Pyridine compound as SARM1 enzyme activity inhibitor and use thereof

Abstract

Provided in the present invention are a compound of formula (I) which can be used as an SARM1 enzyme activity inhibitor and the use of the compound in the treatment of neurodegenerative diseases or neurological diseases or conditions.

Description

Pyridine compounds as SARM1 enzyme activity inhibitors and their applications Technical Field

[0001] The present application relates to compounds that can be used to inhibit SARM1 enzyme activity and the use of these compounds in treating and / or preventing neurodegenerative or neurological diseases or conditions associated with SARM1 enzyme activity. Background Art

[0002] Neurodegenerative diseases are a class of diseases that can severely harm humans, causing devastating damage, such as progressive neuronal cell death. Among the primary neurodegenerative diseases, known are central nervous system diseases such as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis (ALS), and Huntington's disease, as well as peripheral nervous system diseases such as diabetes. While most are associated with aging, the incidence of these diseases increases with age, although some also occur in middle-aged individuals or even younger.

[0003] As a result of research on brain structure and function, the effects of neurotransmitters and neurotrophic factors have been gradually elucidated, but many local causes of neurodegeneration are still unclear. Only for Parkinson's disease, the relationship between this disease and the special neurotransmitter, i.e., dopamine, has been elucidated, and the precursor of dopamine, L-dopa, has been used as a medicine to alleviate neurological symptoms and restore neurological function. However, L-dopa cannot inhibit the development of neurodegeneration, and gradually loses its effect as the disease progresses, i.e., dopamine-based neuronal cell degeneration and defect. Similarly, Alzheimer's disease is also caused by the degeneration and defect of multiple nerve cells such as acetylcholine-based neurons, monoamine-based neurons, etc., and as medicines for the treatment of this disease, cholinesterase inhibitors have been put on the market or are being developed. However, the L-dopa for the treatment of Parkinson's disease is still limited to symptomatic treatment, to temporarily improve neurological symptoms.

[0004] Therefore, there is a lack of effective therapeutic drugs for neurodegenerative diseases.

[0005] Studies have found that axonal damage occurs in a variety of neurodegenerative diseases, accidental injuries, and other neurological diseases. Axonal degeneration can cause structural necrosis and functional disorders in the peripheral nervous system, ultimately leading to acquired or hereditary degenerative diseases of the central nervous system.

[0006] Although there is currently no very effective pharmacological method that can accurately assess the weight of the morbidity caused by axonal degeneration, it has been found in histopathological studies that significant axonal damage degradation has been observed in the early stages of various neuropathies such as Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis, and peripheral neuropathy, indicating that axonal degeneration plays an important role in the development of neuropathy (Fischer et al., Neuro-degenerative Diseases, 2007, 4:431-442). Therefore, by weakening or even blocking axonal degeneration, maintaining the integrity of neuronal structure and function may be a therapeutic solution that benefits various nervous system diseases.

[0007] In the absence of effective therapeutic drugs for neurodegenerative diseases, the existing technology urgently needs to research and develop new compounds, especially small chemical molecules, including compounds that have effects on axonal degeneration. Summary of the Invention

[0008] After long-term research, the present inventors unexpectedly discovered a class of compounds with significant SARM1 enzyme activity inhibition, and found that the compounds can be used to treat or prevent diseases or conditions related to SARM1 enzyme activity, including improving axonal degeneration, and for treating or preventing neurodegenerative diseases and related conditions.

[0009] The SARM1 (Sterile alpha and TIR motif containing 1) protein is composed of three domains: the ARM (Armadillo / HEAT repeat) domain at the nitrogen terminus, two tandem SAM (Sterile alpha motif) domains, and the TIR (Toll / Interleukin Receptor) domain at the carbon terminus. In addition, there is a mitochondrial localization signal peptide at the nitrogen terminus.

[0010] It is known that in wild-type neurons, axonal injury induces NAD + Depletion and axonal degeneration; SARM1 knockout inhibits axonal degeneration, and NAD + Maintained at normal levels, indicating that SARM1 promotes NAD + consumption, exacerbating axonal degeneration.

[0011] The Milbrandt team at the University of Washington School of Medicine prepared the TIR domain of SARM1 (SARM1-TIR) and found that it has NAD + Hydrolase activity. Further rigorous E. coli expression and purification experiments and cell-free expression systems were used to obtain high-purity SARM1-TIR, ultimately proving that SARM1-TIR can catalyze NAD + Produces adenosine 5'-diphosphate ribose (ADPR) and cyclic adenosine 5'-diphosphate ribose (cADPR).

[0012] SARM1 is a multifunctional signaling enzyme that catalyzes the conversion of multiple substrates to NAD + , NADP + and NA to generate signal molecules cADPR, ADPR and NAADP. In a variety of neurodegenerative diseases, SARM1 is activated, leading to NAD + Depletion of SARM1 can activate a new cell death mechanism; knocking out SARM1 can inhibit axonal degeneration and disease progression, and is therefore considered a potential drug target for related neurological diseases, including TBI, AD, CIPN, ASL, etc.

[0013] In the present disclosure, the inventors prepared full-length SARM1 for NAD enzyme activity experiments, and used it to screen and obtain compound molecules with enzyme activity inhibitory ability of the present invention.

[0014] Therefore, based on the above findings, in a first aspect, the present invention provides compounds of formula (I) that can be used as inhibitors of SARM1 enzyme activity:

[0015] or its racemate, enantiomer, diastereomer, deuterated form or pharmaceutically acceptable salt, wherein,

[0016] X1 is selected from -N-, -CH-;

[0017] R1 is independently selected from H, halogen, C1-C3 alkyl, cyano, trifluoromethyl, amino, hydroxy, methoxy, -C(O)NH2, preferably H, F, Cl, methyl, cyano;

[0018] R2 is independently selected from H, halogen, C1-C3 alkyl, cyano, trifluoromethyl, amino, hydroxy, methoxy, C1-C3 alkyl-OC(O)-, C1-C3 alkyl-NHC(O)-, preferably H, F, Cl, methyl, cyano;

[0019] M represents O, S or NH, preferably O;

[0020] L1 and L3 are each independently selected from -CH2-, -O-, -S- and -NH-, provided that at least one of L1 and L3 is -CH2-; wherein the H of -CH2- and -NH- may be substituted by a C1-C3 alkyl group;

[0021] L2 is selected from H, -CH3, -CH2-CH3, -OR3, -NHR4;

[0022] wherein R3 and R4 are independently selected from H and C1-C3 alkyl;

[0023] L1 can be connected to L2 to form a 4-membered ring, a 5-membered ring or a 6-membered ring, preferably a 5-membered ring;

[0024] A is selected from 5- to 10-membered aryl, 5- to 10-membered heteroaryl, 5- to 10-membered cycloalkyl, 5- to 10-membered heterocycloalkyl, 3- to 6-membered cycloalkyl and 5- to 10-membered aryl, 3- to 6-membered heterocycloalkyl and 5- to 10-membered aryl, wherein the heteroaryl or heterocycloalkyl may contain 1 or 2 heteroatoms selected from N, O and S, and the 5- to 10-membered aryl, 5- to 10-membered heteroaryl, 3- to 6-membered cycloalkyl and 5- to 10-membered aryl, 3- to 6-membered heterocycloalkyl and 5- to 10-membered aryl may be selected from 1, 2 or 3 substituted by the following substituents: halogen, C1-C3 alkyl, C1-C3 alkoxy, halo-C1-C3 alkyl, halo-C1-C3 alkoxy, cyano; preferably, the substituents are selected from methyl, methoxy, trifluoromethyl, trifluoromethoxy, F, Cl, Br and cyano; preferably, A is selected from phenyl, naphthyl, thienyl and tetrahydrofurophenyl optionally substituted by 1, 2 or 3 substituents selected from the following: F, Cl, Br, cyano, methyl, methoxy, trifluoromethyl.

[0025] In some embodiments, the compound of formula (I) of the present invention has the following structure:

[0026] in,

[0027] R1, R2, L3, A have the above definitions;

[0028] X2 is selected from CH2, O, NH, preferably CH2.

[0029] In some embodiments, the compound of formula (I) of the present invention has the following structure:

[0030] in,

[0031] R1, R2, L3, A have the above definitions;

[0032] X2 is selected from CH2, O, NH, preferably CH2.

[0033] In some embodiments, the compound of formula (I) of the present invention has the following structure:

[0034] in,

[0035] R1, R2, M, L1, L2, L3, and A have the same meanings as above.

[0036] In some embodiments, the compound of formula (I) of the present invention has the following structure:

[0037] in,

[0038] R1, R2, M, L1, L2, L3, and A have the same meanings as above.

[0039] In some embodiments, the compound of formula (I) of the present invention has the following structure:

[0040] in,

[0041] X1 as defined in claim 1;

[0042] R5, R6, and R7 are each independently selected from H, F, Cl, Br, cyano, methyl, methoxy, and trifluoromethyl.

[0043] In some embodiments of the present invention, a compound is provided, or a racemate, enantiomer, diastereomer, deuterated form, or pharmaceutically acceptable salt thereof, wherein the compound has a structure selected from the group consisting of:

[0044] In this article, when referring to the compound of formula (I), it also includes the pharmaceutically acceptable salts of the compound of formula (I) or its stereoisomers (including its enantiomers, diastereomers, racemates) or its tautomers or its deuterated products. Similarly, when referring to the compounds of formula (II) to formula (V), it also includes its pharmaceutically acceptable salts or its stereoisomers (including its enantiomers, diastereomers, racemates) or its tautomers or its deuterated products.

[0045] In another aspect, the present invention provides use of the compound of formula (I) of the present invention in the preparation of a SARM1 enzyme activity inhibitor or a medicament for treating or preventing a disease or condition related to SARM1 enzyme activity.

[0046] In another aspect, the present invention provides the use of the compound of formula (I) of the present invention in the preparation of a medicament for treating or preventing axonal degeneration-related diseases or disorders.

[0047] In another aspect, the present invention provides use of a compound of formula (I) according to the present invention in the preparation of a medicament for treating or preventing a neurodegenerative disease or a neurological disease or disorder.

