Cyclic peptide-based PCSK9 inhibitor and use

Abstract

Disclosed in the present invention is a cyclic peptide-based PCSK9 inhibitor, and also provided are a method and use of the inhibitor in the treatment of related diseases.

Description

A cyclic peptide PCSK9 inhibitor and its uses Technical Field

[0001] This invention relates to a cyclic peptide PCSK9 inhibitor. Background Technology

[0002] Cardiovascular disease (CVD) has a high prevalence that increases with age, making it the leading cause of death worldwide. Among these diseases, atherosclerotic cardiovascular disease (ASCVD), including coronary heart disease, myocardial infarction, and stroke, causes far more deaths than cancer and other diseases. The primary culprit behind these diseases is elevated levels of low-density lipoprotein cholesterol (LDL-C).

[0003] Currently, statins are mainly used clinically to lower LDL-C to target levels, thereby preventing and treating ASCVD. These drugs primarily exert their lipid-lowering effect by inhibiting the activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-COA), the rate-limiting enzyme in cholesterol biosynthesis in the liver, and by increasing the expression of the low-density lipoprotein receptor (LDL-R) gene. However, for patients with severe hyperlipidemia, familial hyperlipidemia, or statin intolerance, statins may not lower their LDL-C to the expected level, necessitating consideration of alternative or combination therapy. Long-term use of statins can cause common side effects such as muscle pain and injury, liver damage, and potentially elevated blood sugar levels leading to type 2 diabetes. Developing a safer, more effective, and more adherent drug remains a pressing challenge in the field of lipid-lowering therapy.

[0004] In recent years, the discovery of proprotein convertase subtilisin / kexin type 9 (PCSK9), a novel target for non-statin lipid-lowering drugs, has offered new hope for the treatment of various types of refractory hypercholesterolemia and the management of lipids in patients with very high-risk ASCVD. PCSK9, discovered as early as 2003, is a serine protease synthesized by hepatocytes. PCSK9 can bind to LDL-R to form a complex and degrade LDL-R via the lysosomal pathway, thereby reducing the amount of LDL-R on the cell surface and ultimately leading to elevated plasma LDL-C levels. PCSK9 inhibitors significantly reduce LDL-C levels by inhibiting PCSK9 activity.

[0005] There are two main types of PCSK9 inhibitors on the market globally: monoclonal antibodies (MBAs) and small nucleic acid drugs. The first two MBAs were evolocumab (developed by Amgen / Astera) and alirocumab (developed by Sanofi / Regeneron), both launched in 2015. Both are used for patients with hereditary hypercholesterolemia or high-risk ASCVD, especially those intolerant to statins or with poor response to high-intensity statins. This year, domestically produced MBAs have also been launched: Tolecithin from Innovent Biologics, Inusimiximab from Akeso Biopharma, and Ongoreciximab from Junshi Biosciences. However, these MBAs are all administered via subcutaneous injection every two weeks. For patients requiring long-term treatment and lifelong medication, long-term adherence is low, and some side effects may occur: mild hyperglycemia, and some patients may experience allergic reactions to PCSK9 inhibitors, manifesting as rashes, urticaria, itching, etc. In severe cases, it may cause angioedema, and even anaphylactic shock symptoms such as decreased blood pressure, shortness of breath, and arrhythmia. Some patients may experience side effects such as flu-like symptoms. Inclisiran (Leqvio), developed by Alnylam / Novartis, was launched at the end of 2020. It is a siRNA-based small nucleic acid drug used to treat primary hypercholesterolemia (heterozygous familial and non-familial) or mixed dyslipidemia in adults. Leqvio allows for long-acting dosing (two injections in the first three months, followed by one injection every six months), reducing the frequency of medication and improving patient convenience.

[0006] While all three drugs effectively inhibit PCSK9 activity, they require long-term, repeated injections, which is unacceptable for patients with needle phobia. Furthermore, these drugs are expensive and difficult to store and transport. Due to the widespread mutations and codon degeneracy in human genes, the use of siRNA-based small nucleic acid drugs and antisense oligonucleotide inhibitors carries the risk of off-target effects or ineffectiveness. A recent analysis of over 2.14 million individuals suggests that lower LDL-C is not necessarily better. The study showed a U-shaped relationship between LDL-C levels and all-cause mortality in CAD patients, with both extremely low (<50 mg / dL) and high (≥130 mg / dL) LDL-C levels associated with increased mortality risk. Moreover, several studies have questioned the safety of extremely low LDL-C levels; for example, some studies have shown a doubling of hemorrhagic stroke when LDL-C levels are <70 mg / dL, and others have shown that excessively low LDL-C levels increase the risk of diabetes. Therefore, for long-acting small nucleic acid drugs like injectable ones, once a patient receives an injection, even if their LDL-C drops too low during the course of treatment, it is impossible to stop, which increases the risk of death for the patient.

[0007] In conclusion, although the injectable PCSK9 inhibitors already on the market have shown very good clinical efficacy, they cannot meet the needs of all people with high cholesterol. Therefore, the development of orally administered PCSK9 inhibitors is of great clinical significance.

