnan, Guizhou, Sichuan, Hong Kong, and Macau, with an average prevalence of about 2%. Among them, transfusion-dependent thalassemia (TDT), that is, severe patients often develop in infancy and present with chronic progressive hemolytic anemia, which seriously threatens the quality of life and even life of children. If severe β-thalassemia is not treated, most children will die before the age of 5. Standardized lifelong blood transfusion and iron chelation therapy are the main means of treating severe β-thalassemia, splenectomy is palliative therapy, and hematopoietic stem cell transplantation is the only cure for thalassemia, but its application is limited by the lack of blood donations, high medical expenses, and limited medical resources. Recently, gene therapy has also been used to treat β-thalassemia, but it is extremely expensive, the applicable population is very limited, and its long-term safety is unknown. Recently, the fusion protein red blood cell maturation agent Rotacept targeting TGF-β was approved for severe β-thalassemia, but many patients do not respond to this drug, and the side effects such as joint pain and bone pain are serious. Severe thalassemia is a rare disease in both my country and the United States.

[0008] SCD anemia is another very serious, life-threatening hereditary chronic blood disease that is common in Africans and their descendants. There are approximately 100,000 patients in the United States and more than 3 million patients worldwide. SCD anemia is a rare disease in both my country and the United States. The disease is caused by the mutation of the sixth amino acid of the beta-globin of hemoglobin from hydrophilic, negatively charged glutamic acid to lipophilic valine, which causes the hemoglobin in the patient's red blood cell cytoplasm to physically aggregate / polymerize after the oxygen molecules bound to the tissue are exchanged, causing the red blood cells to collapse into a sickle shape; clinically, it manifests as severe hemolytic anemia, and because the deformed red blood cells block capillaries, it leads to local ischemia, pain, and multiple organ failure. The main drug treatment for SCD anemia is hydroxyurea (HU), but HU itself is a chemotherapy drug with certain side effects of bone marrow suppression, and long-term use can lead to leukemia. In addition, a small molecule drug, voxelotor, recently received conditional approval from the U.S. FDA but has not yet received final formal approval. Its mechanism of action is that it can embed into hemoglobin cavities without oxygen molecules to prevent hemoglobin aggregation, but its clinical efficacy is also limited and cannot improve the patient's quality of life (patient reported outcomes (PRO)).

[0009] Generally speaking, there is currently no safe and effective treatment for the above-mentioned rare anemias, such as myelodysplastic syndrome anemia (MDS anemia), β-thalassemia (β-thalassemia), and sickle cell disease (SCD anemia), especially no safe and effective small molecule drugs.

[0010] Therefore, there is a great need for a safe and effective small molecule drug for treating rare anemias, especially rare anemias such as MDS anemia, β-thalassemia, and SCD anemia. Summary of the invention

[0011] In order to find safe and effective small molecule drugs that can be used to treat rare anemias, especially rare anemias such as MDS anemia, β-thalassemia, and SCD anemia, the inventors of the present application have conducted a lot of research and investigation.

[0012] As one of the treatments for common anemias such as anemia in chronic kidney disease (CKD), oral hypoxia inducible factor prolyl hydroxylase inhibitors (HIF-PHI) have been gradually approved by some regulatory agencies. As a new type of small molecule drug, the principle of HIF-PHI in treating chronic renal anemia is to inhibit the degradation of HIF-α by inhibiting hypoxia inducible factor prolyl hydroxylase, thereby allowing HIF-α to combine with HIF-β to form a transcription factor with transcriptional function, such as HIF dimer, thereby activating multiple downstream target genes, including increasing the production of endogenous EPO, thus achieving a similar effect as recombinant erythropoietin in treating anemia in chronic kidney disease. According to public reports, HIF-PHIs that have entered the clinical treatment of CKD renal anemia include Roxadustat, Daprodustat, Vadadustat, Molidustat, Enarodustat, Desidustat, HIF117 (SSS17), HEC53856, DDO-3055 and FG-2216. Even so, not every HIF-PHI can truly safely and effectively treat anemia in chronic kidney disease. For example, so far, in the United States, only Daprodustat has achieved success in the key phase III clinical trial for the indication of anemia in chronic kidney disease with dialysis, and its new drug application (NDA) has been approved by the FDA (U.S. Food and Drug Administration); Daprodustat has not achieved success in the key phase III clinical trial for anemia in non-dialysis chronic kidney disease in the United States, and its NDA has not been approved by the FDA; Roxadustat has not achieved success in the key phase III clinical trial for anemia in dialysis and non-dialysis chronic kidney disease in the United States, and its NDA has not been approved by the FDA in the end. Therefore, even though so many of the above molecules are considered HIF-PHI, it is not obvious whether they can be proven to be safe and effective in anemia in chronic kidney disease.

[0013] Based on the mechanism of HIF-PHI stabilizing HIF, activating EPO and then increasing EPO, HIF-PHI was once considered an ideal alternative to injectable recombinant erythropoietin rEPO for the treatment of anemia in chronic kidney disease; for this reason, all HIF-PHIs currently reported are being developed for the treatment of anemia in chronic kidney disease. Only roxadustat has been tested for the treatment of anemia in MDS, but the efficacy of roxadustat in MDS in the phase III clinical trial was proven to be no significant difference from placebo and failed; see the news report on May 5, 2023: https: / / www.biospace.com / article / releases / fibrogen-announces-results-for-matterhorn-a-phase-3-clinical-study-of-roxadustat-for-the-treatment-of-anemia-in-patients-with-myelodysplastic-syndromes-mds-study-did-not-meet-primary-endpoint / . Other HIF-PHIs are rarely developed for rare anemias, and the reason is easy to understand because the biological mechanisms of rare anemias such as MDS anemia, β-thalassemia, and SCD anemia are very different from those of chronic kidney disease anemia.

[0014] The failure of some of the above-mentioned HIF-PHIs in the key phase III clinical trials and NDAs for chronic kidney disease anemia in the United States, and the success of the key phase III clinical trial of daptostat in the United States for dialysis chronic kidney disease anemia and its approval by the US FDA, have proven that although all hypoxia-inducible factor-proline hydroxylase inhibitors seem to have similar mechanisms of action, their safety and effectiveness in the most common chronic kidney disease anemia are completely different. In other words, the safety and effectiveness of these HIF-PHIs in chronic kidney disease anemia are not obvious, not to mention the safety and effectiveness in rare anemias such as myelodysplastic syndrome anemia (MDS anemia), β-thalassemia (β-thalassemia) and SCD sickle cell disease (SCD, also known as SCD anemia). In addition, the failure of rosuvastatin in the key phase III clinical trial of MDS anemia in 2023 has made the application of various HIF-PHIs in rare anemias such as MDS anemia, β-thalassemia and SCD anemia more unpredictable.

[0015] However, unexpectedly, after in-depth research, the inventors of the present application found that compounds containing indolizine derivatives (such as the compounds synthesized in Example 24 of WO 2018205928) can effectively increase the red blood cell count, hemoglobin content, and hematocrit in MDS anemia model mice, β-thalassemia model mice and SCD anemia model mice, and improve MDS anemia, β-thalassemia and SCD anemia; further unexpectedly, the above-mentioned indolizine derivatives showed surprising effectiveness in MDS anemia model mice, β-thalassemia model mice and sickle cell disease (sickle type) SCD anemia model mice, respectively; therefore, they have the effect of treating MDS anemia, β-thalassemia and SCD anemia.

[0016] In addition, the inventors of the present application have found through further in-depth research that hypoxia-inducible factor-proamide hydroxylase inhibitor HIF-PHI molecules other than roxadustat may have the function of treating MDS anemia or SCD anemia; and hypoxia-inducible factor-proamide hydroxylase inhibitor HIF-PHI molecules including roxadustat may have the function of treating β-thalassemia.

[0017] Therefore, the first aspect of the present application provides the use of a hypoxia inducible factor-proline hydroxylase inhibitor (HIF-PHI) in the manufacture of a medicament for treating rare anemias.

[0018] The second aspect of the present application provides the use of a hypoxia inducible factor-proline hydroxylase inhibitor (HIF-PHI) in the manufacture of a medicament for increasing the red blood cell count, hemoglobin content and / or hematocrit of rare anemias.

[0019] The third aspect of the present application provides a method for treating rare anemias, comprising: administering a therapeutically effective amount of a hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) to a patient in need of treatment.

[0020] The fourth aspect of the present application provides a method for increasing the red blood cell count, hemoglobin content and / or hematocrit of rare anemia, comprising: administering a therapeutically effective amount of a hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) to a patient in need of treatment.

[0021] The fifth aspect of the present application provides a hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) for treating rare anemias.

[0022] The sixth aspect of the present application provides the use of hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) for treating rare anemias. DETAILED DESCRIPTION

[0023] 1. Hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI)

[0024] In various aspects and various embodiments of the present application, the hypoxia-inducible factor-proline hydroxylase inhibitor can be various hypoxia-inducible factor-proline hydroxylase inhibitor compounds known in the art.

[0025] In some preferred embodiments of the present application, the hypoxia-inducible factor-proline hydroxylase inhibitor may be a HIF-PHI compound or a pharmaceutically acceptable salt thereof that has been approved for clinical use or is undergoing clinical trials. For example, the HIF-PHI may be selected from: Roxadustat, Daprodustat, Vadadustat, Molidustat, Enarodustat, Desidustat, HIF117 (SSS17), HEC53856, and DDO-3055; or the HIF-PHI may be selected from: Daprodustat, Vadadustat, Molidustat, Enarodustat, Desidustat, HIF117 (SSS17), HEC53856, and DDO-3055.

[0026] In some embodiments, the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) is roxadustat or a pharmaceutically acceptable salt thereof.

[0027] In some embodiments, the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) is Daprodustat or a pharmaceutically acceptable salt thereof.

[0028] In some embodiments, the hypoxia-inducible factor-prolamide hydroxylase inhibitor (HIF-PHI) is Vadadustat or a pharmaceutically acceptable salt thereof.

[0029] In some embodiments, the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) is Molidustat or a pharmaceutically acceptable salt thereof.

[0030] In some embodiments, the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) is Enarodustat or a pharmaceutically acceptable salt thereof.

[0031] In some embodiments, the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) is Desidustat or a pharmaceutically acceptable salt thereof.

[0032] In some embodiments, the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) is HIF117 (SSS17) or a pharmaceutically acceptable salt thereof.

[0033] In some embodiments, the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) is HEC53856 or a pharmaceutically acceptable salt thereof.

[0034] In some embodiments, the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) is DDO-3055 or a pharmaceutically acceptable salt thereof.

[0035] In some embodiments, the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) is a compound of formula I disclosed in WO2018205928 or a pharmaceutically acceptable salt thereof.

[0036] In some embodiments, the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) is any mixture of the above-mentioned HIF-PHIs.