[0048] Accordingly, the present invention also relates to a method for treating or preventing a neurodegenerative disease or a neurological disease or condition related thereto, comprising administering to a subject in need thereof a compound of formula (I) of the present invention as a SARM1 enzyme activity inhibitor. In particular, the present invention relates to a method for treating or preventing axonal degeneration-related diseases or conditions, comprising administering to a subject in need thereof a compound of formula (I) of the present invention as a SARM1 enzyme activity inhibitor. More particularly, the present invention relates to a method for inhibiting SARM1 enzyme activity, comprising administering to a subject in need thereof a compound of formula (I) of the present invention; more particularly, the present invention relates to a method for inhibiting axonal degeneration, comprising administering to a subject in need thereof a compound of formula (I) of the present invention. The compound of formula (I) of the present invention or a composition thereof can be administered to a desired subject or patient in an effective amount.

[0049] Accordingly, the present invention also relates to the use of the compounds of the present invention or their pharmaceutically acceptable salts or stereoisomers in the preparation of a method for treating or preventing a neurodegenerative disease or a neurological disease or condition. The present invention also relates to the use of the compounds of the present invention or their pharmaceutically acceptable salts or stereoisomers in the preparation of a SARM1 enzyme activity inhibitor. The present invention also relates to the use of the compounds of the present invention or their pharmaceutically acceptable salts or stereoisomers in the preparation of a method for treating or preventing axonal degeneration-related diseases or conditions. Preferably, the neurodegenerative disease or neurological disease or condition or axonal degeneration-related disease or condition is selected from Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis, and peripheral neuropathy. DETAILED DESCRIPTION

[0050] the term

[0051]

[0046] Reference herein to a "compound" having a particular structural formula is generally also intended to encompass pharmaceutically acceptable salts, stereoisomers, diastereomers, enantiomers, racemic mixtures, and isotopic derivatives thereof.

[0052] Pharmaceutically acceptable salts of the present invention may be formed using, for example, the following inorganic or organic acids: "Pharmaceutically acceptable salts" refers to salts that, within the scope of sound medical judgment, are suitable for use in contact with human and mammalian tissues without undue toxicity, irritation, allergic reaction, or the like, at a reasonable benefit / risk ratio. The salts may be prepared in situ during the final isolation and purification of the compounds of the invention, or separately by reacting the free base or free acid with a suitable reagent. For example, the free base may be reacted with a suitable acid. Examples of pharmaceutically acceptable acid addition salts are salts of an amino group (amine group) formed with inorganic acids (e.g., hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, and perchloric acid) or organic acids (e.g., acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid, or malonic acid), or salts formed using other methods known in the art, such as ion exchange. Other pharmaceutically acceptable salts include sodium alginate, ascorbate, benzenesulfonate, adipate, camphorsulfonate, aspartate, benzoate, bisulfate, borate, butyrate, camphorate, citrate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, heptanoate, hexanoate, hydroiodide, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate, and the like.

[0053] The pharmaceutically acceptable salts of the present invention can be prepared by conventional methods, for example, by dissolving the compound of the present invention in a water-miscible organic solvent (e.g., methanol, ethanol, acetone and acetonitrile), adding an excess of an organic acid or an aqueous inorganic acid solution thereto to precipitate the salt from the resulting mixture, removing the solvent and the remaining free acid therefrom, and then isolating the precipitated salt.

[0054] The "stereoisomerism" mentioned in the present invention is divided into conformational isomerism and configurational isomerism. Configurational isomerism can be further divided into cis-trans isomerism and optical isomerism (i.e., optical isomerism). Conformational isomerism refers to a stereoisomerism phenomenon in which the atoms or atomic groups of an organic molecule with a certain configuration have different spatial arrangements due to the rotation or distortion of carbon-carbon single bonds. Common examples include the structures of alkanes and cycloalkanes, such as the chair conformation and boat conformation that appear in the structure of cyclohexane. "Stereoisomers" refer to when the compounds of the present invention contain one or more asymmetric centers, and can thus be racemates and racemic mixtures, single enantiomers, diastereomeric mixtures, and single diastereoisomers. The compounds of the present invention have asymmetric centers, and each asymmetric center will produce two optical isomers. The scope of the present invention includes all possible optical isomers and diastereomeric mixtures and pure or partially pure compounds.

[0055] In some cases, the compounds of the present invention can exist as tautomers, which have different hydrogen attachment points through one or more double bond displacements. For example, a ketone and its enol form are keto-enol tautomers. Amides and imines can also form tautomeric forms. Each tautomer and mixture thereof are included in the compounds of the present invention. Enantiomers, diastereomers, racemates, mesomorphs, cis-trans isomers, tautomers and mixtures thereof of all compounds are included within the scope of the present invention.

[0056] When the compound of the present invention is used in combination with another SARM1 enzyme activity inhibitor for the treatment or prevention of neurodegenerative diseases or related neurological diseases or conditions, or can be used in combination with another active drug for the treatment or prevention of neurodegenerative diseases or related neurological diseases or conditions, it is used to treat or prevent neurodegenerative diseases or related diseases or conditions.

[0057] The compound of the present invention or its mesomer, racemate, enantiomer, diastereomer or its pharmaceutically acceptable salt can be used as an active ingredient by oral or parenteral administration, and the scope of its effective amount is 0.1 to 2000 mg / kg body weight / day, preferably 0.1 to 100 mg / kg body weight / day in the case of mammals including humans (weighing about 70 kg), and is administered every day in single or divided doses, or with or without a predetermined time. The dosage of the active ingredient can be adjusted according to multiple relevant factors (such as the situation of the subject to be treated, the type of disease and severity, the rate of administration and the doctor's opinion). In some cases, the amount less than the above dosage may be suitable.

[0058] The compound of the present invention or its mesoform, racemate, enantiomer, diastereomer or pharmaceutically acceptable salt can be formulated into a pharmaceutical composition. The pharmaceutical composition can comprise the compound of the present invention or its mesoform, racemate, enantiomer, diastereomer or pharmaceutically acceptable salt, and a pharmaceutically acceptable carrier.

[0059] The pharmaceutical composition of the present invention can be formulated into dosage forms for oral administration or parenteral administration (including intramuscular, intravenous and subcutaneous routes, intratumor injection) according to any of the conventional methods, such as tablets, granules, powders, capsules, syrups, emulsions, microemulsions, solutions or suspensions.

[0060] The pharmaceutical composition of the present invention for oral administration can be prepared by mixing the active ingredient with a pharmaceutically acceptable carrier such as cellulose, calcium silicate, magnesium stearate, calcium stearate, corn starch, lactose, sucrose, dextrose, calcium phosphate, stearic acid, surfactants, suspending agents, gelatin, talc, emulsifiers and diluents. Examples of carriers employed in the injectable composition of the present invention are water, glycerides, saline solutions, alcohols, glycols, glucose solutions, ethers (e.g., polyethylene glycol 400), oils, fatty acids, fatty acid esters, surfactants, suspending agents and emulsifiers.

[0061] If not stated otherwise, conventional methods of mass spectrometry, nuclear magnetic resonance, HPLC, protein chemistry, biochemistry, recombinant DNA technology and pharmacology are used. In this application, if not stated otherwise, "or" or "and" may be used to mean "and / or".

[0062] In the specification and claims, a given chemical formula or name shall encompass all stereo and optical isomers and racemates in which such isomers exist. Unless otherwise indicated, all chiral (enantiomers and diastereomers) and racemic forms are within the scope of the present invention. Many geometric isomers of C=C double bonds, C=N double bonds, ring systems, etc. may exist in the compounds, and all such stable isomers are encompassed by the present invention.

[0063] The compounds of the present invention can be isolated in optically active or racemic forms. All methods for preparing the compounds of the present invention and the intermediates prepared therein are considered part of the present invention. When enantiomeric or diastereomeric products are prepared, they can be separated by conventional methods (e.g., by chromatography or fractional crystallization). It should be understood that all isomeric forms that may exist are included in the present invention.

[0064] Unless otherwise defined, when a substituent is marked as "optionally substituted", the substituent is selected from, for example, substituents such as C1-C3 alkyl, C3-C6 cycloalkyl, C5-C 10Aryl, 3-10 membered heterocyclic group, halogen (including F, Cl, Br), hydroxyl, C1-C3 alkoxy, nitro, cyano, oxo (=O), C1-C3 alkyl acyl, C5-C 10 Aryloxy, C1-C3 alkylacyloxy, amino, C1-C3 alkylamino, C5-C 10 Arylamino, C1-C3 alkylthio, etc.

[0065] The term "alkyl" as used herein is intended to include both branched and straight chain saturated aliphatic hydrocarbon groups having the specified number of carbon atoms. 12 Alkyl, C1-C 10 C1-C8 alkyl, C1-C6 alkyl, more preferably C1-C4 alkyl, particularly preferably C1-C3 alkyl. For example, "C1-C6 alkyl" means an alkyl group having 1 to 6 carbon atoms. Examples of alkyl groups include, but are not limited to, methyl (Me), ethyl (Et), propyl (e.g., n-propyl and isopropyl), butyl (e.g., n-butyl, isobutyl, tert-butyl) and pentyl (e.g., n-pentyl, isopentyl, neopentyl). For the C1-C6 alkyl group in the present invention, 12 In the case of an alkyl group, 1 to 4 -CH2- units are optionally replaced by O atoms, S atoms or -NH-.

[0066] The term "carbonyl" refers to an organic functional group composed of carbon and oxygen atoms connected by a double bond (C=O or C(O)).

[0067] The term "aryl" refers to an aromatic 5- to 10-membered ring structure. An aromatic carbocyclic ring is an aromatic carbocyclic ring structure in which all ring members are composed of carbon atoms. A non-aromatic carbocyclic ring is a non-aromatic carbocyclic ring structure in which all ring members are composed of carbon atoms.

[0068] The term "heteroaryl" as used herein refers to a 5-10 membered aromatic group containing 1, 2 or 3 heteroatoms selected from O, N, S

[0069] "Halo" or "halogen" includes fluorine, chlorine, bromine and iodine; preferably fluorine, chlorine and bromine, more preferably fluorine and chlorine.

[0070] The term "heterocycle" refers to a monocyclic or bicyclic heterocycloalkyl system, and may also include spiro heterocycles or bridged heterocycloalkyls. A monocyclic heterocycloalkyl refers to a 3-8 membered, preferably 3-6 membered, saturated or unsaturated, but non-aromatic, cyclic alkyl system containing at least one heteroatom selected from O, N, S, or P (preferably containing 1, 2, or 3 heteroatoms selected from O, N, and S). A bicyclic heterocycloalkyl system refers to a heterocycloalkyl fused to a phenyl group, a cycloalkyl group, a cycloalkenyl group, a heterocycloalkyl group, or a heteroaryl group. An aromatic heterocycle refers to an aromatic heterocycle system, and a non-aromatic heterocycle refers to a non-aromatic heterocycle system.