[0008] Oral PCSK9 inhibitors will provide a more convenient and effective treatment option for people with high cholesterol, and are the goal of many innovative drug development companies. Over the past decade, numerous oral small-molecule PCSK9 inhibitors have been discovered and entered clinical trials, but most have progressed slowly, with none yet reaching Phase 3. Recently, Merck reported on its highly active oral cyclic peptide PCSK9 inhibitor, MK-0616, with excellent Phase 2 clinical results, and quickly launched three Phase 3 clinical trials, bringing new hope for the development of oral PCSK9 inhibitors. Summary of the Invention

[0009] This invention involves in-depth structural analysis, design, synthesis, and screening of PCSK9 inhibitors interacting with PCSK9 and LDL-R complexes, and has successfully identified a series of novel cyclic peptide PCSK9 inhibitors with extremely high activity and superior oral efficacy.

[0010] In one aspect, the present invention provides a compound of formula I or a pharmaceutically acceptable salt thereof:

[0011] R1 is selected from -CH3, -C(O)-(CH2)n1-N +(CH3)3,-C(O)-(CH2CH2O)n2-CH2CH2N + (CH3)3, n1 is selected from integers from 1 to 14, and n2 is selected from integers from 1 to 5.

[0012] Among them, R1' and R1” are independently selected from none, hydrogen, and CH3, respectively.

[0013] R2 is selected from hydrogen, halogen, -SCH3, -SCH2X, and -SCHX2.

[0014] R3 is selected from hydrogen and halogens.

[0015] Where A is selected from:

[0016] R4 and R5 are independently selected from single-chain, C1-C5 straight-chain alkyl, and phenyl groups, respectively. R4 and R5 are not both single-chain, nor are they both phenyl groups.

[0017] Among them, R 41 R 42 Each of the following is independently selected from unsubstituted, hydrogen-containing, deuterium-containing, substituted, or unsubstituted C1-C3 alkyl groups and halogens; or, when R4 is selected from a straight-chain alkyl group, R... 41 R 42 The carbon atom attached to it forms a C3-C5 cycloalkyl group.

[0018] Among them, R 51 R 52 Each of the following is independently selected from unsubstituted, hydrogen-containing, deuterium-containing, substituted, or non-substituted C1-C3 alkyl groups and halogens; or, when R5 is selected from C1-C5 straight-chain alkyl groups, R... 51 R 52 The carbon atom attached to it forms a C3-C5 cycloalkyl group.

[0019] Where n3 is selected from 0 or integers from 1 to 4; n4 is selected from 0 or 1.

[0020] In the substituted C1-C3 alkyl group, the substituent is selected from deuterium or halogen.

[0021] Furthermore, when R1 is not -CH3, either R1' or R1" is selected as none, and the other is selected as hydrogen; or, when R1 is -CH3, both R1' and R1" are -CH3.

[0022] Furthermore, R1 is selected from -C(O)-(CH2)n1-N + (CH3)3,-C(O)-(CH2CH2O)n2-CH2CH2N + (CH3)3. That is, the present invention may be selected from the following structures or their pharmaceutically acceptable salts:

[0023] Furthermore, R1 is selected from -C(O)-(CH2)n1-N + (CH3)3.

[0024] Furthermore, R3 is selected from hydrogen, meaning that the present invention may be selected from the following structures or pharmaceutically acceptable salts thereof:

[0025] Furthermore, R2 is selected from hydrogen, -SCHX2, -SCX3, that is, the present invention can be selected from the following structures or their pharmaceutically acceptable salts:

[0026] The structure or group shown in A can exist in each of the above structural formulas.

[0027] Furthermore, when R4 is a one-key, R 41 R 42 It is none; or, when R5 is a one-key, R 51 R 52 It is none.

[0028] Furthermore, when R4 is a straight-chain alkyl group, R 41 R 42 It is hydrogen, deuterium, substituted or unsubstituted C1-C3 alkyl, halogen, or R 41 R 42 Carbon atoms attached to the same carbon atom and bonded to it form a C3-C5 cycloalkyl group; or, when R5 is a straight-chain alkyl group, R... 51 R 52 It is hydrogen, deuterium, substituted or unsubstituted C1-C3 alkyl, halogen, or R 51 R 52 Carbon atoms attached to the same carbon atom and bonded to it form C3-C5 cycloalkyl groups.

[0029] Furthermore, when R4 is phenyl, R 41 R 42 It is hydrogen, deuterium, substituted or unsubstituted C1-C3 alkyl, or halogen; or, when R5 is phenyl, R 51 R 52 It is hydrogen, deuterium, substituted or unsubstituted C1-C3 alkyl, or halogen.

[0030] Furthermore, R4 and R5 are independently selected from single-chain, C1-C2 straight-chain alkyl, and phenyl groups, respectively. R4 and R5 are not both single-chain, nor are they both phenyl groups.

[0031] Furthermore, R 41 R 42Each of the following is independently selected from: none, H, D, -CH3, halogen, -CH2X, -CHX2, -CX3, -CH2D, -CHD2, and -CD3.