[0037] In the present application, unless otherwise specified, when any compound or its pharmaceutically acceptable salt is mentioned or referred to by structural formula, name or code, for example, "compound of Formula I" (synonymous with "compound represented by general formula I"), "rosuvastatin", or "(6'-hydroxy-8'-oxo-3'-phenyl-8'H-spiro[cyclopentane-1,5'-indolizine]-7'-carbonyl)glycine", etc., various forms of these compounds are included, such as their tautomers, optical isomers, geometric isomers, or isomer mixtures (such as racemic mixtures), isotope-modified compounds thereof, and solvates or hydrates of these compounds, as well as their various solid forms (such as various crystalline forms or amorphous forms).

[0038] The term "optical isomer" means that when a compound has one or more chiral centers, each chiral center may exist in the R configuration or the S configuration, and the various isomers thus constituted are optical isomers. Optical isomers include all diastereomers, enantiomers, mesomers, racemates or mixtures thereof. For example, optical isomers can be separated by chiral chromatography or by chiral synthesis.

[0039] The term "geometric isomer" means that when a double bond exists in a compound, the compound may exist as cis isomers, trans isomers, E isomers and Z isomers. Geometric isomers include cis isomers, trans isomers, E isomers, Z isomers or a mixture thereof.

[0040] The term "tautomer" refers to an isomer that is produced by the rapid movement of an atom in a molecule between two positions. Those skilled in the art will appreciate that tautomers can be mutually converted and may reach an equilibrium state and coexist under a certain state. For example, the "compound of formula I" described herein also encompasses any tautomer of the compound of formula I. Specifically, the inventors have discovered that the compound of formula I may exist in the following tautomers Ia, Ib, Ic, or Id:

[0041]

[0042] An "isotopically modified compound" refers to a compound in which any atom in the compound is replaced by an isotope atom thereof. The "isotopically modified compound" in the present application includes all pharmaceutically acceptable isotopically modified compounds of a compound, in which one or more atoms are replaced by atoms having the same atomic number as the atoms usually found in nature but a different atomic mass or mass number.

[0043] Examples of isotopes suitable for inclusion in the compounds of the present application include isotopes of hydrogen such as 2 H(D) and 3 H(T), isotopes of carbon, such as 11 C. 13 C and 14 C, isotopes of chlorine, such as 36 Cl, isotopes of fluorine, such as 18 F, isotopes of iodine, such as 123 I and 125 I, isotopes of nitrogen, such as 13 N and 15 N, isotopes of oxygen, such as 15 O. 17 O and 18 O, and isotopes of sulfur such as 35 S.

[0044] Certain isotopically modified compounds (e.g., those containing radioactive isotopes) are useful in drug and / or substrate tissue distribution studies. Deuterium (i.e., the isotope) is preferred because of its ease of incorporation and convenient means of detection. 2 H), tritium (i.e. 3 H) and carbon-14 (i.e. 14 C) is particularly useful for this purpose.

[0045] Using deuterium (i.e. 2 H) or tritium (i.e. 3 Substitution with heavier isotopes of H) may afford certain therapeutic advantages resulting from greater metabolic stability (for example, increased in vivo half-life or reduced dosage requirements) and hence may be preferred in some circumstances.

[0046] Using positron emitting isotopes such as 11 C. 18 F. 15 O and 13 N) substitution can be used in positron emission topography (PET) research to detect the substrate receptor occupancy state.

[0047] Isotopically modified compounds can generally be prepared by conventional techniques known to those skilled in the art or by using an appropriate isotopically labeled reagent in place of the non-labeled reagent previously used in an analogous manner to those described in the Examples and Preparations appended herein.

[0048] Certain compounds of the present application can exist in unsolvated forms as well as solvated forms, including hydrated forms.

[0049] Certain compounds of the present application may exist in different crystalline or amorphous forms, and any form thereof is included within the scope of the present application.

[0050] In the present application, "pharmaceutically acceptable salt" refers to the inorganic or organic acid addition salt or organic or inorganic base addition salt of the compound suitable for use in mammals (i.e., having safety and effectiveness when used). These salts can be prepared in situ during the final separation and purification of the compound, or by reacting the pure compound in free form with a suitable organic or inorganic acid or base, and separating the formed salt to prepare. Typical salts include hydrobromide, hydrochloride, sulfate, bisulfate, nitrate, acetate, oxalate, valerate, oleate, palmitate, stearate, laurate, borate, benzoate, lactate, phosphate, toluenesulfonate, citrate, maleate, fumarate, succinate, tartrate, glucoheptonate, lactobionate, lauryl sulfonate, etc. These salts can include salts based on alkali and alkaline earth metals (e.g., sodium, lithium, potassium, calcium, magnesium, etc.) cations, as well as non-toxic ammonium, quaternary ammonium, and amine cations (including, but not limited to, ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine, and the like).

[0051] In some preferred embodiments of the present application, the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) is a compound of formula I disclosed in WO2018205928 or a pharmaceutically acceptable salt thereof:

[0052]

[0053] In some embodiments, in the compound represented by formula I:

[0054] R 1 and R 2 independently selected from cyano, alkyl, heterocyclic, alkenyl, alkynyl, aryl, heteroaryl and acyl; wherein the above alkyl, heterocyclic, alkenyl, alkynyl, aryl, heteroaryl, acyl are optionally substituted by one or more substituents, wherein the substituents are independently selected from halogen, cyano, hydroxyl, amino, carboxyl, acyl, alkyl, heterocyclic, alkenyl, alkynyl, aryl, heteroaryl, =O, =S, -SH, R 10 O-、R 10 S-, R 10 (O=)S- and R 10 (O=)2S-, where R 10 is alkyl, heterocyclyl, alkenyl, alkynyl, aryl, or heteroaryl; or R 1 and R 2 Put together to form a ring;

[0055] R 3 is selected from the group consisting of alkyl, heterocyclyl, alkenyl, alkynyl, aryl and heteroaryl;

[0056] R 4 and R 5 is hydrogen;

[0057] R 6 and R 6’ is hydrogen;

[0058] R 7 is hydrogen;

[0059] R 8 is selected from hydrogen and alkyl;

[0060] X is an oxygen atom.

[0061] In some embodiments, in the compound represented by formula I:

[0062] R 1 and R 2independently selected from cyano, C1-C12 alkyl, C2-C12 alkenyl, C2-C12 alkynyl, C6-C14 aryl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, heterocycloalkyl containing 3 to 8 ring atoms, heterocycloalkenyl containing 3 to 8 ring atoms, heteroaryl containing 5 to 14 ring atoms, C1-C12 alkyl-C(=O)- and C2-C12 alkenyl-C(=O)-; wherein the above-mentioned C1-C12 alkyl, C2-C12 alkenyl, C2-C12 alkynyl, C6-C14 aryl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, heterocycloalkyl containing 3 to 8 ring atoms, heterocycloalkenyl containing 3 to 8 ring atoms, heteroaryl containing 5 to 14 ring atoms, C1-C12 alkyl-C(=O)- and C2-C12 alkenyl-C(=O)- -C12 alkynyl, C6-C14 aryl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, heterocycloalkyl containing 3 to 8 ring atoms, heterocycloalkenyl containing 3 to 8 ring atoms, heteroaryl containing 5 to 14 ring atoms, C1-C12 alkyl-C(=O)-, C2-C12 alkenyl-C(=O)- are optionally substituted with 1 to 3 substituents, wherein the substituents are independently selected from hydroxy, halogen, cyano, amino, carboxyl, C1-C6 alkyl, C3-C8 cycloalkyl, C2-C6 alkenyl-, C2-C6 alkynyl-, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, C6-C14 aryl, heteroaryl containing 5 to 14 ring atoms, C1-C6 alkyl-O-, C3-C8 cycloalkyl-O-, C2-C6 alkenyl-O-, C2-C6 alkynyl-O-, C3-C8 cycloalkenyl-O-, C6-C14 aryl-O-, heteroaryl containing 5 to 14 ring atoms, C1-C6 alkyl-S-, C3-C8 cycloalkyl -S-, C2-C6 alkenyl-S-, C2-C6 alkynyl-S-, C3-C8 cycloalkenyl-S-, C6-C14 aryl-S-, heteroaryl-S- containing 5 to 14 ring atoms, heterocycloalkyl containing 3 to 8 ring atoms, heterocycloalkenyl containing 3 to 8 ring atoms, =O, =S, -SH, -CF3, -CO2, C1-C6 alkyl, C1-C6 alkyl-S-, C1-C6 alkyl(O=)S- and C1-C6 alkyl(O=)2S-; or

[0063] R 1 and R 2 taken together to form an optionally substituted cycloalkane, cycloalkene, heterocycloalkane or heterocycloalkene ring containing 3 to 8 ring atoms;

[0064] R 3is selected from the group consisting of C1-C12 alkyl, C2-C12 alkenyl, C2-C12 alkynyl, C6-C14 aryl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, heterocycloalkyl containing 3 to 8 ring atoms, heterocycloalkenyl containing 3 to 8 ring atoms and heteroaryl containing 5 to 14 ring atoms; wherein the above-mentioned C1-C12 alkyl, C2-C12 alkenyl, C2-C12 alkynyl, C6-C14 aryl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, heterocycloalkyl containing 3 to 8 ring atoms, heterocycloalkenyl containing 3 to 8 ring atoms and heteroaryl containing 5 to 14 ring atoms The cycloalkyl group is a C-C8 cycloalkyl, a C3-C8 cycloalkenyl group, a C3-C8 cycloalkyl group, a C3-C8 cycloalkyl group, a C3-C8 cycloalkyl group, a C2-C6 alkenyl group, a C2-C6 alkynyl group, a C3-C8 cycloalkyl group, a C3-C8 cyclo ... C6-C14 aryl, heteroaryl containing 5 to 14 ring atoms, C1-C6 alkyl-O-, C3-C8 cycloalkyl-O-, C2-C6 alkenyl-O-, C2-C6 alkynyl-O-, C3-C8 cycloalkenyl-O-, C6-C14 aryl-O-, heteroaryl containing 5 to 14 ring atoms, C1-C6 alkyl-S-, C3-C8 cycloalkyl-S-, C2-C6 alkenyl-S-, C 2-C6 alkynyl-S-, C3-C8 cycloalkenyl-S-, C6-C14 aryl-S-, heteroaryl-S- containing 5 to 14 ring atoms, heterocycloalkyl containing 3 to 8 ring atoms, heterocycloalkenyl containing 3 to 8 ring atoms, =O, =S, -SH, -CF3, -CO2, C1-C6 alkyl, C1-C6 alkyl-S-, C1-C6 alkyl(O=)S- and C1-C6 alkyl(O=)2S-;

[0065] R 4 and R 5 is hydrogen;

[0066] R 6 and R 6’ is hydrogen;

[0067] R 7 is hydrogen;

[0068] R 8is selected from hydrogen, C1-C12 alkyl and C3-C8 cycloalkyl; wherein C1-C12 alkyl and C3-C8 cycloalkyl are optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxy, halogen, cyano, amino, carboxyl, C1-C6 alkyl, C3-C8 cycloalkyl, C2-C6 alkenyl-, C2-C6 alkynyl-, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, C6-C14 aryl, heteroaryl containing 5 to 14 ring atoms, C1-C6 alkyl-O-, C3-C8 cycloalkyl-O-, C2-C6 alkenyl-O-, C2-C6 alkynyl-O-, C3-C8 cycloalkenyl-O-, C6-C14 aryl-O- and heteroaryl containing 5 to 14 ring atoms;

[0069] X is an oxygen atom.