[0071] When any variable occurs more than one time in any constituent or formula for a compound, its definition at each occurrence is independent of its definition at every other occurrence. Thus, for example, if a group is shown to be substituted with 0-3 R groups, then said group may be optionally substituted with up to three R groups, and at each occurrence R is independently selected from the definition of R. Furthermore, combinations of substituents and / or variables are permissible only if such combinations result in stable compounds.

[0072] As used herein, the term "effective amount" means an amount of a drug or pharmaceutical agent (i.e., a compound of the present invention) that will cause a biological or medical response in a tissue, system, animal, or human being, such as that sought by a researcher or clinician. In addition, the term "therapeutically effective amount" means an amount that results in improved treatment, cure, prevention, or alleviation of a disease, condition, or side effect, or reduces the rate of progression of a disease or condition, compared to a corresponding subject that has not received the above amount. An effective amount can be given in one or more administrations, applications, or dosages and is not intended to be limited by a specific formulation or route of administration. The term also includes within its scope an effective amount that enhances normal physiological function.

[0073] As used herein, the term "treating" includes any effect that results in improvement of a condition, disease, disorder, etc., such as alleviation, reduction, modulation, improvement, or elimination, or amelioration of the symptoms thereof.

[0074] The term "pharmaceutically acceptable" is used herein to refer to those compounds, substances, compositions and / or dosage forms that are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and animals without excessive toxicity, irritation, allergic response and / or other problems or complications, and commensurate with a reasonable benefit / risk ratio.

[0075] As used herein, the phrase "pharmaceutically acceptable carrier" means a pharmaceutical substance, composition, or vehicle, such as a liquid or solid filler, diluent, excipient, manufacturing aid (e.g., a lubricant, talc, magnesium stearate, calcium stearate, or zinc stearate, or stearic acid), or solvent encapsulating substance, which is involved in carrying or transporting the subject compound from one organ or part of the body to another. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient.

[0076] The term "pharmaceutical composition" means a composition comprising a compound of the present invention and at least one other pharmaceutical carrier. "Pharmaceutical carrier" refers to a medium generally accepted in the art for delivering biologically active agents to animals (particularly mammals), including (i.e.) adjuvants, excipients or vehicles such as diluents, preservatives, fillers, flow regulators, disintegrants, wetting agents, emulsifiers, suspending agents, sweeteners, flavoring agents, fragrances, antibacterial agents, antifungal agents, lubricants and dispersants, depending on the mode of administration and the nature of the dosage form.

[0077] The compounds or pharmaceutical compositions of the present invention can improve a disease, symptom, or condition after administration, particularly improving its severity, delaying its onset, slowing its progression, or reducing its duration, regardless of whether the administration is fixed or temporary, continuous or intermittent, and can be attributed to or related to the administration.

[0078] Route of administration

[0079] Suitable routes of administration for the compounds or pharmaceutical compositions of the present invention include, but are not limited to, oral, intravenous, rectal, aerosol, parenteral, ocular, pulmonary, transdermal, vaginal, auricular, nasal, and topical administration. In addition, parenteral administration, by way of example only, includes intramuscular, subcutaneous, intravenous, intramedullary, intraventricular, intraperitoneal, intralymphatic, and intranasal injections.

[0080] In some embodiments, the administration of the compound of the present invention is topical administration. In other specific embodiments, the compound of the present invention can be administered by implantation (e.g., subcutaneous or intramuscular) or by intramuscular injection. In addition, in other specific embodiments, the compound of the present invention is administered by a targeted drug delivery system.

[0081] In the pharmaceutical compositions of the present invention, pharmaceutical carriers can be formulated based on a variety of factors well within the purview of those skilled in the art. These factors include, but are not limited to: the type and nature of the active agent being formulated; the subject to whom the composition containing the active agent is to be administered; the intended route of administration of the composition; and the targeted therapeutic indication. Pharmaceutical carriers include aqueous and non-aqueous liquid media and various solid and semisolid dosage forms.

[0082] Such carriers may include a variety of ingredients and additives in addition to the active agent, which are included in the formulation for various reasons known to those skilled in the art, such as stabilizing the active agent, binding agents, etc. Descriptions of suitable pharmaceutical carriers and factors involved in carrier selection can be found in a number of readily available sources, such as Allen LV Jr. et al. Remington: The Science and Practice of Pharmacy (2 Volumes), 22 nd Edition(2012),Pharmaceutical Press.

[0083] The compounds are usually administered in admixture with suitable pharmaceutical diluents, excipients or carriers (collectively referred to herein as pharmaceutical carriers) appropriately selected according to the intended administration form (e.g., oral tablets, capsules, elixirs and syrups) and in accordance with conventional pharmaceutical practice.

[0084] While it is possible for a compound of the present invention to be administered alone, it is preferable to administer the compound as a pharmaceutical formulation (composition).

[0085] Kit / Product Packaging

[0086] Kits / product packaging are also described herein for use in treating the aforementioned indications. These kits can consist of a carrier, a pouch, or a container box, each of which can be divided into compartments to accommodate one or more containers, such as vials, test tubes, and the like, each containing a single component of the method. Suitable containers include bottles, vials, syringes, and test tubes. The containers can be made of acceptable materials such as glass or plastic.

[0087] For example, a container may contain one or more compounds described herein, either as a pharmaceutical composition or in admixture with other ingredients described herein. The container may have a sterile delivery port (e.g., an IV bag or bottle with a stopper pierceable by a hypodermic needle). Such a kit may include a compound and instructions, labeling, or operating instructions for use as described herein.

[0088] A typical kit may include one or more containers, each containing one or more materials (e.g., reagents, concentrated stock solutions, and / or instruments) to suit the commercialization and user needs of the compound. These materials include, but are not limited to, buffers, diluents, filters, needles, syringes, delivery devices, bags, containers, bottles, and / or test tubes, accompanied by a list of contents and / or instructions for use, including instructions for use within the packaging. The complete set of instructions should be included.

[0089] The features described above, or in the embodiments, may be combined in any combination. All features disclosed in this specification may be used in any combination, and each feature disclosed in this specification may be replaced by any alternative feature that serves the same, equivalent, or similar purpose. Therefore, unless otherwise specified, the features disclosed are merely general examples of equivalent or similar features.

[0090] The present invention will be further described below in conjunction with specific examples. It should be understood that these examples are intended to illustrate the present invention only and are not intended to limit the scope of the invention. The experimental methods in the following examples, for which specific conditions are not specified, are generally based on conventional conditions or the conditions recommended by the manufacturer. Unless otherwise stated, all percentages, ratios, proportions, or parts are by weight.

[0091] The units used in the present invention for weight-volume percentages are well known to those skilled in the art, for example, referring to the weight of solute in 100 ml of solution. Unless otherwise defined, all technical and scientific terms used herein have the same meanings as those familiar to those skilled in the art. In addition, any methods and materials similar or equivalent to those described herein can be applied to the present invention. The preferred embodiments and materials described herein are for illustrative purposes only.

[0092] In the terms used in the present invention, "neurodegenerative disease" and "neurodegenerative disease" have the same meaning; "axonal degeneration" and "axonal degeneration" have the same meaning. Those skilled in the art will understand that the terms have commonly understood meanings.

[0093] The descriptions provided herein, including those provided in the Examples, are not intended to limit the scope of any claims. The following non-limiting examples are provided to further illustrate the present invention. Based on this disclosure, those skilled in the art will appreciate that many changes may be made to the specific embodiments disclosed and still achieve the same or similar results without departing from the spirit and scope of the present teachings.

[0094] Unless otherwise noted, all materials / reagents were obtained from commercial suppliers and used without further purification. Reactions were monitored by LC-MS and / or thin-layer chromatography (TLC) on silica gel 60F254 (0.2 mm) pre-coated glass backing and observed using UV light. The structures of the following example compounds were characterized by nuclear magnetic resonance (NMR) and / or mass spectroscopy (MS).

[0095] 1 H NMR spectra (400 MHz) were recorded at room temperature on a Bruker spectrometer using TMS or residual solvent peaks as internal standards. Chemical shift values ​​or peak shape multiples are given in (δ), and coupling constants (J) are given in absolute values ​​in Hertz (Hz). 1The multiplicity in HNMR spectra is abbreviated as follows: s (singlet), d (doublet), t (triplet), q (quartet), m (multiplet), br or broad (broadened).

[0096] Preparative HPLC purification was performed on a Shimadzu LC-6AD. All purifications were performed using a Shim-pack PREP-DDS(H)KIT column. The mobile phase consisted of water (containing 0.1% HCO₂H₂) and acetonitrile; all reagents used were HPLC grade. The flow rate was 10 ml / min.

[0097] LC-MS was performed on an Agilent 1260 infinity II; mobile phase: A: water (0.1% trifluoroacetic acid), B: ACN; 3.5 min column run; column: YMC-Triart C18 50*3 mm, 3 um; flow rate: 1.8 ml / min; oven temperature: 40°C; gradient: 5-100 (ACN%).

[0098] Preparative TLC was performed on Whatman LK6F Silica Gel 60A plates of size 20 x 20 cm with a thickness of 500 μm.

[0099] The following examples are intended to illustrate embodiments of the present invention but are not intended to limit the same in any way.

[0100] Synthesis route of intermediate BB1

[0101] Step 1: Intermediate BB1-C tert-butyl 3-(4-fluorobenzyl)-2-oxopyrrolidine-1-carboxylate

[0102] The intermediate BB1-A tert-butyl 2-oxopyrrolidine-1-carboxylate (7404 mg, 40 mmol) was dissolved in THF (120 mL), cooled to -78°C under nitrogen, and lithium bis(trimethylsilyl)amide (42 mmol, 42 mL) was added. The reaction was controlled at temperature for 1 hour. The intermediate BB1-B 1-(bromomethyl)-4-fluorobenzene (7518 mg, 40 mmol) was dissolved in tetrahydrofuran (40 mL) and added to the reaction solution within 5 minutes. The reaction was controlled at temperature for 1 hour. Saturated ammonium chloride (30 mL) was added to quench the reaction. The organic solvent was removed by concentration under reduced pressure. The aqueous phase was extracted with ethyl acetate (50 mL x 3). The combined organic phases were washed with water (50 mL x 2) and saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was separated and purified by silica gel column chromatography (eluent: 27% ethyl acetate in n-hexane solution) to obtain a white solid intermediate BB1-C. tert-Butyl 3-(4-fluorobenzyl)-2-oxopyrrolidine-1-carboxylate (5240 mg, 18.22 mg), yield: 42.2%.

[0103] 1 H NMR (400MHz, Chloroform-d) δ7.21(dd,J=7.4,1.4Hz,1H),7.11(s,1H),7.06(d,J=7.5Hz,1H),4.67(s,3H),3.19(s,4H).

[0104] Step 2: Intermediate BB1 ​​3-(4-fluorobenzyl)pyrrolidin-2-one

[0105] The intermediate BB1-C tert-butyl 3-(4-fluorobenzyl)-2-oxopyrrolidine-1-carboxylate (5240 mg, 18.22 mmol) was dissolved in dichloromethane (20 mL), cooled to 0°C, and a solution of hydrochloric acid in 1.4-dioxane (45 mL, 182.2 mmol) was added. The mixture was stirred at room temperature for 1 hour and concentrated to dryness under reduced pressure. The residue was separated and purified by silica gel column chromatography (eluent: 7.6% methanol in dichloromethane) to obtain the intermediate BB1 ​​3-(4-fluorobenzyl)pyrrolidin-2-one (2950 mg) as a white solid in a yield of 84%.

[0106] LC_MS:(ES + ):m / z 194.07[M+H] +

[0107] Synthesis route of intermediate BB2

[0108] Step 1: Intermediate BB2 3-(4-fluorophenyl)propionamide

[0109] To a solution of compound BB2-A 3-(4-fluorophenyl)propanoic acid (200 mg, 1.189 mmol) in dichloromethane (5 mL) at 0°C under nitrogen was added oxalyl chloride (301.9 mg, 2.379 mmol), and the reaction was stirred continuously. After one hour, the mixture was concentrated under reduced pressure. The resulting residue was dissolved in tetrahydrofuran (5 mL), and aqueous ammonia (1 mL) was added. The reaction was stirred at room temperature for half an hour. LCMS confirmed the reaction was complete. The reaction mixture was quenched with water (10 mL) and extracted with ethyl acetate (10 mL x 2). The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: 7% methanol in dichloromethane) to afford the intermediate BB2 3-(4-fluorophenyl)propionamide (180 mg) as a white solid.

[0110] LC_MS:(ES + ):m / z 168.1[M+H] + .

[0111] Synthesis route of intermediate BB3

[0112] Step 1: Intermediate BB3-C methyl 2-cyano-3-(4-fluorophenyl)propionate

[0113] Intermediate BB3-A methyl cyanoacetate (3.93 g, 0.04 mol) was dissolved in DMSO (20 mL). Potassium carbonate (5.48 g, 0.04 mol) was added and stirred at room temperature for 30 min. Then, intermediate BB3-B 4-fluorobenzyl bromide (5.0 g, 0.026 mol) was added at 0°C and stirred at room temperature for 6 h. The reaction was monitored for completion by TLC. The reaction solution was poured into water (60 mL) and extracted twice with ethyl acetate (50 mL). The combined organic phases were washed three times with saturated brine (60 mL), dried over anhydrous sodium sulfate, filtered, and spin-dried. The crude product was isolated and purified by silica gel column chromatography (eluent: PE:EtOAc = 20:1 → 15:1 → 10:1) to obtain intermediate BB3-C methyl 2-cyano-3-(4-fluorophenyl)propanoate (1.13 g, 20% yield) as a colorless oil.

[0114] 1 H NMR (400MHz, CDCl3): δ7.26-7.23(m,2H),7.06-7.02(m,2H),3.80(s,3H),3.74-3.70(m,1H),3.28-3.16(m,1H).

[0115] Step 2: Intermediate BB3-D methyl 2-(aminomethyl)-3-(4-fluorophenyl)propionate

[0116] The intermediate BB3-C methyl 2-cyano-3-(4-fluorophenyl)propionate (1.13 g, 5.4 mmol) was dissolved in ethanol (20 mL) and Raney nickel (1.5 g) was added. The mixture was reacted overnight at room temperature under a hydrogen balloon. The reaction was monitored for completion by TLC. The reaction solution was filtered through celite, the filter cake was washed with ethanol (20 mL), and the filtrate was collected and dried to give the crude intermediate BB3-D methyl 2-(aminomethyl)-3-(4-fluorophenyl)propionate (1.1 g, crude product) as a colorless oil. This product was used directly in the next reaction without purification.

[0117] LC_MS:(ES + ):m / z 212.10[M+H] + .

[0118] Step 3: Intermediate BB3 3-[(4-fluorophenyl)methyl]azetidin-2-one

[0119] Intermediate BB3-D methyl 2-(aminomethyl)-3-(4-fluorophenyl)propanoate (1.1 g, 5.2 mmol) was dissolved in anhydrous ether (10 mL) and cooled to 0°C under N2. CH3MgI (7 mL, 20.8 mmol, 3 mol / L) was then slowly added dropwise to the reaction mixture. The ice bath was removed and the mixture was stirred at room temperature overnight. LCMS monitored the reaction for completion. The reaction mixture was quenched with saturated sodium bicarbonate (20 mL) under ice bath, filtered, and the filter cake was washed with EtOAc. The mixture was separated and the aqueous phase was extracted with EtOAc (20 mL x 2). The organic phases were combined and washed once with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and spin-dried. The crude product was purified by column chromatography (mobile phase DCM: CH3OH = 200:1→150:1), then stirred with n-hexane (5 mL) for 30 min and filtered to give the intermediate BB3 3-[(4-fluorophenyl)methyl]azetidin-2-one (125 mg, yield 13%) as a white solid.

[0120] LC_MS:(ES + ):m / z 180.00[M+H] + .

[0121] 1H NMR (400MHz, CDCl3): δ7.21-7.17(m,2H),6.99(t,J=8.8Hz,2H),5.66(brs,1H),3.53-3.5 1(m,1H),3.39(t,J=5.6Hz,1H),3.13-3.08(m,1H),3.04-3.02(m,1H),2.97-2.92(m,1H).

[0122] Synthesis route of intermediate BB4

[0123] Step 1: Compound BB4-C tert-butyl 3-(2-fluorobenzyl)-2-oxopiperidine-1-carboxylate

[0124] To a solution of compound BB4-A 1-(tert-butoxycarbonyl)-2-piperidone (1.06 g, 5.29 mmol) in tetrahydrofuran (20 mL) was added dropwise a solution of lithium N,N-diisopropylamine in tetrahydrofuran (4.0 mL, 2 M) under a nitrogen atmosphere at -78°C. The resulting mixture was stirred at -78°C for 1 hour, and then a solution of compound BB4-B 4-fluorobenzyl bromide (1.0 g, 5.29 mmol) in tetrahydrofuran (10 mL) was slowly added dropwise. After reacting at -78°C for 1 hour, saturated ammonium chloride solution (20 mL) was added to quench the reaction, and the mixture was partitioned with dichloromethane (15 mL × 3). The organic phase was collected and concentrated under reduced pressure. The crude product was isolated and purified by silica gel column chromatography (25% ethyl acetate in petroleum ether) to give compound BB4-C tert-butyl 3-(2-fluorobenzyl)-2-oxopiperidine-1-carboxylate (500 mg) as a white solid.

[0125] LC_MS:(ES + ):m / z 252.1[M+H-56] + .

[0126] Step 2: Compound BB4 3-(2-fluorobenzyl)piperidin-2-one

[0127] To a solution of BB4-C tert-butyl 3-(2-fluorobenzyl)-2-oxopiperidine-1-carboxylate (500 mg, 1.627 mmol) in dichloromethane (15 mL) was slowly added dropwise a 1,4-dioxane solution (2 mL, 4 M). The mixture was stirred on ice for 1.5 hours. TLC monitored the reaction for completion. The reaction mixture was concentrated under reduced pressure to afford crude BB4 3-(2-fluorobenzyl)piperidin-2-one (300 mg) as a white solid.

[0128] LC_MS:(ES+):m / z 208.1[M+H]+ .

[0129] Synthesis route of intermediate BB5

[0130] Step 1: Compound BB5 3-(4-fluorobenzyl)pyrrolidin-2-one-3-d

[0131] To a solution of compound BB5-A (1 g, 3.41 mmol) in tetrahydrofuran (15 mL) was added LiHMDS (4.09 ml, 4.09 mmol) at 0°C. The reaction mixture was stirred at 0°C for 0.5 hours. Deuterated methanol (5 mL) was added, and the mixture was allowed to warm to room temperature with stirring for 2 hours. LCMS monitored the reaction for completion. The reaction mixture was poured into saturated ammonium chloride solution (30 mL) and extracted with ethyl acetate (20 mL x 2). The combined organic phases were washed with saturated brine (30 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (eluent: 0%-5% methanol in dichloromethane) to afford compound BB5 3-(4-fluorobenzyl)pyrrolidin-2-one-3-d (500 mg) as a white solid.

[0132] LC_MS:(ES + ):m / z 195.1[M+H] + .

[0133] 1 H NMR (400MHz, CDCl3) δ7.22-7.15(m,2H),7.02-6.95(m,2H),6.36(s,1H),3.32-3.21(m,2 H),3.17(d,J=14.0Hz,1H),2.70(t,J=10.6Hz,1H),2.19-2.11(m,1H),1.88-1.80(m,1H).

[0134] Synthesis route of intermediate BB6

[0135] Step 1: Compound BB6 2-(bromomethyl)-5-chlorothiophene

[0136] Phosphorus tribromide (1.01 g, 3.735 mmol) was added to a solution of compound BB6-A (5-chlorothiophen-2-yl)methanol (370 mg, 2.49 mmol) in dichloromethane (15 mL) at 0°C. The reaction was stirred at 0°C for 1 hour. TLC (PE:EtOAc = 10:1) monitored the reaction for completion. The reaction mixture was poured into ice water (15 mL) and extracted with dichloromethane (15 mL x 2). The combined organic layers were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: 10% ethyl acetate in petroleum ether) to afford compound BB6 2-(bromomethyl)-5-chlorothiophene (360 mg) as a colorless oil.