[0070] In some preferred embodiments of the present application, in the compound represented by general formula I, R 1 and R 2 are independently selected from cyano and unsubstituted C1-C6 alkyl; or R 1 and R 2 Taken together they form an optionally substituted cycloalkane or heterocycloalkane ring containing 3 to 8 ring atoms.

[0071] In some preferred embodiments of the present application, in the compound represented by general formula I, R 1 and R 2 are independently selected from methyl and ethyl; or R 1 and R 2 They are combined to form a cyclopropane ring, a cyclobutane ring, a cyclopentane ring, a cyclohexane ring, a methylcyclohexane ring, a cycloheptane ring, a tetrahydrofuran ring or a tetrahydropyran ring.

[0072] In some preferred embodiments of the present application, in the compound represented by general formula I, R 1 and R 2 Independently selected from cyano and unsubstituted C1-C6 alkyl.

[0073] In some preferred embodiments of the present application, in the compound represented by general formula I, R 1 and R 2 Independently selected from methyl and ethyl.

[0074] In some preferred embodiments of the present application, in the compound represented by general formula I, R 1 and R 2 Taken together they form an optionally substituted heterocycloalkane ring containing 3 to 8 ring atoms.

[0075] In some preferred embodiments of the present application, in the compound represented by general formula I, R 1and R 2 Together they form a tetrahydrofuran ring or a tetrahydropyran ring.

[0076] In some preferred embodiments of the present application, in the compound represented by general formula I, R 1 and R 2 Taken together they form an optionally substituted cycloalkane ring containing 3 to 8 ring atoms.

[0077] In some preferred embodiments of the present application, in the compound represented by general formula I, R 1 and R 2 Together they form a cyclopropane ring, a cyclobutane ring, a cyclopentane ring, a cyclohexane ring, a methylcyclohexane ring or a cycloheptane ring.

[0078] In some preferred embodiments of the present application, in the compound represented by general formula I, R 3 Selected from C1-C6 alkyl, C6-C14 aryl, C3-C8 cycloalkyl and heteroaryl containing 5 to 14 ring atoms; wherein the above-mentioned C1-C6 alkyl, C6-C14 aryl, C3-C8 cycloalkyl are optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxyl, halogen, cyano, amino, carboxyl, C6-C14 aryl and C1-C6 alkyl.

[0079] In some preferred embodiments of the present application, in the compound represented by general formula I, R 3 Selected from methyl, ethyl, propyl, butyl, phenyl, benzyl, tolyl, methoxyphenyl, chlorophenyl, fluorophenyl, bromophenyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and pyridyl.

[0080] In some preferred embodiments of the present application, in the compound represented by general formula I, R 3 Selected from C1-C6 alkyl; wherein the above C1-C6 alkyl is optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxyl, halogen, cyano, amino, carboxyl, C6-C14 aryl and C1-C6 alkyl.

[0081] In some preferred embodiments of the present application, in the compound represented by general formula I, R 3 Selected from methyl, ethyl, propyl and butyl.

[0082] In some preferred embodiments of the present application, in the compound represented by general formula I, R 3 Selected from C6-C14 aryl; wherein the above C6-C14 aryl is optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxyl, halogen, cyano, amino, carboxyl, C6-C14 aryl and C1-C6 alkyl.

[0083] In some preferred embodiments of the present application, in the compound represented by general formula I, R 3 The group consisting of phenyl, benzyl, tolyl, methoxyphenyl, chlorophenyl, fluorophenyl, and bromophenyl is preferably selected from phenyl.

[0084] In some preferred embodiments of the present application, in the compound represented by general formula I, R 3 Selected from C3-C8 cycloalkyl; wherein the above C3-C8 cycloalkyl is optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxyl, halogen, cyano, amino, carboxyl, C6-C14 aryl and C1-C6 chain alkyl.

[0085] In some preferred embodiments of the present application, in the compound represented by general formula I, R 3 Selected from cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.

[0086] In some preferred embodiments of the present application, in the compound represented by general formula I, R 3 is selected from heteroaryl groups containing 5 to 14 ring atoms.

[0087] In some preferred embodiments of the present application, in the compound represented by general formula I, R 3 Selected from pyridyl.

[0088] In some preferred embodiments of the present application, in the compound represented by general formula I, R 8 is selected from hydrogen and C1-C6 alkyl; wherein the C1-C6 alkyl is optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxy, halogen, cyano, amino, carboxyl, C1-C6 alkyl and C1-C6 alkyl-O-.

[0089] In some preferred embodiments of the present application, in the compound represented by general formula I, R 8 Selected from hydrogen, methyl, ethyl, propyl, butyl and pentyl.

[0090] In some preferred embodiments of the present application, in the compound represented by general formula I, R 8 Selected from hydrogen.

[0091] In some preferred embodiments of the present application, in the compound represented by general formula I, R 8 Selected from C1-C6 alkyl; wherein the C1-C6 alkyl is optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxy, halogen, cyano, amino, carboxyl, C1-C6 alkyl and C1-C6 alkyl-O-.

[0092] In some preferred embodiments of the present application, in the compound represented by general formula I, R 8 Selected from methyl, ethyl, propyl, butyl and pentyl.

[0093] In some preferred embodiments of the present application, in the compound represented by general formula I, R 1 and R 2 taken together to form an optionally substituted cycloalkane ring containing 3 to 8 ring atoms; R 3 is selected from C6-C14 aryl; wherein the above C6-C14 aryl is optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxyl, halogen, cyano, amino, carboxyl, C6-C14 aryl and C1-C6 alkyl; and R 8 is selected from hydrogen and C1-C6 alkyl; wherein the C1-C6 alkyl is optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxy, halogen, cyano, amino, carboxyl, C1-C6 alkyl and C1-C6 alkyl-O-.

[0094] In some preferred embodiments of the present application, in the compound represented by general formula I, R 1 and R 2 Each is independently selected from C1-C6 alkyl, wherein the above C1-C6 alkyl is optionally substituted by 1 to 3 C1-C6 alkyl substituents; or R 1 and R 2 Taken together, they form a cyclopropane ring, a cyclobutane ring, a cyclopentane ring, a cyclohexane ring, a methylcyclohexane ring, a cycloheptane ring or a tetrahydropyran ring; R 3 is selected from C6-C14 aryl; wherein the above C6-C14 aryl is optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxyl, halogen, cyano, amino, carboxyl, C6-C14 aryl and C1-C6 alkyl; and R 8 is selected from hydrogen and C1-C6 alkyl; wherein the C1-C6 alkyl is optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxy, halogen, cyano, amino, carboxyl, C1-C6 alkyl and C1-C6 alkyl-O-.

[0095] In some preferred embodiments of the present application, in the compound represented by general formula I, R 1 and R 2 Together they form a cyclopropane ring, a cyclobutane ring, a cyclopentane ring, a cyclohexane ring, a methylcyclohexane ring, and a cycloheptane ring; R 3is selected from C6-C14 aryl; wherein the above C6-C14 aryl is optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxyl, halogen, cyano, amino, carboxyl, C6-C14 aryl and C1-C6 alkyl; and R 8 Selected from hydrogen.

[0096] In some particularly preferred embodiments of the present application, the compound represented by general formula I is a compound (6'-hydroxy-8'-oxo-3'-phenyl-8'H-spiro[cyclopentane-1,5'-indolizine]-7'-carbonyl)glycine, wherein R 1 and R 2 Together with the C atoms to which they are attached, they form a cyclopentane ring (i.e., R 1 and R 2 -CH2CH2CH2CH2-), R 3 is phenyl, R 4 and R 5 Hydrogen, R 6 and R 6’ is hydrogen, R 7 is hydrogen, R 8 is a compound of formula I wherein R is hydrogen and X is an oxygen atom.

[0097] In the above different preferred embodiments, the preferred options of each substituent can be combined with each other, and various combinations thereof are within the scope of the present application.

[0098] To avoid ambiguity, the following definitions are given for various terms used in this article. Unless otherwise specified, the meanings of the terms used in this article are as follows.

[0099] The term "hydroxy" refers to -OH.

[0100] The term "halogen" or "halo" refers to -F, -Cl, -Br, or -I.

[0101] The term "amino" refers to -NH2.

[0102] The term "cyano" refers to -CN.

[0103] The term "carboxy" refers to -C(=O)OH.

[0104] The term "substituted" means that one or more (preferably 1 to 5, more preferably 1 to 3) hydrogen atoms in the group are independently replaced by a corresponding number of substituents.

[0105] The term "independently" means that when there are more than one substituent, these substituents may be the same or different.

[0106] The term "optional" or "optionally" means that the event it describes can occur or not occur. For example, a group "optionally substituted" means that the group can be unsubstituted or substituted.

[0107] The term "heteroatom" as used herein refers to oxygen (O), nitrogen (N), or S (O) m (wherein m can be 0, 1 or 2, i.e. a sulfur atom S, or a sulfoxide group SO, or a sulfonyl group S(O)2).

[0108] The term "alkyl" refers to a group formed when a saturated hydrocarbon composed of only two elements, C and H, loses a hydrogen atom from any carbon atom. The "alkyl" described herein includes chain alkyl groups such as straight chain alkyl groups and branched chain alkyl groups; it also includes cycloalkyl groups such as monocyclic alkyl groups, spirocyclic alkyl groups, condensed cycloalkyl groups and bridged cycloalkyl groups. Preferably, the "alkyl" in this application refers to chain alkyl groups. Alkyl groups may be unsubstituted or substituted.

[0109] The "alkyl" described herein includes optionally substituted chain alkyl groups, which preferably have 1 to 20 carbon atoms, more preferably 1 to 12 carbon atoms, and most preferably 1 to 6 carbon atoms; for example, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl, n-hexyl, chloromethyl, fluoroethyl, trifluoromethyl or 1,1,1-trifluoroethyl, etc.

[0110] The "alkyl" described herein also includes optionally substituted cycloalkyl (e.g., C3-C20 cycloalkyl or C3-C12 cycloalkyl or C3-C8 cycloalkyl), for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, decalinyl, norpinyl, adamantyl, fluorocyclopropyl, 2-iodocyclobutyl, 2,3-dimethylcyclopentyl, 2,2-dimethoxycyclohexyl and 3-phenylcyclopentyl, etc.