[0137] Synthesis route of intermediate BB7

[0138] Step 1: Compound BB7 2-(bromomethyl)-5-fluoropyridine

[0139] Phosphorus tribromide (1.437 g, 5.31 mmol) was added to a solution of compound BB7-A (5-fluoropyridin-2-yl)methanol (450 mg, 3.54 mmol) in dichloromethane (10 mL) at 0°C. The mixture was stirred at 25°C for 1 hour. TLC (PE:EtOAc = 5:1) monitored the reaction completion. The reaction mixture was poured into ice water (15 mL), adjusted to pH 8-9 with saturated NaHCO₃ solution, and extracted with dichloromethane (20 mL x 2). The combined organic layers were washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: 20% ethyl acetate in petroleum ether) to afford compound BB7 2-(bromomethyl)-5-fluoropyridine (505 mg) as a colorless oil.

[0140] Synthesis route of intermediate BB8

[0141] Step 1: Compound BB8-C(E)-2-oxo-3-(pyridin-4-ylmethylene)pyrrolidine-1-carboxylic acid tert-butyl ester

[0142] Compound BB8-A tert-butyl 2-oxopyrrolidine-1-carboxylate (925 mg, 5 mmol) was dissolved in tetrahydrofuran (30 mL), cooled to -78°C under nitrogen, lithium bistrimethylsilylamide (5.25 mL, 5.25 mmol) was added, and the reaction was incubated for 1 hour. Then, compound BB8-B 4-pyridinecarboxaldehyde (535 mg, 5 mmol) dissolved in tetrahydrofuran (7 mL) was added, and the reaction was incubated for 2.5 hours. Two drops of saturated ammonium chloride were added to quench the reaction, and the mixture was stirred and warmed to room temperature. Anhydrous sodium sulfate was added, and the mixture was filtered and concentrated to dryness. The residue was separated and purified by silica gel column chromatography (eluent: dichloromethane solution containing 8.8% methanol) and preparative HPLC to obtain a crude yellow solid compound BB8-C(E)-tert-butyl 2-oxo-3-(pyridin-4-ylmethylene)pyrrolidine-1-carboxylate (51 mg, 3.7%).

[0143] Step 2: Compound BB8-D tert-butyl 2-oxo-3-(pyridin-4-ylmethyl)pyrrolidine-1-carboxylate

[0144] The compound BB8-C(E)-2-oxo-3-(pyridin-4-ylmethylene)pyrrolidine-1-carboxylic acid tert-butyl ester (51 mg, 0.185 mmol) was dissolved in methanol (5 mL), and 10% palladium carbon (10 mg) was added. The mixture was reacted at room temperature for 16 h under a hydrogen atmosphere, filtered, and concentrated under reduced pressure. The residue was separated and purified by preparative thin layer chromatography (developing solvent: dichloromethane solution containing 7% methanol) to obtain the white solid compound BB8-D 2-oxo-3-(pyridin-4-ylmethyl)pyrrolidine-1-carboxylic acid tert-butyl ester (40 mg, 77%).

[0145] LC_MS:(ES + ):m / z 277.1[M+H] + .

[0146] Step 3: Compound BB8 3-(pyridin-4-ylmethyl)pyrrolidin-2-one hydrochloride

[0147] Compound BB8-D tert-butyl 2-oxo-3-(pyridin-4-ylmethyl)pyrrolidine-1-carboxylate (40 mg, 0.18 mmol) was dissolved in anhydrous dichloromethane (1 mL), cooled to 0°C under nitrogen protection, and hydrogen chloride / 1,4-dioxane solution (1 mL, 4 M) was added to react for 30 minutes. The mixture was concentrated under reduced pressure to give compound BB8 3-(pyridin-4-ylmethyl)pyrrolidin-2-one hydrochloride (32 mg) as a white solid.

[0148] LC_MS:(ES + ):m / z 177.1[M+H] + .

[0149] Synthesis route of intermediate BB9

[0150] Step 1: Compound BB9 1-(bromomethyl)-4-methoxybenzene

[0151] Compound BB9-A (4-methoxyphenyl)methanol (1 g, 7.237 mmol) was dissolved in dichloromethane (10 mL). The reaction mixture was placed in an ice bath and phosphorus tribromide (687 μL) was slowly added dropwise with stirring. The mixture was allowed to return to room temperature and stirred for 1 hour. After the reaction was complete, the solvent was removed by concentration under reduced pressure. Dichloromethane (50 mL) was added for dilution and further concentration was performed to obtain crude compound BB9 (1-(bromomethyl)-4-methoxybenzene) (800 mg) as a colorless oil. This was used directly in the next step without further purification.

[0152] Synthesis route of intermediate BB10

[0153] Step 1: Compound BB10-B (5-chloropyridin-2-yl)methanol

[0154] To a solution of compound BB10-A (5-chloropyridine-2-carboxaldehyde) (1.0 g, 7.06 mmol) in methanol (15 mL) was added sodium borohydride (400 mg, 10.60 mmol) portionwise under ice-cooling conditions. The resulting mixture was stirred at room temperature for 2.5 hours. After completion of the reaction as monitored by TLC, the reaction mixture was concentrated under reduced pressure and quenched by the addition of saturated ammonium chloride solution (5 mL). The pH was adjusted to neutral with saturated sodium bicarbonate solution, and the mixture was extracted with dichloromethane (15 mL x 2). The organic phase was collected and concentrated under reduced pressure to afford the crude compound BB10-B (5-chloropyridin-2-yl)methanol (1.1 g) as a white solid.

[0155] LC_MS:(ES + ):m / z 144.0[M+H] + .

[0156] Step 2: Compound BB10 2-(Bromomethyl)-5-chloropyridine

[0157] Phosphorus tribromide (2.9 g, 10.60 mmol) was added dropwise to a solution of compound BB10-B (5-chloropyridin-2-yl)methanol (1.1 g, 7.06 mmol) in dichloromethane (15 mL) under ice-cooling conditions. The resulting mixture was stirred at room temperature for 2.5 hours. After completion of the reaction, as monitored by TLC, the reaction mixture was poured into saturated aqueous sodium bicarbonate solution and partitioned between dichloromethane (15 mL x 3). The organic phase was collected and concentrated under reduced pressure. The resulting crude product was isolated and purified by silica gel column chromatography (eluting with 9% ethyl acetate in petroleum ether) to afford compound BB10 2-(bromomethyl)-5-chloropyridine (800 mg) as a colorless oil.

[0158] LC_MS:(ES + ):m / z 207.9[M+H] + .

[0159] Example 1: Synthesis route of compound 2

[0160] Step 1: Compound 2-C 5-iodo-2-(pyridin-4-yl)pyridazin-3(2H)-one

[0161] To a solution of Compound 2-A (5-iodopyridazin-3(2H)-one) (2.5 g, 11.26 mmol), Compound 2-B (4-pyridineboronic acid) (4.15 g, 33.78 mmol), and cupric acetate (409 mg, 2.25 mmol) in N,N-dimethylformamide (40 mL) was added pyridine (5.34 g, 67.56 mmol) at room temperature. The reaction mixture was heated to 80°C and stirred in air for 18 hours. The reaction was complete, as monitored by TLC. The reaction solution was cooled to room temperature, quenched with water (40 mL), and extracted with ethyl acetate (40 mL x 2). The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting crude product was separated and purified by silica gel column chromatography (eluent: 0%-10% methanol in dichloromethane) to give 2-C 5-iodo-2-(pyridin-4-yl)pyridazin-3(2H)-one (1.4 g) as a light purple solid.

[0162] LC_MS:(ES + ):m / z 300.0[M+H] + .

[0163] 1 H NMR (400MHz, DMSO) δ8.73 (s, 2H), 8.38 (d, J = 2.0Hz, 1H), 7.81 (d, J = 2.0Hz, 1H), 7.74-7.69 (m, 2H).

[0164] Step 2: Compound 2 5-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl)-2-(pyridin-4-yl)pyridazin-3(2H)-one

[0165] Under nitrogen, to a solution of 2-C 5-iodo-2-(pyridin-4-yl)pyridazin-3(2H)-one (50 mg, 0.167 mmol), intermediate BB1 ​​3-(4-fluorobenzyl)pyrrolidin-2-one (32.3 mg, 0.167 mmol), Xant-Phos (9.7 mg, 0.0167 mmol), and cesium carbonate (108.8 mg, 0.334 mmol) in 1,4-dioxane (3 mL) was added Pd2(dba)3 (15.3 mg, 0.0167 mmol). The reaction system was purged with nitrogen three times and heated to 100°C for 2 hours. LCMS monitored the reaction for completion. The reaction solution was filtered through celite and concentrated under reduced pressure. The crude product was separated and purified by preparative thin-layer chromatography (eluent: 5% methanol in dichloromethane) to give compound 2 5-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl)-2-(pyridin-4-yl)pyridazin-3(2H)-one (24.5 mg) as a white solid.

[0166] LC_MS:(ES + ):m / z 365.9[M+H] + .

[0167] 1 H NMR(400MHz, CDCl3)δ9.24(d,J=2.5Hz,1H),8.82-8.70(m,2H),7.82(dd,J=4.8 ,1.5Hz,2H),7.27-7.16(m,2H),7.09-6.98(m,2H),6.45(d,J=2.5Hz,1H),3.70 -3.53(m,2H),3.24(dd,J=13.8,4.3Hz,1H),2.98(ddd,J=18.0,8.8,4.3Hz,1H) ,2.86(dd,J=13.8,8.6Hz,1H),2.36-2.25(m,1H),1.96(dq,J=12.9,8.8Hz,1H).

[0168] Example 2: Synthesis route of compound 23

[0169] Step 1: Compound 23-C 4-bromo-2H-[1,4'-bipyridyl]-2-one

[0170] To a solution of Compound 23-A (4-bromopyridin-2(1H)-one) (500 mg, 2.873 mmol) and Compound 23-B (1.059 g, 8.619 mmol) in DMF (10 mL) were added copper acetate (104.4 mg, 0.5747 mmol) and pyridine (681.7 mg, 8.619 mmol). The mixture was heated to 80°C in an open atmosphere for 12 hours. LCMS monitored the reaction for completion. The reaction solution was extracted with ethyl acetate (20 mL x 2) and water (25 mL). The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The resulting crude product was isolated and purified by silica gel column chromatography (eluent: 5% methanol in dichloromethane) to afford Compound 23-C (450 mg) as a white solid, 4-bromo-2H-[1,4'-bipyridyl]-2-one.

[0171] LC_MS:(ES + ):m / z 250.9[M+H] + .