[0111] "C1-C6 chain alkyl" is also called "lower chain alkyl", which refers to a subset of alkyl, which refers to straight chain alkyl and branched chain alkyl having 1 to 6 carbon atoms, including, for example, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl, n-hexyl, etc.

[0112] The "alkyl" described herein is optionally substituted by one or more substituents, wherein the substituents are independently selected from halogen, cyano, nitro (-NO2), hydroxyl, amino, carboxyl, acyl, alkyl, heterocyclyl, alkenyl, alkynyl, aryl, heteroaryl, =O, =S, -SH, R 16 O-、R 16 S-, R 16 (O=)S-、R 16 (O=)2S-, where R 16is an alkyl group, a heterocyclyl group, an alkenyl group, an alkynyl group, an aryl group, or a heteroaryl group.

[0113] Preferably, the "alkyl" described herein is optionally substituted by 1 to 3 substituents, wherein the substituents are independently selected from hydroxy, halogen, nitro, cyano, amino, carboxyl, C1-C6 alkyl, C3-C8 cycloalkyl, C2-C6 alkenyl-, C2-C6 alkynyl-, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, C6-C14 aryl, heteroaryl containing 5 to 14 ring atoms, C1-C6 alkyl-O-, C3-C8 cycloalkyl-O-, C2-C6 alkenyl-O-, C2-C6 alkynyl-O-, C3-C8 cycloalkenyl-O-, C6-C14 aryl- O-, heteroaryl-O- containing 5 to 14 ring atoms, C1-C6 alkyl-S-, C3-C8 cycloalkyl-S-, C2-C6 alkenyl-S-, C2-C6 alkynyl-S-, C3-C8 cycloalkenyl-S-, C6-C14 aryl-S-, heteroaryl-S- containing 5 to 14 ring atoms, heterocycloalkyl containing 3 to 8 ring atoms, heterocycloalkenyl containing 3 to 8 ring atoms, =O, =S, -SH, -CF3, -CO2, C1-C6 alkyl, C1-C6 alkyl-S-, C1-C6 alkyl(O=)S- and C1-C6 alkyl(O=)2S-

[0114] The term "alkenyl" refers to any hydrocarbon compound consisting of only two elements, C and H, and containing one or more carbon-carbon double bonds but no carbon-carbon triple bonds or aromatic bonds, formed after any carbon atom loses a hydrogen atom. The "alkenyl" described herein includes alkenyls such as straight-chain alkenyls and branched-chain alkenyls; and also includes cycloalkenyls such as monocyclic alkenyls, spirocyclic alkenyls, fused-cyclic alkenyls and bridged-cyclic alkenyls. Alkenyls may be unsubstituted or substituted.

[0115] The “alkenyl” described herein includes optionally substituted alkenyl groups, preferably having 2 to 20 carbon atoms, more preferably having 2 to 12 carbon atoms, and most preferably having 2 to 6 carbon atoms; for example, ethenyl, 1-propenyl, 2-propenyl, 1-butenyl, 2-butenyl, isobutenyl, 1-pentenyl, 2-pentenyl, isopentenyl, 1-hexenyl, 2-hexenyl, 3-hexenyl, isohexenyl, 1-heptenyl, 2-heptenyl, 3-heptenyl, 1-octenyl, 2-octenyl, 3-octenyl, 4-octenyl, 1-nonenyl, 1-decenyl, 1-undecenyl, 1-dodecenyl, and the like.

[0116] The term "alkenyl" described herein also includes optionally substituted cycloalkenyl groups (e.g., C3-C20 cycloalkenyl or C3-C12 cycloalkenyl or C3-C8 cycloalkenyl), for example, cyclobutenyl, cyclopentenyl, cyclohexenyl, cycloheptenyl, cyclobutadienyl, cyclopentadienyl, cycloheptatrienyl, and the like.

[0117] The "alkenyl" described herein is optionally substituted by one or more (eg, 1 to 3) substituents, wherein the selection and preferred range of the substituents are the same as those of the "alkyl" substituents.

[0118] The term "alkynyl" refers to a group formed when a hydrocarbon consisting of only two elements, C and H, and containing one or more carbon-carbon triple bonds but no aromatic bonds, loses a hydrogen atom from any carbon atom. The "alkynyl" described herein includes chain alkynyls such as straight chain alkynyls and branched chain alkynyls; and also includes cycloalkynyls such as monocyclic alkynyls, spirocyclic alkynyls, fused cyclic alkynyls and bridged cyclic alkynyls. The alkynyl group may optionally contain one or more carbon-carbon double bonds. The alkynyl group may be unsubstituted or substituted.

[0119] The “alkynyl” described herein includes optionally substituted alkynyl groups, preferably having 2 to 20 carbon atoms, more preferably having 2 to 12 carbon atoms, and most preferably having 2 to 6 carbon atoms; for example, ethynyl, propynyl, 1-butynyl, 2-butynyl, isobutynyl, 1-pentynyl, 2-pentynyl, isopentenyl, 3-methyl-3-butynyl, 1-hexynyl, 2-hexynyl, 3-hexynyl, 3-heptynyl, 1-octynyl, 1-nonynyl, 1-decynyl, 1-undecanynyl, 1-dodecynyl, and the like.

[0120] The term "alkynyl" described herein also includes optionally substituted cycloalkynyl (eg, C8-C18 cycloalkynyl), for example, cyclooctynyl and the like.

[0121] The "alkynyl" described herein is optionally substituted by one or more (eg, 1 to 3) substituents, wherein the selection and preferred range of the substituents are the same as those of the "alkyl" substituents.

[0122] The term "heterocyclyl" refers to a monocyclic or polycyclic compound substituent that is saturated or contains a carbon-carbon double bond or a carbon-carbon triple bond; it contains 3 to 20 ring atoms (preferably 3 to 12 ring atoms, more preferably 3 to 8 ring atoms), wherein one or more ring atoms are selected from heteroatoms, and the remaining ring atoms are carbon; any of the rings are non-aromatic. The "heterocyclyl" described herein also includes spiro heterocyclyl, fused heterocyclyl and bridged heterocyclyl. The heterocyclyl group may be unsubstituted or substituted. The heterocyclyl group may be a heterocycloalkyl, heterocycloalkenyl or heterocycloalkynyl. Examples of suitable monocyclic heterocycles include, but are not limited to, piperidinyl, pyrrolidinyl, piperazinyl, azetidinyl, aziridine, morpholinyl, thietidinyl, oxolanyl (tetrahydrofuranyl), oxahexyl (tetrahydropyranyl), etc.

[0123] It should be understood that the "heterocyclyl" described herein is optionally substituted by one or more (eg, 1-3) substituents, wherein the selection and preferred range of the substituents are the same as those of the "alkyl" substituents.

[0124] The term "aryl" refers to a 6- to 14-membered all-carbon monocyclic or fused polycyclic group having a conjugated π electron system. The aryl ring may be fused with a heteroaromatic ring, a heterocyclic ring, a cycloalkane, a spirocycloalkane, a fused cycloalkane, a bridged cycloalkane, a cycloalkene, a spirocycloalkene, a fused cycloalkene, a bridged cycloalkene, a cycloalkyne, a spirocycloalkyne, a fused cycloalkyne or a bridged cycloalkyne. The aryl group may be unsubstituted or substituted. Examples include, but are not limited to, phenyl, naphthyl, 2-methylphenyl, 3-methylphenyl, 4-methylphenyl, methoxyphenyl (e.g., 2-methoxyphenyl, 3-methoxyphenyl, 4-methoxyphenyl), chlorophenyl (e.g., 2-chlorophenyl, 3-chlorophenyl, 4-chlorophenyl), fluorophenyl (e.g., 2-fluorophenyl, 3-fluorophenyl, 4-fluorophenyl), bromophenyl (e.g., 2-bromophenyl, 3-bromophenyl, 4-bromophenyl), 2-chloro-3-methylphenyl, 2-chloro-4-methylphenyl, 2-chloro-5-methylphenyl, 3-chloro-2-methylphenyl, 3-chloro-4-methylphenyl, 4-chloro-2-methylphenyl, 4-chloro-3-methylphenyl, 5-chloro-2-methylphenyl, 2,3-dichlorophenyl, 2,5-dichlorophenyl, 3,4-dichlorophenyl, 2,3-dimethylphenyl, 3,4-dimethylphenyl, and the like.

[0125] The "aryl" described herein is optionally substituted by 1 to 4 or 1 to 3 substituents, wherein the selection and preferred range of the substituents are the same as those of the "alkyl" substituents.

[0126] The term "heteroaryl" refers to an aromatic system containing 5 to 18 ring atoms (preferably 5 to 14 ring atoms) and wherein 1 to 4 ring atoms are heteroatoms selected from oxygen, nitrogen and sulfur. The heteroaryl itself can be fused with an aromatic ring, a heterocycle, a cycloalkane, a spirocycloalkane, a condensed cycloalkane, a bridged cycloalkane, a cycloalkene, a spirocycloalkene, a condensed cycloalkene, a bridged cycloalkene, a cycloalkyne, a spirocycloalkyne, a condensed cycloalkyne or a bridged cycloalkyne. It can be unsubstituted or substituted. Examples of heteroaryl include, but are not limited to, thienyl, furanyl, pyrrolyl, pyridyl, pyrimidinyl, imidazolyl, pyrazinyl, oxazolyl, thiazolyl, benzothienyl, benzofuranyl, benzoxazolyl, benzimidazolyl, indolyl, quinolyl, isoquinolyl and quinazolinyl etc.

[0127] The "heteroaryl" described herein is optionally substituted by 1-4 or 1-3 substituents, wherein the selection and preferred range of the substituents are the same as those of the "alkyl" substituents.

[0128] The term "acyl" as used herein refers to RC(=O)-, wherein R is C1-C18 (preferably C1-C12, more preferably C1-C6) alkyl. Examples of "acyl" include, but are not limited to, formyl, acetyl, benzoyl, nicotinoyl, propionyl, isobutyryl, oxalyl, and the like.

[0129] The acyl group RC(=O)- is optionally substituted by one or more (eg, 1 to 3) substituents, wherein the selection and preferred range of the substituents are the same as those of the "alkyl" group.

[0130] The term "cyclization" as used herein refers to the formation of a cyclic structure such as a cycloalkane ring, a cycloalkene ring, a cycloalkyne ring, an aromatic ring, a heterocycloalkane ring, a heterocycloalkene ring, a heterocycloalkyne ring, a heteroaromatic ring, etc., wherein the cyclic structure may be a monocyclic, bicyclic or polycyclic structure, including fused rings, bridged rings, and spirocyclic structures thereof. In particular, the substituent R herein 1 and R 2 The formed ring is preferably a 3- to 12-membered ring, particularly preferably a 3- to 12-membered cycloalkane ring, cycloalkene ring, heterocycloalkane ring, heterocycloalkene ring, and most preferably a 3- to 8-membered cycloalkane ring, cycloalkene ring, heterocycloalkane ring, and heterocycloalkene ring, such as a cyclopropane ring, a cyclobutane ring, a cyclopentane ring, a cyclohexane ring, a cycloheptane ring, a tetrahydrofuran ring, a tetrahydropyran ring, etc. The ring structure is optionally substituted with one or more (e.g., 1-3) substituents, wherein the selection and preferred range of the substituents are the same as those of the substituents of "alkyl".

[0131] In this article, the numerical ranges related to the number of substituents, the number of carbon atoms, and the number of ring atoms represent a list of all integers within the range, and the range is only used as a simplified representation. For example:

[0132] "1 to 4 substituents" means 1, 2, 3 or 4 substituents;

[0133] "1 to 3 substituents" means 1, 2 or 3 substituents;

[0134] "3 to 12 membered ring" means a 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12 membered ring;

[0135] "3 to 8 membered ring" means a 3, 4, 5, 6, 7 or 8 membered ring;

[0136] “1-12 carbon atoms” or “C1-C12” means 1 (C1), 2 (C2), 3 (C3), 4 (C4), 5 (C5), 6 (C6), 7 (C7), 8 (C8), 9 (C9), 10 (C10), 11 (C11) or 12 carbon atoms (C12);

[0137] “1-6 carbon atoms” or “C1-C6” means 1 (C1), 2 (C2), 3 (C3), 4 (C4), 5 (C5) or 6 carbon atoms (C6);

[0138] “2-6 carbon atoms” or “C2-C6” means 2 (C2), 3 (C3), 4 (C4), 5 (C5) or 6 carbon atoms (C6);

[0139] "C3-C8" means 3 (C3), 4 (C4), 5 (C5), 6 (C6), 7 (C7) or 8 carbon atoms (C8);

[0140] "3 to 8 ring atoms" means 3, 4, 5, 6, 7 or 8 ring atoms.

[0141] Therefore, numerical ranges related to the number of substituents, the number of carbon atoms, and the number of ring atoms also include any subranges thereof, and each subrange is also deemed to be disclosed herein.

[0142] In some particularly preferred embodiments of the present application, the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) is a compound disclosed in WO2018205928 (6'-hydroxy-8'-oxo-3'-phenyl-8'H-spiro[cyclopentane-1,5'-indolizine]-7'-carbonyl)glycine or a pharmaceutically acceptable salt thereof. The compound (6'-hydroxy-8'-oxo-3'-phenyl-8'H-spiro[cyclopentane-1,5'-indolizine]-7'-carbonyl)glycine is wherein R 1 and R 2 Together they form a cyclopentane ring (i.e. -CH2CH2CH2CH2-), R 3 is phenyl, R 4 and R 5 Hydrogen, R 6 and R 6’ is hydrogen, R 7 is hydrogen, R 8 is a compound of formula I wherein R is hydrogen and X is an oxygen atom.

[0143] 2. Drug Use or Treatment Method

[0144] The first aspect of the present application provides the use of a hypoxia inducible factor-proline hydroxylase inhibitor (HIF-PHI) in the manufacture of a medicament for treating rare anemias.

[0145] The second aspect of the present application provides the use of a hypoxia inducible factor-proline hydroxylase inhibitor (HIF-PHI) in the manufacture of a medicament for increasing the red blood cell count, hemoglobin content and / or hematocrit of rare anemias.

[0146] The third aspect of the present application provides a method for treating rare anemias, comprising: administering a therapeutically effective amount of a hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) to a patient in need of treatment.

[0147] The fourth aspect of the present application provides a method for increasing the red blood cell count, hemoglobin content and / or hematocrit of rare anemia, comprising: administering a therapeutically effective amount of a hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) to a patient in need of treatment.

[0148] The fifth aspect of the present application provides a hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) for treating rare anemias.

[0149] The sixth aspect of the present application provides the use of hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) for treating rare anemias.

[0150] Herein, the term "patient" means all mammals, and includes humans. Examples of patients include humans, cows, dogs, cats, goats, sheep, mice, pigs, rabbits, rats or mice.

[0151] Herein, the rare anemia may be selected from myelodysplastic syndrome anemia (MDS anemia), beta-thalassemia (β-thalassemia) or sickle cell disease (sickle cell anemia or SCD anemia).

[0152] The hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) of the present application can also be used to treat anemia complications or ischemic diseases, such as ischemic cerebrovascular disease (ischemic cerebrovascular disease), ischemic renal disease (ischemic renal disease, IRD), ischemic cardiomyopathy (ischemic cardiomyopathy, ICM), etc.

[0153] When using the hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) of the present application to treat rare anemias, it is usually necessary to administer a therapeutically effective amount of the active ingredient (i.e., HIF-PHI) to the patient. Herein, the term "therapeutically effective amount" is an amount that can effectively delay or eliminate the patient's symptoms or improve the patient's health status when the active ingredient mentioned in the present application is administered to the patient. The specific application dosage can be determined by the doctor according to the patient's specific circumstances. The precise dosage to be used depends not only on the route of administration, the disease, the severity of the disease to be treated, and various physical factors related to the individual being treated, but can also be determined according to the judgment of the health care practitioner. In vitro or in vivo tests can be used to help determine the optimal dosage range.

[0154] For example, the compound of formula I of the present application can be administered to a patient at a dosage of about 0.01-4000 mg, or 0.05-2000 mg, or 0.1-1000 mg, or 0.1-500 mg, or 5-500 mg, 5-300 mg, 5-200 mg, 5-100 mg, 10-30 mg, 5-15 mg, 5-10 mg, 10-15 mg, 30-50 mg per day. However, the specific dosage used can vary. It is known to those skilled in the art how to determine the optimal dosage for a particular patient.

[0155] For therapeutic use, HIF-PHI is usually administered to a patient in the form of a pharmaceutical composition comprising at least one of the above compounds as an active ingredient, optionally together with a pharmaceutically acceptable adjuvant and / or excipient and a pharmaceutically acceptable solid or liquid carrier.

[0156] The pharmaceutical composition of the present application can be formulated into various pharmaceutical dosage forms suitable for oral administration as required. Therefore, if a solid carrier is used, the preparation can be tableted, placed in a hard gel capsule in the form of powder or particles, or in the form of lozenges or lozenges. Solid carriers can include conventional excipients, such as adhesives, fillers, tableting lubricants, disintegrants, wetting agents, etc. If necessary, the tablet can be coated with conventional technology films. If a liquid carrier is used, the preparation can be in the form of syrup, emulsion, soft gel capsule, aqueous or non-aqueous liquid suspension, or can be a dry product reconstituted with water or other suitable carriers before use. Liquid preparations can include conventional additives, such as suspending agents, emulsifiers, wetting agents, non-aqueous carriers (including edible oils), preservatives, and flavoring agents and / or coloring agents.

[0157] These pharmaceutical compositions (or pharmaceutical preparations) may also include various excipients, such as preservatives, wetting agents, emulsifiers and dispersants. The inhibition of the effects of microorganisms may be ensured by various antibacterial and antifungal agents (e.g., parabens, chlorobutanol, phenol, sorbic acid, etc.). Isotonic agents may also be included, such as sugar, sodium chloride, etc. The absorption of injectable pharmaceutical dosage forms may be extended by using delayed absorption agents (e.g., aluminum monostearate and gel).

[0158] Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules. In such solid dosage forms, the active compound is mixed with at least one inert excipient (or carrier) (e.g., sodium citrate or dicalcium phosphate), which may also include: (a) fillers or admixtures (e.g., starch, lactose, sucrose, glucose, mannitol and silicic acid); (b) binders (e.g., carboxymethylcellulose, alginate, gelatin, polyvinyl pyrrolidone, sucrose and gum arabic); (c) humectants (e.g., glycerol); (d) disintegrants (e.g., agar); The invention can be prepared by mixing (a) agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain synthetic silicates, sodium carbonate); (e) solution retarder (e.g., paraffin); (f) absorption promoter (e.g., quaternary ammonium compounds); (g) wetting agents (e.g., cetyl alcohol and glycerol monostearate); (h) adsorbents (e.g., kaolin and bentonite) and (i) lubricants (e.g., talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate) or mixtures thereof.

[0159] Solid pharmaceutical compositions (or pharmaceutical formulations) of a similar type may also be used as fillers in soft and hard-filled gelatin capsules using excipients such as lactose and high molecular weight polyethylene glycols.

[0160] Solid dosage forms (e.g., tablets, dragees, capsules, pills and granules) can be prepared with coatings and shells (e.g., enteric coatings and others known in the art). They may contain opacifiers, and they may also be pharmaceutical compositions (or pharmaceutical preparations) that release the active compound or compounds in a delayed manner in a certain part of the intestinal tract. The active ingredient may also be in microencapsulated form, if appropriate, with one or more of the above-mentioned excipients.

[0161] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, dispersions, syrups and elixirs. In addition to the active compound, the liquid dosage form may contain inert diluents (e.g., water or other solvents), solubilizers and emulsifiers (e.g., ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide ammonium), oils (specifically, cottonseed oil, peanut oil, corn oil, olive oil, castor oil, sesame oil), glycerol, tetrahydrofuran alcohol, polyethylene glycol and fatty acid esters of sorbitan or mixtures of these substances, etc., commonly used in the art.

[0162] Besides these inert diluents, the pharmaceutical composition (or pharmaceutical preparation) may also include, for example, wetting agents, emulsifying and suspending agents, perfuming agents, flavoring agents and perfuming agents.

[0163] In addition to the active compounds, suspensions may contain suspending agents such as ethoxylated isostearyl alcohols, polyoxyethylene sorbitol, sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar and tragacanth, or mixtures of these substances, and the like.

[0164] The amount of the HIF-PHI compound in the pharmaceutical composition (or pharmaceutical preparation) can be appropriately determined by those skilled in the art according to needs.

[0165] In some embodiments, the amount of HIF-PHI or a pharmaceutically acceptable salt thereof contained in each unit of the pharmaceutical preparation is about 1-1000 mg, about 1-10 mg, about 10 mg, about 5 mg, about 4.5 mg, about 1-3 mg, about 2-4 mg, about 3-5 mg, about 4-6 mg, about 5-7 mg, about 6-8 mg, about 7-9 mg, about 8-10 mg, about 9-11 mg, about 10-12 mg, about 4.5-5 mg, 20 mg, 30 mg, 30 -100 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg, about 80 mg, about 90 mg, about 10-30 mg, about 30-50 mg, about 50-70 mg, about 10-90 mg, about 10-800 mg, about 10-700 mg, about 10-600 mg, about 10-500 mg, about 10-400 mg, about 10-300 mg, or about 10-200 mg, or an amount within any range defined by any of these values.