[0172] 1 H NMR (400MHz, CDCl3) δ8.80(d,J=5.8Hz,2H),7.40(dd,J=4.7,1.5Hz,2H),7.22(d,J=7.4Hz,1H),6.96(d,J=2.0Hz,1H),6.50(dd,J=7.4,2.1Hz,1H).

[0173] Step 2: Compound 23 4-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl)-2H-[1,4'-bipyridyl]-2-one

[0174] To a solution of compound 23-C (4-bromo-2H-[1,4'-bipyridyl]-2-one) (50 mg, 0.199 mmol), intermediate BB1 ​​(38.5 mg, 0.199 mmol), Xant-Phos (11.5 mg, 0.0199 mmol), and cesium carbonate (129.8 mg, 0.398 mmol) in 1,4-dioxane (3 mL) was added Pd2(dba)3 (18.2 mg, 0.0199 mmol) under nitrogen. The reaction system was purged with nitrogen three times and heated to 100°C for 2 hours. LCMS monitored the reaction for completion. The reaction solution was filtered through celite and concentrated under reduced pressure. The crude product was separated and purified by preparative thin-layer chromatography (eluent: 5% methanol in dichloromethane) to give compound 23 4-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl)-2H-[1,4'-bipyridyl]-2-one (30.2 mg) as a white solid.

[0175] LC_MS:(ES + ):m / z 364.1[M+H] + .

[0176] 1 H NMR (400MHz, CDCl3) δ8.78(s,2H),7.66(d,J=6.5Hz,1H),7.44(s,2H),7.30(s,1H),7.20(s,2H),7.03(d,J=7.5Hz,2H) ,6.15(s,1H),3.62(d,J=4.9Hz,2H),3.23(d,J=13.3Hz,1H),2.94(s,1H),2.88-2.69(m,1H),2.22(s,1H),1.88(s,1H).

[0177] Example 3: Synthesis route of compound 10

[0178] Step 1: Compound 10 3-(4-fluorophenyl)-N-(6-oxo-1-(pyridin-4-yl)-1,6-dihydropyridazin-4-yl)propionamide

[0179] Under nitrogen, to a solution of 2-C 5-iodo-2-(pyridin-4-yl)pyridazin-3(2H)-one (50 mg, 0.167 mmol), intermediate BB2 3-(4-fluorophenyl)propionamide (27.9 mg, 0.167 mmol), Xant-Phos (9.7 mg, 0.0167 mmol), and cesium carbonate (108.8 mg, 0.334 mmol) in 1,4-dioxane (3 mL) was added Pd2(dba)3 (15.3 mg, 0.0167 mmol). The reaction system was purged with nitrogen three times and heated to 100°C for 2 hours. LCMS monitored the reaction for completion. The reaction solution was filtered through celite and concentrated under reduced pressure. The crude product was separated and purified by preparative thin-layer chromatography (eluent: 5% methanol in dichloromethane) to give Compound 10 3-(4-fluorophenyl)-N-(6-oxo-1-(pyridin-4-yl)-1,6-dihydropyridazin-4-yl)propanamide (23.8 mg) as a white solid.

[0180] LC_MS:(ES + ):m / z 339.1[M+H] + .

[0181] 1H NMR (400MHz, DMSO) δ10.63(s,1H),8.68(dd,J=4.7,1.6Hz,2H),8.07(d,J=2.4Hz,1H),7.73(dd,J=4.7,1.6Hz, 2H),7.37(d,J=2.4Hz,1H),7.33-7.27(m,2H),7.16-7.09(m,2H),2.92(t,J=7.5Hz,2H),2.73(t,J=7.5Hz,2H).

[0182] Example 4: Synthesis route of compound 26

[0183] Step 1: Compound 26 3-(4-fluorophenyl)-N-(2-oxo-2H-[1,4'-bipyridyl]-4-yl)propionamide

[0184] To a solution of compound 23-C 4-bromo-2H-[1,4'-bipyridyl]-2-one (50 mg, 0.199 mmol), intermediate BB2 3-(4-fluorophenyl)propionamide (33.3 mg, 0.199 mmol), Xant-Phos (11.5 mg, 0.0199 mmol), and cesium carbonate (129.8 mg, 0.398 mmol) in 1,4-dioxane (3 mL) was added Pd2(dba)3 (18.2 mg, 0.0199 mmol) under nitrogen. The reaction system was purged with nitrogen three times and heated to 100°C for 2 hours. LCMS monitored the reaction for completion. The reaction solution was filtered through celite and concentrated under reduced pressure. The crude product was separated and purified by preparative thin-layer chromatography (eluent: 5% methanol in dichloromethane) to give compound 263-(4-fluorophenyl)-N-(2-oxo-2H-[1,4'-bipyridyl]-4-yl)propanamide (28.3 mg) as a white solid.

[0185] LC_MS:(ES+):m / z 338.3[M+H] +

[0186] 1H NMR (400MHz, DMSO) δ10.22(s,1H),8.70(dd,J=4.6,1.6Hz,2H),7.66(d,J=7.6Hz,1H),7.51(dd,J=4.6,1.6Hz,2H),7.28(dd,J=8.6,5. 6Hz,2H),7.12(dd,J=12.3,5.5Hz,2H),6.90(d,J=2.2Hz,1H),6.46(dd,J=7.6,2.3Hz,1H),2.90(t,J=7.6Hz,2H),2.68(t,J=7.6Hz,2H)

[0187] Example 5: Synthesis route of compound 31

[0188] Step 1: Compound 31 5-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl-3-d)-2-(pyridin-4-yl)pyridazin-3(2H)-one

[0189] Under nitrogen, to a solution of 2-C 5-iodo-2-(pyridin-4-yl)pyridazin-3(2H)-one (80 mg, 0.267 mmol), BB5 3-(4-fluorobenzyl)pyrrolidin-2-one-3-d (52.0 mg, 0.267 mmol), Xant-Phos (15.4 mg, 0.0267 mmol), and cesium carbonate (174.0 mg, 0.534 mmol) in 1,4-dioxane (3 mL) was added Pd2(dba)3 (24.4 mg, 0.0267 mmol). The reaction system was purged with nitrogen three times and heated to 100°C for 2 hours. LCMS monitored the reaction for completion. The reaction solution was filtered through celite and concentrated under reduced pressure. The crude product was separated and purified by preparative thin-layer chromatography (eluent: 5% methanol in dichloromethane) to give Compound 31 5-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl-3-d)-2-(pyridin-4-yl)pyridazin-3(2H)-one (42 mg) as a white solid.

[0190] LC_MS:(ES + ):m / z 366.6[M+H] + .

[0191] 1H NMR (400MHz, CDCl3) δ9.23(d,J=2.5Hz,1H),8.73(dd,J=4.8,1.5Hz,2H),7.82(dd,J=4.7,1.6Hz,2H),7.25-7.15(m,2H),7.07-6.97(m,2H),6.45(d, J=2.5Hz,1H),3.69-3.53(m,2H),3.27-3.19(m,1H),2.86(dd,J=13.9,5.4 Hz, 1H), 2.30 (ddd, J=12.7, 7.4, 3.2Hz, 1H), 1.95 (dt, J=12.9, 8.6Hz, 1H).

[0192] Example 6: Synthesis route of compound 35

[0193] Step 1: Compound 35-C 1-(4-fluorobenzyl)imidazolidin-2-one

[0194] To a solution of Compound 35-A imidazolidin-2-one (1.0 g, 11.63 mmol) in N,N-dimethylformamide (20 mL) was added sodium hydroxide (465 mg, 11.63 mmol, 60% wt). The reaction mixture was cooled to 0°C, and Compound 35-B 1-(bromomethyl)-4-fluorobenzene (2.2 g, 11.63 mmol) was slowly added portionwise. The reaction mixture was allowed to warm to room temperature and stirred for 4 hours. TLC confirmed the reaction completion. The reaction mixture was diluted with water (50 mL) and extracted with ethyl acetate (20 mL x 2). The organic layers were combined, washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was isolated and purified by silica gel column chromatography (eluent: 5% methanol in dichloromethane) to afford Compound 35-C 1-(4-fluorobenzyl)imidazolidin-2-one (200 mg) as a yellow solid.

[0195] LC_MS:(ES + ):m / z 195.1[M+H] + .

[0196] Step 2: Compound 35 5-(3-(4-fluorobenzyl)-2-oxoimidazolidin-1-yl)-2-(pyridin-4-yl)pyridazin-3(2H)-one

[0197] To a solution of compound 35-C 1-(4-fluorobenzyl)imidazolidin-2-one (50 mg, 0.258 mmol) in 1,4-dioxane (2 mL) were added cesium carbonate (167 mg, 0.515 mmol), compound 2-C 5-iodo-2-(pyridin-4-yl)pyridazin-3(2H)-one (77 mg, 0.258 mmol), tris(dibenzylideneacetone)dipalladium (23 mg, 0.026 mmol), and 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (13 mg, 0.026 mmol). The atmosphere was purged with nitrogen three times, and the reaction mixture was stirred at 100°C for 4 hours. TLC monitored the reaction for completion. The reaction mixture was filtered, the filtrate concentrated, diluted with dichloromethane (3 mL), washed with saturated brine (2 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated and purified by silica gel column chromatography (eluent: 5% methanol in dichloromethane solution) to give yellow solid compound 35 5-(3-(4-fluorobenzyl)-2-oxoimidazolidin-1-yl)-2-(pyridin-4-yl)pyridazin-3(2H)-one (52 mg). LC_MS: (ES + ):m / z 366.1[M+H] + .

[0198] Example 7: Synthesis route of compound 36

[0199] Step 1: Compound 36-A (6-oxo-1-(pyridin-4-yl)-1,6-dihydropyridazin-4-yl)carbamic acid tert-butyl ester

[0200] The synthesis of this step uses the intermediate 2-C 5-iodo-2-(pyridin-4-yl)pyridazin-3(2H)-one and tert-butyl carbamate as raw materials, and refers to the synthesis process of the second step of Example 1.

[0201] LC_MS:(ES + ):m / z 289.1[M+H] + .