[0166] In some embodiments, the amount of HIF-PHI or a pharmaceutically acceptable salt thereof contained in each unit of the pharmaceutical preparation is about 0.01-0.2 mg, about 0.2-0.4 mg, about 0.4-0.6 mg, about 0.6-0.8 mg, about 0.8-1 mg, about 1-1.2 mg, about 1.2-1.4 mg, about 1.4-1.6 mg, about 1.6-1.8 mg, about 1.8-2 mg, about 2-2.2 mg, about 2.2-2.4 mg, about 2.4-2.6 mg, about 2.6-2.8 mg, about 2.8-3 mg, about 3-3.2 mg, about 3.2-3.4 mg, about 3.4-3.6 mg, about 3.6-3.8 mg, about 3.8-4 mg, about 3.4-3.6 mg, about 3.8-3.8 mg, about 3.8-4 mg. 9-4.1mg, about 4-4.2mg, about 0.2-0.4mg, about 0.2-0.6mg, about 0.2-0.8mg, about 0.2-1mg, about 0.2-1.2mg, about 0.2-1.4mg, about 0.2-1.6mg, about 0.2-1.8mg, about 0.2-2.0mg, 0.2-2.5mg, about 0.2-3.0mg, about 0.2-3.5mg, about 0.2-4.0mg, about 5-10mg, about 10-15mg, about 15-20mg, about 20-25mg, about 25-30mg, about 30-40mg, about 40-50mg, about 50-60mg, about 60-70mg, about 70-80mg, about 80- 90mg, about 90-100mg, about 100-120mg, about 120-140mg, about 140-150mg, about 150-160mg, about 160-180mg, about 180-200mg, about 200-220mg, about 220-240, about 10-500mg, about 50-400mg, about 50-300mg, about 100-250mg, about 1-10mg, about 10-200mg, about 10-150mg, about 10-100mg, about 10-180mg, about 10-160mg, about 10-140mg, about 10-120mg, about 10-100mg, about 10-20mg, about 20-30mg, about 3 In some embodiments, the present invention relates to an amount of at least one embodiment of the present invention, wherein the amount of the at least one embodiment of the present invention is at least one embodiment of the present invention. The amount of the at least one embodiment of the present invention can be 0.0 to 40 mg, about 40 to 50 mg, about 50 to 60 mg, about 60 to 70 mg, about 70 to 80 mg, about 80 to 90 mg, about 90 to 100 mg, about 100 to 120 mg, about 120 to 140 mg, about 140 to 160 mg, about 160 to 180 mg, about 180 to 200 mg, about 200 to 220 mg, about 220 to 240, about 240 to 250 mg, about 250 to 260 mg, about 260 to 280 mg, about 280 to 300 mg, about 300 to 350 mg, about 350 to 400 mg, about 25 mg, about 50 mg, about 100 mg, about 250 mg, or an amount within any range defined by any of these values.

[0167] In some embodiments, the amount of HIF-PHI or a pharmaceutically acceptable salt thereof contained in each unit of the pharmaceutical preparation is about 0.1% (w / w) to about 10% (w / w), about 0.05% (w / w) to about 5% (w / w), about 0.2% (w / w) to about 15% (w / w), about 0.5% (w / w) to about 30% (w / w), about 1% (w / w) to about 25% (w / w), about 2% (w / w), about 3% (w / w), about 4% (w / w), about 5% (w / w), at least about 10% (w / w), at least about 20% (w / w), at least about 50% (w / w), at least about 70% (w / w). % (w / w), at least about 80%, about 10% (w / w) to about 30% (w / w), about 10% (w / w) to about 20% (w / w), about 20% (w / w) to about 30% (w / w), about 30% (w / w) to about 50% (w / w), about 30% (w / w) to about 40% (w / w), about 40% (w / w) to about 50% (w / w), about 50% (w / w) to about 80% (w / w), about 50% (w / w) to about 60% (w / w), about 70% (w / w) to about 80% (w / w), or about 80% (w / w) to about 90% (w / w), or a ratio within any range defined by any of these ratios.

[0168] The preparation of the pharmaceutical composition (or pharmaceutical preparation) can adopt methods known or commonly used in the art, which generally include the process of mixing the active ingredient HIF-PHI with a pharmaceutically acceptable carrier, adjuvant or excipient, and optional post-treatment or processing steps (such as drying, granulation, encapsulation, etc.).

[0169] Example

[0170] The embodiments described below are part of the embodiments of the present invention, rather than all of the embodiments. The detailed description of the embodiments of the present invention is not intended to limit the scope of the invention claimed for protection, but merely represents selected embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without departing from the principles of the present invention and without making creative work are within the scope of protection of the present invention.

[0171] For the sake of simplicity, some materials, equipment and method steps commonly used in the art are not specified in the examples. All process methods and analytical test processes (and related parameters) not specifically specified in the examples are carried out according to those commonly used by those skilled in the art; materials, reagents and equipment without specific sources are conventional laboratory materials and equipment and can be obtained through commercial channels.

[0172] Example 1

[0173] (6'-Hydroxy-8'-oxo-3'-phenyl-8'H-spiro[cyclopentane-1,5'-indolizine]-7'-carbonyl)glycine ( 1 )

[0174]

[0175] The compound was synthesized according to the method of Example 24 in WO2018205928 1 .

[0176] The structures of the compounds were determined by liquid chromatography-mass spectrometry (LCMS) or nuclear magnetic resonance (NMR). NMR chemical shifts (δ) were expressed in 10 -6 The unit of (ppm) is used for NMR measurement. Bruker-500 NMR instrument is used, and the measurement solvent is deuterated dimethyl sulfoxide (DMSO-D6), deuterated chloroform (CDCl3), etc., and the internal standard is tetramethylsilane (TMS). LCMS is measured by Shimadzu LCMS-2020 or Thermo UltiMate3000.

[0177] The thin layer chromatography silica gel plate uses Shandong Yantai Huanghai HSGF254 or Shandong Qingdao GF254 silica gel plate. Column chromatography generally uses Shandong Yantai Huanghai silica gel 200 to 300 mesh silica gel as a carrier.

[0178] All starting materials used in this example were purchased from chemical suppliers or synthesized by literature methods.

[0179] The abbreviations that may be used in this embodiment are as follows:

[0180] DMSO-d6: dimethyl sulfoxide with all six hydrogen atoms replaced by deuterium

[0181] CDCl3: deuterated chloroform

[0182] CAS: Chemical Abstracts Registration Number

[0183] NMR: Nuclear Magnetic Resonance

[0184] LCMS: Liquid chromatography-mass spectrometry

[0185] ESI: Electrospray ionization

[0186] ppm: parts per million

[0187] δ: NMR chemical shift

[0188] TMS: Tetramethylsilane

[0189] s: NMR singlet

[0190] d: NMR doublet

[0191] t: NMR triplet

[0192] br: NMR broad peak

[0193] CDI: Carbonyldiimidazole

[0194] DCC: N,N'-dicyclohexylcarbodiimide

[0195] NBS: N-bromocyclosuccinimide

[0196] Compound 1 The main synthetic steps are as follows.

[0197] first step

[0198] 1-(2-phenyl-1H-pyrrol-1-yl)cyclopentane-1-carboxylic acid ( 1a )

[0199]

[0200] Compound 3-(1,3-dioxacyclohexane-2-yl)-1-phenylpropane-1-one (prepared according to the method described in patent document WO / 2011 / 042477) (500 mg) and 1-amino-cyclopentane-1-carboxylic acid (421 mg) were refluxed in 8 ml of acetic acid for 16 hours. Then, after the reaction was cooled, the acetic acid was dried as much as possible; water and ethyl acetate were added to dilute; the organic phase was collected, and the organic phase was washed several times with a dilute sodium chloride aqueous solution until the aqueous phase was close to neutral; then the ethyl acetate layer was dried with anhydrous sodium sulfate, filtered, concentrated, and purified by column chromatography to obtain the compound 1a LCMS ESI(+):256(M+1) + . 1 H NMR(500MHz,DMSO-d6)δ(ppm):12.95(br,s,1H),7.35-7.33(m,3H),7.25-7.22(m,2H),7.00-6 .99(m,1H),6.04(m,1H),5.95(m,1H),2.14-2.11(m,2H),2.01-1.97(m,2H),1.60-1.57(m,4H).

[0201] Step 2

[0202] 2-(1-(2-phenyl-1H-pyrrol-1-yl)cyclopentane-1-carbonyl)malonate ( 1b )

[0203]

[0204] The second step compound in Example 15 of WO2018205928 is used 15b The synthetic route is 15a Replace with 1a , you can get 1b LCMS ESI(+):370(M+1) + . 1 H NMR (500MHz, DMSO-d6) δ7.28–7.22(m,3H),7.20–7.15(m,2H),7.11(dd,J=3.2,1.8Hz,1H),6.15(t,J=3.3Hz,1H),5.93(d d,J=3.5,1.7Hz,1H),5.13(s,1H),3.56(s,6H),2.06(d,J=8.0Hz,2H),1.59(t,J=12.2Hz,2H),1.33(s,2H),1.16(s,2H).

[0205] Step 3

[0206] 6'-Hydroxy-8'-oxo-3'-phenyl-8'H-spiro[cyclopentane-1,5'-indolizine]-7'-carboxylic acid methyl ester

[0207] ( 1c )

[0208]

[0209] The third step compound in Example 15 of WO2018205928 is used 15c The synthetic route is 15b Replace with 1b , you can get 1c LCMS ESI(+):338(M+1) + . 1 H NMR(500MHz,DMSO-d6)δ13.84(s,1H),7.54-7.43(m,5H),7.09(d,J=4.0Hz,1H),6.35(d,J=4.0H z,1H),3.81(s,3H),2.30-2.21(m,2H),2.10-2.01(m,2H),1.48-1.38(m,2H),1.00-0.89(m,2H).

[0210] Step 4

[0211] (6'-Hydroxy-8'-oxo-3'-phenyl-8'H-spiro[cyclopentane-1,5'-indolizine]-7'-carbonyl)glycine ( 1 )

[0212]

[0213] The fourth step compound in Example 15 of WO2018205928 is used 15 The synthetic route is 15c Replace with 1c , the compound can be obtained 1 LCMS ESI(+):381(M+1) + . 1 H NMR (500MHz, DMSO-d6) δ (ppm) 17.96 (s, 1H), 12.97 (s, 1H), 9.89-9.87 (d, J = 5.5Hz, 1H), 7.53-7.46 (m, 5H), 7.08-7.07 (d, 1H) ,6.37-6.36(d,1H),4.08-4.07(d,J=5.5Hz,2H),2.35-2.20(m,2H),2.18-2.10(m,2H),1.60-1.46(m,2H),0.98-0.85(m,2H).

[0214] Example 2

[0215] Inhibitory effects of compounds on HIF PHD-2

[0216] In this example and the following examples, the compound prepared in Example 1 was used. 1 Tests are performed and compounds are referred to as "Comp-1" 1 .

[0217] Prepare the following stock solutions: HEPES buffer (pH 7.4, 50 mM), DMSO / HEPES solution of HIF-1α peptide (sequence: DLDLEMLAPYIPMDDDFQL; Genscript) (0.5 mM; preparation method: first prepare a 20 mM DMSO solution of the peptide, and then dilute to 40 times the volume with the aforementioned HEPES buffer), ascorbic acid solution (10 mM), FeSO4 hydrochloric acid solution (0.1 mM, hydrochloric acid concentration 10 μM), 2-oxoglutaric acid solution (2-OG: 1 mM), EDTA solution (pH 8.0; 0.5M), 8 concentrations of Comp-1 / DMSO solution (concentrations are 4000μM, 1333μM, 444.4μM, 148.1μM, 49.4μM, 16.5μM, 5.5μM and 1.82μM). The above concentrations of reagents were added to 9 Eppendorf tubes: 50μL HEPES solution, 10μL ascorbic acid solution, 10μL FeSO4 hydrochloric acid solution, 10μL 2-OG solution, 10μL HIF-1α peptide solution; then 5μL of the above 8 concentrations of Comp-1 / DMSO solution and 5μL DMSO were added respectively; finally, 5μL PHD2 enzyme (Active Motif; concentration: 0.2μg / μL) was added to each tube; the reaction system was incubated at 30℃ in the dark for 2 hours; finally, 10μL EDTA solution was added to each tube to terminate the reaction. The liquid of each reaction system was filtered with a 0.45 μm filter, and then HIF-peptide and hydroxylated HIF-peptide were separated by high performance liquid chromatography. The concentrations of HIF-peptide and hydroxylated HIF-peptide were quantified by ultraviolet absorption detector, and their ratio was calculated. IC was calculated by Graph Prism 6.0 software. 50 ; IC of Comp-1 to HIF PHD-2 50 It is 2.36μM.

[0218] Example 3

[0219] Effects of HIF-PH inhibitor and EPO on MDS model mice

[0220] During the entire experimental period, the animals were kept in animal cages with 24-hour continuous central air conditioning, temperature set at 20-26 degrees Celsius, relative air humidity set at 40-70%, and 12-hour light and dark alternation. All animals were free to eat and drink sterilized drinking water. The C57 mice used in the experiment contain three knock-in genes in the genome: Vav1-Cre, LSL and c-Myc (T58A / S62E). Animals with positive all three genes can be used for the experiment. Vav1-Cre expression can induce LSL site cutting, thereby expressing the tTA gene. After adding Doxycycline induction, tTA can bind to the promoter of the c-Myc (T58A / S62E) gene, thereby driving the expression of the c-Myc (T58A / S62E) gene, and the animals show MDS phenotype. When the mice were about 8 weeks old, the model was established. The mice were fed with special feed (1 kg of feed contained 200 mg of doxycycline) for 4 weeks. On days 0, 14, and 28, the mice were anesthetized and 70 μl of blood was collected from the eyes. The contents of RBC, Hb, and HCT in the blood were detected to confirm that the MDS model was successfully established. Then, the mice were randomly divided into 3 groups, 10 in each group: vehicle control group (0.1 M meglumine), low-dose Comp-1 group (10 mg / kg), and high-dose Comp-1 group (30 mg / kg); the volume of drug administration or solvent was 6.67 mL / kg; the frequency of drug administration or solvent administration in each group was oral gavage three times a week (TIW), and the drug administration lasted for 4 weeks, that is, the 4th to 8th week was the drug administration treatment period. On the 14th day (the 6th week of the experiment) and the 28th day (the 8th week of the experiment) of drug administration, the mice were anesthetized and 70 μl of blood was collected from the eyes. The collected blood samples were placed in heparin-anticoagulant tubes to detect the content of RBC, Hb, and HCT in the blood. After the last blood sampling of each animal, the animal was euthanized and the corresponding organ tissues were collected and stored for later use. Detection instrument: Element HT5 Veterinary Hematology Analyzer; Supplier: HESKA. Graphpad prism 9.0 was used for data analysis.

[0221] Experimental results:

[0222] (1) Effect on RBC: After oral administration of Comp-1, the RBC in the blood increased significantly. After 2 weeks of administration, the RBC values ​​of 30 mg / kg and 10 mg / kg were significantly higher than those of the vehicle group (7.11M / μL vs 6.33M / μL vs 5.57M / μL). After 4 weeks of administration, the RBC in the treatment group continued to increase (8.46M / μL vs 7.17M / μL vs 5.27M / μL), and the RBC of the 30 mg / kg group animals returned to a level close to that before modeling (8.46M / μL vs 9.01M / μL). See the table below:

[0223]

[0224] (2) Effect on Hb: After oral administration of Comp-1, the Hb of MDS mice increased significantly. After 2 weeks of administration, the Hb values ​​of 30 mg / kg and 10 mg / kg were significantly higher than those of the vehicle group (13.56 g / dL vs 10.97 g / dL vs 8.72 g / dL). After 4 weeks of administration, the Hb of the treatment group continued to increase (14.91 g / dL vs 11.77 g / dL vs 8.19 g / dL), and the Hb of the 30 mg / kg group animals returned to the pre-modeling level (14.91 g / dL vs 14.37 g / dL). See the table below:

[0225]

[0226] (3) Effect on HCT hematocrit: After oral gavage of Comp-1, the HCT of MDS mice increased significantly. After 2 weeks of administration, the HCT values ​​of 30 mg / kg and 10 mg / kg were significantly higher than those of the vehicle group (43.13% vs 34.36% vs 27.69%). After 4 weeks of administration, the HCT of the treatment group continued to rise (47.41% vs 37.52% vs 27.13%), and the HCT of the animals in the 30 mg / kg group recovered and was even slightly higher than the level before modeling (47.41% vs 43.32%). See the table below:

[0227]

[0228] It can be seen that the test substance Comp-1 at 10 and 30 mg / kg can effectively increase the RBC, Hb, and HCT values ​​in the blood of MDS mice, and significantly improve the anemia symptoms of MDS mice.

[0229] As a control, the same experiment was repeated using Roxadustat (30 mpk, oral gavage, TIW / three times a week) and EPO (50 U / 30 g). Neither Roxadustat nor EPO could improve the RBC, Hb, and HCT values ​​of this MDS mouse model.

[0230] Example 4

[0231] Effects of Comp-1 on homozygous β-thalassemia mice

[0232] During the entire experimental period, the animals were kept in animal cages with 24-hour continuous central air conditioning, temperature set at 20-26 degrees Celsius, relative humidity set at 40-70%, and 12-hour light and dark alternation. All animals were free to eat (radiation sterilized feed) and drink water (prepared by ultrapure water). The mouse strain used in the experiment was: Hbbd 3th / d3th Mice; female and male, 9-15 weeks old, weighing 25-30 grams. One male and two females were used for breeding, and the male mice were selected with the genotype Hbb. d3th / d3th The female mice were selected with the genotype Hbb d3th / + Mice. When the offspring mice were one week old, the toes were cut to extract tissue DNA, which was subsequently amplified by PCR and separated by agarose gel electrophoresis for the purpose of gene identification. Before the experiment, a secondary genotyping of mice of appropriate age was performed, and mice with no obvious differences in weight and appearance were selected as experimental mice. Solvent 1 (0.1M meglumine aqueous solution): An appropriate amount of meglumine (0.586g) was dissolved in an appropriate amount of deionized water (30mL) and stirred to prepare the solvent. After the solvent is prepared, it is divided into daily doses and stored at 5±3°C. Stir at room temperature for at least 10 minutes before administration and continue stirring during administration.

[0233] Comp-1: Weigh the required amount of Comp-1, dissolve it completely in an appropriate volume of solvent 1, and stir and sonicate to make a solution. After the dissolution is observed by the naked eye, gradually add the solvent until the specified volume. Before daily administration, take the preparation (including the solvent) out of the refrigerator and allow it to reach room temperature. The preparation is stirred at room temperature for at least 10 minutes before administration and stirred throughout the administration process. Information such as animal grouping, dosage, and experimental time length is shown in the table below. The route of administration is oral gavage, the dosage volume is 10mL / kg, and the frequency of administration is daily for a total of 42 days.

[0234] Group Number of animals Animal Gender drug Time of administration Dosage, mg / kg G1 2+3 female + male solvent 1 Day 1-42 0 G2 2+3 female + male Comp-1 Day 1-42 5 G3 2+3 female + male Comp-1 Day 1-42 10 G4 2+3 female + male Comp-1 Day 1-42 15

[0235] CBC test

[0236] On days 0, 15, 29, and 43 of administration, 50 μl of blood was collected from the mice through the eye sockets, stored at 4°C, and restored to room temperature before testing. Then, the blood was gently mixed and added to the prepared diluent (blood: diluent = 1:6); the pre-dilution mode of the instrument was used for testing to obtain data. The statistics of the data were processed and analyzed by the software Microsoft Office Excel (Microsoft Corporation, version number 2013, Redmond, WA, USA) and Graphpad prism 7.0. The red blood cell count and proportion were reported in the form of two significant figures. The blood cell count and proportion between groups and at different time points within the group were analyzed using T-test.

[0237] Experimental results:

[0238] (1) Effect on RBC count: The RBC count gradually increased after drug administration, and the 15mpk (mpk: mg / kg) drug administration group reached the highest point on the 29th day, and then stabilized. The hematocrit and hemoglobin concentration stabilized after the 29th day, and the drug administration group was higher than the control group, with an obvious dose-response relationship. Compared with before drug administration, the RBC counts of the 10 and 15mpk groups gradually increased at D15, D29, and D43, and were significantly dose-dependent. Although there was no significant difference between the 5mpk group and the group before drug administration at each time point after drug administration, the RBC count showed a slow upward trend; the RBC counts of the 10 and 15mpk groups increased significantly at D15, D29, and D43 compared with before drug administration, and the increase in the RBC count of the 15mpk group was more significant. See the table below:

[0239]

[0240] (2) Effect on HGB content: The HGB content of each medication group increased with the increase of medication time. Compared with the vehicle group, the 15mpk group had significant statistical significance at D29 and D43 (*P<0.05; ***P<0.001). The HGB content of the 10 and 15mpk groups at D15, D29, and D43 increased significantly, and showed obvious dose dependence, which was statistically significant compared with the pre-administration period; the HGB content of each group at each time point after medication was compared with that before medication, #, P<0.05; ##, P<0.01; ###, P<0.001; ####, P<0.0001. See the table below:

[0241]

[0242]

[0243] (3) Effect on HCT hematocrit: 5, 10, and 15 mpk of Comp-1 can increase the hematocrit HCT percentage. With the increase of administration time, HCT gradually increased, and there was a certain degree of dose dependence. Compared with the vehicle group, at the end point of the experiment D43, the three dose groups were statistically significant (low dose group P < 0.05, medium dose group P < 0.01, high dose group P < 0.01). The effect of the high dose group on increasing HCT at D43 was significantly stronger than that of the medium and low dose groups (P < 0.05, P < 0.01). After administration, the HCT% of the low, medium, and high dose groups increased significantly compared with their respective pre-administration results, and there was a clear dose dependence (#, P < 0.05; ##, P < 0.01; ###, P < 0.001; ####, P < 0.0001). See the table below:

[0244]

[0245] It can be seen that Comp-1 can significantly increase the number of red blood cells, hemoglobin content, and hematocrit in thalassemia mice, indicating that Comp-1 has a therapeutic effect on anemia in homozygous d3th thalassemia genotype mice.