[0202] Step 2: Compound 36-B 5-amino-2-(pyridin-4-yl)pyridazin-3(2H)-one

[0203] To a dichloromethane (3 mL) solution containing compound 36-A (6-oxo-1-(pyridin-4-yl)-1,6-dihydropyridazin-4-yl)carbamic acid tert-butyl ester (210 mg, 0.728 mmol) was added hydrogen chloride / 1,4-dioxane (3 mL, 4 M) at room temperature. The reaction solution was stirred at 25 ° C for 1 hour. LCMS monitored the reaction to be complete. The reaction solution was concentrated, saturated sodium bicarbonate solution (10 mL) was added, stirred for 30 minutes, filtered, the filter cake was collected, and dried under reduced pressure to obtain a yellow solid compound 36-B 5-amino-2-(pyridin-4-yl)pyridazin-3(2H)-one (118 mg). LC_MS: (ES + ):m / z 189.1[M+H] + .

[0204] Step 3: Compound 36 1-(4-fluorobenzyl)-3-(6-oxo-1-(pyridin-4-yl)-1,6-dihydropyridazin-4-yl)urea

[0205] To a solution of compound 36-C (4-fluorophenyl)methanamine (20 mg, 159.82 μmol) and triethylamine (48.5 mg, 479.46 μmol) in N,N-dimethylformamide (1 mL) was added N,N'-carbonyldiimidazole (25.9 mg, 159.82 μmol) at room temperature and stirred at room temperature for half an hour. Compound 36-B (5-amino-2-(pyridin-4-yl)pyridazin-3(2H)-one) (30.1 mg) was then added to the reaction mixture, which was heated to 80°C and stirred for 3 hours. LCMS confirmed the reaction was complete. The reaction mixture was diluted with water (10 mL) and extracted with ethyl acetate (8 mL x 2). The organic layers were combined, washed with saturated brine (15 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was separated and purified by preparative thin-layer chromatography (developing solvent: dichloromethane solution containing 5% methanol) to obtain a white solid compound, Compound 36 1-(4-fluorobenzyl)-3-(6-oxo-1-(pyridin-4-yl)-1,6-dihydropyridazin-4-yl)urea (2.33 mg). LC_MS: (ES + ):m / z 340.1[M+H] + .

[0206] Example 8: Preparation of Compound 130 and Compound 131

[0207] Compound 2 (100 mg) was dissolved in acetonitrile:methanol = 1:1 and separated by SFC to give Compound 130 (38 mg) and Compound 131 (35 mg). The separation method was as follows: mobile phase: n-hexane:isopropanol = 65:35, flow rate: 8 mL / min, pressure: 4 MPa, detection wavelength: 254 nm, column temperature: room temperature, chromatographic column: Phenomenex Chrial ND column (250 mm*21 mm, 2 mm), retention time (peak 1) = 45 minutes, retention time (peak 2) = 54 minutes.

[0208] Peak 1: LC_MS: (ES + ):m / z 365.1[M+H] + .

[0209] Peak 2: LC_MS: (ES + ):m / z 365.1[M+H] + .

[0210] Example 9: Synthesis route of compound 154

[0211] Step 1: Compound 154-B 5-iodo-2-(tetrahydro-2H-pyran-2-yl)pyridazin-3(2H)-one

[0212] Compound 154-A, 5-iodopyridazin-3(2H)-one (9580 mg, 43.17 mmol), was added to tetrahydrofuran (72 mL). Pyridinium p-toluenesulfonate (2061 mg, 8.2 mmol) and 3,4-dihydro-2H-pyran (7.6 mL, 86.34 mmol) were then added. The mixture was stirred for five minutes and then heated to 65°C and refluxed for five hours. 3,4-dihydro-2H-pyran (4.9 mL, 55.7 mmol) was then added and refluxed overnight. The mixture was cooled to room temperature, filtered, and rinsed with tetrahydrofuran (20 mL x 3). The filtrate was concentrated to dryness to afford crude compound 154-B, 5-iodo-2-(tetrahydro-2H-pyran-2-yl)pyridazin-3(2H)-one (14.65 g), a pale yellow viscous liquid, which was used directly in the next step.

[0213] LC_MS:(ES + ):m / z 306.9[M+H] + .

[0214] Step 2: Compound 154-C 5-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl)-2-(tetrahydro-2H-pyran-2-yl)pyridazin-3(2H)-one

[0215] A solution of crude compound 154-B (5-iodo-2-(tetrahydro-2H-pyran-2-yl)pyridazin-3(2H)-one (43.17 mmol), intermediate BB1 ​​(3-(4-fluorobenzyl)pyrrolidin-2-one) (7473 mg, 38.7 mmol), tris(dibenzylideneacetone)dipalladium (1967 mg, 2.15 mmol), 4,5-bis(diphenylphosphine)-9,9-dimethylxanthene (2485 mg, 4.3 mmol), and cesium carbonate (35 g, 107.5 mmol) in 1.4-dioxane (172 mL) was heated to 80°C under a nitrogen atmosphere for 1 hour. The reaction was complete as determined by LCMS. The mixture was cooled to room temperature, filtered under nitrogen atmosphere, washed with 1.4-dioxane (30 mL*2), and concentrated to dryness to give a gray crude compound 154-C 5-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl)-2-(tetrahydro-2H-pyran-2-yl)pyridazin-3(2H)-one (16.0 g), which was directly carried out to the next step.

[0216] LC_MS:(ES + ):m / z 372.1[M+H] + .

[0217] Step 3: Compound 154-D 5-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl)pyridazin-3(2H)-one

[0218] The crude compound 154-C 5-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl)-2-(tetrahydro-2H-pyran-2-yl)pyridazin-3(2H)-one (16.0 g) was dissolved in dichloromethane (50 mL) and stirred until completely dissolved. The mixture was cooled to 0°C under nitrogen protection. HCl / Dioxane (120 mL, 480 mmol) was added dropwise and the temperature was raised to room temperature for 1 hour. The mixture was concentrated to dryness under reduced pressure, washed twice with toluene (100 mL*2) and three times with dichloromethane (100 mL*3). The residue was added to dichloromethane (200 mL) and slurried for 2 hours, filtered, and the filter cake was washed with dichloromethane (50 mL*2). The residue was collected and added with dichloromethane (100 mL) and water (100 mL). The pH was adjusted to greater than 8 with saturated sodium bicarbonate aqueous solution, filtered, washed with pure water (100 mL*2), and the solid was collected and dried to give a light yellow solid compound 154-D5-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl)pyridazin-3(2H)-one (7470 mg).

[0219] LC_MS:(ES + ):m / z 288.1[M+H] + .

[0220] Step 4: Compound 154 4-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl)-6H-[1,4'-bipyridazine]-6-one

[0221] To a dioxane solution (2 mL) containing compound 154-D 5-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl)pyridazin-3(2H)-one (20 mg, 0.07 mmol), compound 154-E 4-bromopyridazine hydrobromide (16.6 mg, 0.07 mmol), Xant-Phos (4.02 mg, 0.007 mmol), and cesium carbonate (113.1 mg, 0.35 mmol) was added Pd2(dba)3 (3.2 mg, 0.0035 mmol) under nitrogen at room temperature. The reaction mixture was purged with nitrogen three times. The reaction mixture was heated to 110°C and stirred for 18 hours. The reaction was complete, monitored by TLC. The reaction mixture was cooled to room temperature, filtered, and the filtrate concentrated. The crude product was separated and purified by preparative thin layer chromatography (eluted with 5% methanol in dichloromethane) to give compound 154 4-(3-(4-fluorobenzyl)-2-oxopyrrolidin-1-yl)-6H-[1,4'-bipyridazine]-6-one (0.6 mg)

[0222] LC_MS:(ES + ):m / z 366.2[M+H] + .

[0223] Example 10: Preparation of Compound 138 and Compound 139

[0224] Compound 7 (15 mg) was dissolved in acetonitrile:methanol = 1:1 and separated by SFC to give compounds 138 (5 mg) and 139 (6 mg). The separation method was as follows: mobile phase: n-hexane:isopropanol = 70:30, flow rate: 8 mL / min, pressure: 4 MPa, detection wavelength: 254 nm, column temperature: room temperature, chromatographic column: Phenomenex Chrial ND column (250 mm*21 mm, 2 mm), retention time (peak 1) = 45 minutes, retention time (peak 2) = 49 minutes.

[0225] Peak 1: LC_MS: (ES + ):m / z 381.1[M+H] + .

[0226] Peak 2: LC_MS: (ES + ):m / z 381.1[M+H] + .

[0227] Example 11: Preparation of Compound 146 and Compound 147

[0228] Compound 88 (12 mg) was dissolved in acetonitrile:methanol = 1:1 and separated by SFC to give compounds 138 (4 mg) and 139 (4 mg). The separation method was as follows: mobile phase: n-hexane:isopropanol = 60:40, flow rate: 8 mL / min, pressure: 4 MPa, detection wavelength: 254 nm, column temperature: room temperature, chromatographic column: Phenomenex Chrial ND column (250 mm*21 mm, 2 mm), retention time (peak 1) = 30 minutes, retention time (peak 2) = 38 minutes.

[0229] Peak 1: LC_MS: (ES + ):m / z 400.1[M+H] + .

[0230] Peak 2: LC_MS: (ES + ):m / z 400.1[M+H] + .

[0231] The synthetic routes of the following compounds are as follows, referring to the synthetic methods of intermediate BB1 ​​and Example 1, using corresponding reagents for synthesis:

[0232] The synthetic routes of the following compounds are as follows, referring to the synthetic methods of intermediate BB1 ​​and Example 2, using corresponding reagents for synthesis:

[0233] The synthetic routes of the following compounds are as follows, referring to the synthetic methods of intermediate BB2 and Example 3, using corresponding reagents for synthesis:

[0234] The synthetic routes of the following compounds are as follows, referring to the synthetic methods of intermediate BB2 and Example 4, using corresponding reagents for synthesis:

[0235] The synthetic routes of the following compounds are as follows, referring to the synthetic method of Example 1, using corresponding reagents for synthesis:

[0236] The following compounds were prepared by referring to the SFC separation method of Example 8, Example 10, and Example 11 using the corresponding raw materials:

[0237] The following compounds can be synthesized by referring to the synthesis method of intermediate BB1 ​​and Example 1 using corresponding reagents:

[0238] The following compounds can be synthesized by referring to the synthesis method of intermediate BB2 and Example 3 using corresponding reagents:

[0239] The following compounds can be synthesized by referring to the synthesis method of intermediate BB1 ​​and Example 2 using corresponding reagents:

[0240] The following compounds can be synthesized by referring to the synthesis method of intermediate BB2 and Example 4 using corresponding reagents:

[0241] The following compounds can be synthesized by referring to the synthesis method of Example 2 using corresponding reagents:

[0242] The following compounds can be prepared by referring to the SFC separation method of Example 8, Example 10, and Example 11 using the corresponding raw materials:

[0243] The following compounds can be synthesized by referring to the synthesis method of Example 9 using corresponding reagents:

[0244] Biological activity test example:

[0245] Biological Activity Example 1: Preparation of SARM1 Enzyme

[0246] Preparation of test compounds:

[0247] The test compounds were prepared at a stock concentration of 200 uM or 10 mM (in DMSO) and further diluted to the desired compound concentration for in vitro SARM1 enzyme assays and inhibitor screening.

[0248] Expression and purification of SARM1 protein

[0249] (1) Plasmid construction

[0250] The gene sequence of SARM1 protein was amplified by PCR, and the PCR amplification product was constructed into the pcDNA3.1-HIS-C plasmid. The above construction was completed by Youbao Bio (Hunan, China).

[0251] (2) Plasmid transfection

[0252] HEK293 cells were cultured in a 10 cm dish using DMEM, and the next day the cells were transfected with 15 μg of pcDNA3.1-SARM1-HIS-C expression plasmid.

[0253] (3) Cell collection and protein extraction

[0254] 48 hours after transfection, cells were collected and the SARM1 protein expressed in the cells was obtained by diginotin cleavage for in vitro activity assays.

[0255] The cells were digested with trypsin-EDTA and centrifuged at 1000 rpm for 5 minutes. The cells were washed once with PBS and then resuspended in PBS containing 100 μM diginotin (0.6 mL PBS / 10 cm2 of cells) for 5 minutes. The cells were then treated with trypan blue and observed under a microscope, indicating that >90% of the cells had been lysed. The cells were then centrifuged at 5000 rpm for 10 minutes, and the supernatant containing SARM1 protein was collected.

[0256] Biological Activity Example 2: In vitro biochemical test for inhibition of SARM1 enzyme activity (IC 50 ):

[0257] First, 200 μM of the compound was added to a 50 mM Tris-HCl (pH 7.5) solution containing 0.05 μg / ml SARM1, and then half was added to an equal volume of 50 mM Tris-HCl (pH 7.5) solution containing 0.05 μg / ml SARM1 and mixed. The drug was diluted 6 times in this way, with final concentrations of 200, 100, 50, 25, 12.5, 6.25, 3.125 μM, or 200, 50, 12.5, 3.125, 0.78, 0.195, 0.049 μM. The control group was incubated at room temperature for 10 minutes without adding inhibitors.

[0258] Then, 50 μM NAD and 50 μM PC6 as substrates and 50 μM NMN as an activator were added to the SARM1 protein after incubation with the inhibitor and reacted at room temperature for 30 minutes. The concentrations of each component are the final concentrations in the reaction system.

[0259] During the reaction, PC6 fluorescence kinetics were monitored using a microplate reader, with excitation and emission wavelengths at 390 nm and 520 nm, respectively. The reaction rate was used to represent protein activity, and the 50% inhibitory concentration (50%) was calculated. Higher reaction rates indicate greater protein activity and lower compound inhibition efficiency.

[0260] The dosage curve of compounds inhibiting SARM1 NAD enzyme activity was prepared using the above method.

[0261] The IC values ​​of these compounds in the assay are provided in Table 1 below. 50 Range:

[0262] IC for inhibition of SARM1 enzyme activity 50 Range: A<0.1μM; B:0.1-1μM; C:1-10μM

[0263] Table 1

[0264] Biological Activity Example 3: Axon Degeneration Index

[0265] For experimental methods, please refer to Gerdts J et al.

[0266] Mouse DRG explant culture

[0267] Dorsal root ganglia were dissected from two-month-old C57BL / 6 male mice and placed in a 35 mm culture dish containing pre-chilled HBSS buffer. Excess tissue from each dorsal root ganglion was removed using a scalpel and dissected into 2-3 explants. These were seeded into wells of a 96-well plate coated with 100 ng / mL Matrigel (Corning). 100 μL of DRG culture medium (Neurobasal medium (Gibco) containing 2% B-27 (Gibco), 1% GlutaMax (Gibco), and 10 ng / mL NGF (MCE)) was added and the plates were cultured in a 37°C incubator for one week. The medium was changed on the fourth day, and experiments were performed on the eighth day.

[0268] Axonal Wallerian degeneration assay

[0269] Dissolve 0.1 μL of 10 mM test compound in 100 μL DRG culture medium to prepare a new medium containing 10 μM test compound. The medium in the 96-well plate was aspirated and added to the new medium containing the test compound. The culture medium was then incubated with the DRG explants in a 37°C incubator for 2 hours to protect them. After incubation, axons were severed in an area adjacent to the tissue explant using a glass electrode. Brightfield images of axons in the same area were taken at 0 and 24 hours using a 20x microscope (Olympus). Axonal integrity was assessed using an in-house developed script.

[0270] The compounds of the present invention exhibited neuroprotective effects in an explant axon Wallerian degeneration assay at a concentration of 10 μM, and were classified according to the degree of protection: extreme protection (A) > 50% and partial protection (B) 20%-50%. The results are listed in Table 2 below.

[0271] Table 2

Claims

1. Compound of formula (I): or its racemate, enantiomer, diastereomer, deuterated substance or pharmaceutically acceptable salt, in, X1 is selected from -N-, -CH-; R1 is independently selected from H, halogen, C1-C3 alkyl, cyano, trifluoromethyl, amino, hydroxy, methoxy, -C(O)NH2, preferably H, F, Cl, methyl R2 is independently selected from H, halogen, C1-C3 alkyl, cyano, trifluoromethyl, amino, hydroxy, methoxy, C1-C3 alkyl-OC(O)-, C1-C3 alkyl-NHC(O)-, preferably H, F, Cl, methyl, cyano; M represents O, S or NH, preferably O; L1 and L3 are each independently selected from -CH2-, -O-, -S- and -NH-, provided that at least one of L1 and L3 is -CH2-; wherein the H of -CH2- and -NH- may be substituted by a C1-C3 alkyl group; L2 is selected from H, -CH3, -CH2-CH3, -OR3, -NHR4; wherein R3 and R4 are independently selected from H and C1-C3 alkyl; L1 can be connected to L2 to form a 4-membered ring, a 5-membered ring or a 6-membered ring, preferably a 5-membered ring; A is selected from 5- to 10-membered aryl, 5- to 10-membered heteroaryl, 5- to 10-membered cycloalkyl, 5- to 10-membered heterocycloalkyl, 3- to 6-membered cycloalkyl and 5- to 10-membered aryl, 3- to 6-membered heterocycloalkyl and 5- to 10-membered aryl, wherein the heteroaryl or heterocycloalkyl may contain 1 or 2 heteroatoms selected from N, O and S, and the 5- to 10-membered aryl, 5- to 10-membered heteroaryl, 3- to 6-membered cycloalkyl and 5- to 10-membered aryl, 3- to 6-membered heterocycloalkyl and 5- to 10-membered aryl may be selected from 1, 2 or 3 substituted by the following substituents: halogen, C1-C3 alkyl, C1-C3 alkoxy, halo-C1-C3 alkyl, halo-C1-C3 alkoxy, cyano; preferably, the substituents are selected from methyl, methoxy, trifluoromethyl, trifluoromethoxy, F, Cl, Br and cyano; preferably, A is selected from phenyl, naphthyl, thienyl and tetrahydrofurophenyl optionally substituted by 1, 2 or 3 substituents selected from the following: F, Cl, Br, cyano, methyl, methoxy, trifluoromethyl.

2. The compound of formula (I) according to claim 1, or its racemate, enantiomer, diastereomer, deuterated form or pharmaceutically acceptable salt thereof, wherein the compound of formula (I) has the following structure: in, R1, R2, L3, A as defined in claim 1; X2 is selected from CH2, O, NH, preferably CH2.

3. The compound of formula (I) according to claim 1, or its racemate, enantiomer, diastereomer, deuterated form or pharmaceutically acceptable salt thereof, wherein the compound of formula (I) has the following structure (III): in, R1, R2, L3, A as defined in claim 1; X2 is selected from CH2, O, NH, preferably CH2.

4. The compound of formula (I) according to claim 1, or its racemate, enantiomer, diastereomer, deuterated form or pharmaceutically acceptable salt thereof, wherein the compound of formula (I) has the following structure (IV): in, R1, R2, M, L1, L2, L3, A are as defined in claim 1.

5. The compound of formula (I) according to claim 1, or its racemate, enantiomer, diastereomer, deuterated form or pharmaceutically acceptable salt thereof, wherein the compound of formula (I) has the following structure (V): in, R1, R2, M, L1, L2, L3, A are as defined in claim 1.

6. The compound of formula (I) according to claim 1, or its racemate, enantiomer, diastereomer, deuterated form or pharmaceutically acceptable salt thereof, wherein the compound of formula (I) has the following structure: in, X1 as defined in claim 1; R5, R6, and R7 are each independently selected from H, F, Cl, Br, cyano, methyl, methoxy, and trifluoromethyl.

7. A compound, or a racemate, enantiomer, diastereomer, deuterated form, or pharmaceutically acceptable salt thereof, wherein the compound has a structure selected from the group consisting of:

8. Use of the compound according to any one of claims 1 to 7, or its racemate, enantiomer, diastereomer, deuterated form, or pharmaceutically acceptable salt thereof, in the preparation of a SARM1 enzyme activity inhibitor or a medicament for treating or preventing a disease or condition associated with SARM1 enzyme activity.

9. Use of the compound according to any one of claims 1 to 7, or its racemate, enantiomer, diastereomer, deuterated form, or pharmaceutically acceptable salt thereof, in the preparation of a medicament for treating or preventing axonal degeneration-related diseases or disorders.

10. Use of the compound according to any one of claims 1 to 7, or its racemate, enantiomer, diastereomer, deuterated form, or pharmaceutically acceptable salt thereof, in the preparation of a medicament for treating or preventing neurodegenerative diseases or neurological diseases or conditions.

11. The method according to claim 8, 9 or 10, wherein the disease or disorder associated with SARM1 enzyme activity, neurodegenerative disease, neurological disease or disorder, or axonal degeneration-related disease or disorder is selected from Alzheimer's disease, Parkinson's disease, multiple sclerosis, amyotrophic lateral sclerosis and peripheral neuropathy.