[0246] Example 5

[0247] Effects of HIF PH inhibitor and EPO on SCD anemia mice

[0248] During the entire experimental period, the animals were kept in animal cages with 24-hour continuous central air conditioning, temperature set at 20-26 degrees Celsius, relative humidity set at 40-70%, and 12-hour light and dark alternation. All animals were free to eat and drink sterilized drinking water. The animal strain was B6.Cg-Hbb tm2(HBG1,HBB*)Tow / Hba tm1(HBA)Tow / ShmtJ mice are Townse SCD model mice that have human hemoglobin knocked in to replace mouse endogenous hemoglobin. Two weeks after birth, the mice were numbered by puncturing their ears, and the punctured ear tissues were collected for PCR testing to determine the genotype of the mice. tm2(HBG1,HBB*)Tow / Hba tm1(HBA)Tow / ShmtJ mice were grouped. When the mice were about 4 weeks old, blood was collected from the eyes to detect RBC, Hb and HCT values ​​to determine whether the animals had anemia symptoms. Blood samples were collected every other week. Mice with anemia symptoms were selected for administration. The animals were randomly divided into a vehicle control group (0.1M meglumine aqueous solution; oral gavage; volume: 6.67mL / kg), a Comp-1 group (30mpk; oral gavage; volume: 6.67mL / kg) and an EPO group (50U / 30g body weight; SC injection; MyBioSource, cat.#:MBS650094, erythropoietin alpha); the drugs were administered three times a week (TIW) for 4 consecutive weeks. 70μL of blood was collected from the eyes on days 14 and 28 of administration; the collected blood samples were placed in heparin-anticoagulant tubes. 15 μL of blood sample was drawn from the anticoagulation tube and added to the Element HT5 Veterinary Hematology Analyzer instrument. RBC, Hb and HCT in the blood were detected according to the instrument operating instructions. Graphpad prism 9.0 was used for data analysis.

[0249] Experimental results:

[0250] From the arithmetic mean values ​​of various indicators (red blood cell count, hemoglobin content and hematocrit) of mice in each group before administration in the three tables below, it can be seen that the mice in each group were highly anemic, indicating that the model was successfully established.

[0251] (1) Effect on RBC count: As can be seen from the table below, the RBC count of animals in the vehicle control group continued to decline over time, indicating that the degree of anemia was worsening. Even for the EPO positive control mice, which can be used for anemia such as CKD, their RBC count continued to decline over time and the degree of anemia was worsening. The RBC counts of the EPO group and the vehicle control group were close at the same time point, indicating that EPO could not induce an increase in RBC count. In contrast, the RBC count of the Comp-1 group (30 mg / kg) increased significantly after 2 and 4 weeks of administration compared with before administration (8.88 M / μL, 9.09 M / μL vs 6.34 M / μL).

[0252]

[0253] (2) Effect on HGB hemoglobin content: As can be seen from the table below, as time progressed, the hemoglobin content of mice in the vehicle control group continued to decline, and the degree of anemia continued to worsen. Even in the EPO positive control mice, as time progressed, the hemoglobin content continued to decline, and the degree of anemia continued to worsen. The red blood cell counts of the EPO group and the vehicle control group were close at the same time point, indicating that EPO could not induce an increase in hemoglobin content. In contrast, the hemoglobin content of the Comp-1 group (30 mg / kg) increased significantly after 2 and 4 weeks of administration (11.63 g / dL, 12.10 g / dL vs 8.47 g / dL), and was close to normal levels.

[0254]

[0255] (3) Effect on HCT hematocrit: As can be seen from the table below, with the progress of time, the hematocrit level of mice in the vehicle control group continued to decline, and the degree of anemia continued to deepen; even for mice in the EPO positive control group, with the progress of time, the hematocrit level continued to decline, and the degree of anemia continued to deepen. The hematocrit levels of the EPO group and the vehicle control group were close at the same time point, indicating that EPO could not increase the hematocrit level; while the hematocrit level of the Comp-1 group (30 mg / kg) increased significantly after 2 and 4 weeks of administration compared with before administration (40.05%, 41.45% vs 27.20%), and was close to the normal level.

[0256]

[0257] As controls, the same experiment was repeated using Roxadustat (30 mpk, 60 mkp, oral gavage, TIW / three times a week) and Daprodustat (30 mpk, oral gavage, TIW / three times a week).

[0258] At the end of the four-week experiment, both dose groups of Roxadustat failed to improve RBC, Hb, and HCT; the 2022 Nature article (Nautre 2022, vol 610, 783: https: / / www.nature.com / articles / s41586- 022-05312-w ) It was found that Roxadustat can induce fetal hemoglobin at the cellular level, indicating that Roxadustat may be used for SCD, but this experiment showed that the performance of Roxadustat in SCD model mice was completely unexpected, and Roxadustat had no therapeutic effect on SCD mice.

[0259] Daprodustat can slightly improve RBC, Hb, and HCT. At the end of the four-week experiment, RBC increased from 6.14M / μL to 7.54M / μL, Hb increased from 9.1g / dL to 10.9g / dL, and HCT increased from 31.8% to 35.9%. One of the six mice in the Daprodustat group died two weeks after administration, so the baseline data of the experiment was the average of six mice, and the data at the end of four weeks was the average of five mice. The results showed that Daprodustat had a slight effect on the SCD model mice, but was far inferior to Comp-1.

[0260] From the above effects, it can be seen that although EPO is known to be used for anemia such as CKD chronic kidney disease, it has no therapeutic effect in SCD model mice. Roxaudstat can induce fetal hemoglobin at the cellular level, but it is completely ineffective in SCD model mice. Daprodustat can slightly improve RBC, Hb and HCT, but Comp-1 shows a surprising effect.

[0261] Although the embodiments of the present application have been illustrated and described, it is not intended that these embodiments illustrate and describe all possible forms of the present application. More specifically, the words used in this specification are merely descriptive and non-restrictive, and it should be understood that various changes can be made without departing from the spirit and scope of the present application.

Claims

1. Use of a hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) in the manufacture of a medicament for treating rare anemias.

2. Use of a hypoxia-inducible factor-proline hydroxylase inhibitor (HIF-PHI) in the manufacture of a medicament for increasing red blood cell count, hemoglobin content and / or hematocrit in rare anemias.

3. A method for treating rare anemia, comprising: A therapeutically effective amount of a hypoxia inducible factor-prolamide hydroxylase inhibitor (HIF-PHI) is administered to a patient in need of treatment.

4. A method for increasing the red blood cell count, hemoglobin content and / or hematocrit of a rare anemia, comprising: A therapeutically effective amount of a hypoxia inducible factor-prolamide hydroxylase inhibitor (HIF-PHI) is administered to a patient in need of treatment.

5. The use or method according to any one of claims 1 to 4, wherein: The rare anemia is selected from myelodysplastic syndrome anemia (MDS anemia), beta-thalassemia (β-thalassemia) or sickle cell disease (sickle cell anemia or SCD anemia).

6. The use or method of any one of claims 1 to 4, wherein the hypoxia inducible factor-prolyl hydroxylase inhibitor (HIF-PHI) is Comp-1.

7. The use or method according to any one of claims 1 to 4, wherein: The HIF-PHI is a compound of formula I or a pharmaceutically acceptable salt or a tautomer thereof: in: R 1 and R 2 independently selected from cyano, alkyl, heterocyclic, alkenyl, alkynyl, aryl, heteroaryl, acyl; wherein the above alkyl, heterocyclic, alkenyl, alkynyl, aryl, heteroaryl, acyl are optionally substituted by one or more substituents, wherein the substituents are independently selected from halogen, cyano, hydroxyl, amino, carboxyl, acyl, alkyl, heterocyclic, alkenyl, alkynyl, aryl, heteroaryl, =O, =S, -SH, R 10 O-、R 10 S-, R 10 (O=)S-、R 10 (O=)2S-, where R 10 is alkyl, heterocyclyl, alkenyl, alkynyl, aryl, or heteroaryl; or R 1 and R 2 Put together to form a ring; R 3 is selected from alkyl, heterocyclyl, alkenyl, alkynyl, aryl, and heteroaryl; R 4 and R 5 is hydrogen; R 6 and R 6’ is hydrogen; R 7 is hydrogen; R 8 is selected from hydrogen and alkyl; X is an oxygen atom.

8. The use or method according to claim 7, wherein in the compound of formula I, R 1 and R 2 Together with the C atoms to which they are attached, they form a cyclopentane ring (i.e., R 1 and R 2 is -CH2CH2CH2CH2-), R 3 is phenyl, R 4 and R 5 is hydrogen, R 6 and R 6’ is hydrogen, R 7 is hydrogen, R 8 is hydrogen, and X is an oxygen atom, that is, the compound of formula I is (6'-hydroxy-8'-oxo-3'-phenyl-8'H-spiro[cyclopentane-1,5'-indolizine]-7'-carbonyl)glycine.

9. The use or method according to any one of claims 1 to 4, wherein: When the rare anemia is myelodysplastic syndrome anemia (MDS anemia), the HIF-PHI is selected from: Daprodustat, Vadadustat, Molidustat, Enarodustat, Desidustat, HIF117 (SSS17), HEC53856, DDO-3055 and FG-2216.

10. The use or method according to any one of claims 1 to 4, wherein: When the rare anemia is β-thalassemia (β-thalassemia), the HIF-PHI is selected from: roxadustat, Daprodustat, Vadadustat, Molidustat, Enarodustat, Desidustat, HIF117 (SSS17), HEC53856, DDO-3055 and FG-2216.

11. The use or method according to any one of claims 1 to 4, wherein: When the rare anemia is sickle cell disease (sickle cell anemia or SCD anemia), the HIF-PHI is selected from: Daprodustat, Vadadustat, Molidustat, Enarodustat, Desidustat, HIF117 (SSS17), HEC53856 and DDO-3055.

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