[0032] Furthermore, R 51 R 52 They are independently selected from none, H, D, -CH3, halogen, -CH2X, -CHX2, -CX3, -CH2D, -CHD2, and -CD3, respectively.

[0033] Furthermore, A is selected from one of the following structures:

[0034] (a1)

[0035] (a2)

[0036] (a3)

[0037] Furthermore, R 41 R 42 Each is independently selected from hydrogen, methyl, or halogen.

[0038] Furthermore, R 51 R 52 Each is independently selected from hydrogen.

[0039] Of the above-mentioned halogens, F is preferred.

[0040] Where n4 is selected from 0, n3 is selected from 0 or an integer from 1 to 4. For example, n4 is selected from 0, n3 is selected from 0; n4 is selected from 0, n3 is selected from 1; n4 is selected from 0, n3 is selected from 2; n4 is selected from 0, n3 is selected from 3; n4 is selected from 0, n3 is selected from 4.

[0041] Where n4 is selected from 1, n3 is selected from 0 or an integer from 1 to 4. For example, n4 is selected from 1 and n3 is selected from 0; n4 is selected from 1 and n3 is selected from 1; n4 is selected from 1 and n3 is selected from 2; n4 is selected from 1 and n3 is selected from 3; n4 is selected from 1 and n3 is selected from 4.

[0042] Wherein, n3 is further selected from 0, 1 or 2.

[0043] Furthermore, the structural formula of the compound is selected from one of the following:

[0044] Furthermore, the structural formula of the compound is selected from one of the following:

[0045] In one aspect, the present invention also provides a pharmaceutical composition comprising any of the foregoing compounds or a pharmaceutically acceptable salt thereof.

[0046] The pharmaceutical composition is selected from oral preparations and injectable preparations.

[0047] An oral formulation may be considered a preferred embodiment of the present invention.

[0048] In one aspect, the present invention also provides the use of the above-described compounds or pharmaceutically acceptable salts thereof in the preparation of PCSK9 inhibitors.

[0049] In one aspect, the present invention also provides a method for inhibiting PCSK9, which involves administering an effective dose of any of the aforementioned compounds or a pharmaceutically acceptable salt thereof to a patient.

[0050] In this invention, the patient is a patient with hypercholesterolemia.

[0051] In one aspect, the invention also provides a method of treating diseases by administering to a patient an effective dose of any of the foregoing compounds or a pharmaceutically acceptable salt thereof; said diseases are selected from atherosclerosis, and / or hypercholesterolemia, and / or coronary heart disease, and / or metabolic syndrome, and / or acute coronary syndrome. These diseases are PCSK9-related. Attached Figure Description

[0052] Figure 1. LDL-C concentrations in animals on day 7 (compounds 2451, 2453, 205A, 2421 and compounds 2401, 2460, 2461, 2462, 2455, 2457, 2416)

[0053] Figure 2. LDL-C and TG concentrations in animals on day 17 after 14 days of continuous oral administration. Detailed Implementation

[0054] In the following description, known structural representations are used and include conventional stereochemical symbols for certain asymmetric carbon centers.

[0055] In most cases, the absolute configuration of the example compounds has not been determined, but the absolute configuration has been assigned by simulation to specific example compounds prepared using the same or similar reaction conditions and starting reagents and separated under the same chromatographic conditions, having a known stereochemical configuration (determined by X-ray crystallization). Unless otherwise indicated in the presented information, the specific allocation of configurations presented structurally herein refers to a statement used to identify an excess of a particular stereoisomer of a particular compound prepared herein, and is not necessarily presented herein as a statement of an absolute determination of the stereochemical structure of that compound.

[0056] In this invention, if the stereochemical configuration is not indicated in the structural formula of a compound, it does not mean that the compound does not have a stereochemical configuration.

[0057] When obtaining a mixture of isomers, conventional methods can be used, such as chromatography or crystallization, or the mixture can be separated by using a stereochemically homogeneous starting material for the synthesis or by stereoselective synthesis to prepare individual stereoisomers in a significant percentage enantiomeric excess. Separation of the mixture of stereoisomers can be performed at intermediate steps during compound synthesis, or this separation can be performed on the final racemic product.

[0058] Absolute stereochemistry is determined by X-ray crystallization of the crystallized product or intermediate, which is derivatized as needed using a reagent containing a stereosymmetry center of known configuration. Unless otherwise specified, this invention includes all such isomers, as well as salts, solvates (including hydrates), or solvated salts of such racemates, enantiomers, diastereomers, and mixtures thereof.

[0059] This invention also includes isotopically labeled compounds of the invention that are structurally equivalent to those compounds described herein. The invention is intended to include all suitable isotopic variants of the compounds. Examples of preferred isotopes include, but are not limited to, isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, iodine, fluorine, and chlorine, such as, but not limited to: 2 H, 3 H, 11 C 13 C 14 C 13 N、 15 N、 15 O、 17 O、 18 O、 31 P, 32 P, 35 S, 18 F and 36 Cl、 123 I and 125 I. Additionally, other isotopes can be incorporated using known methods. Deuterium (D) is the main hydrogen isotope found in nature. Enriching deuterium can provide certain therapeutic advantages, such as increasing in vivo half-life or reducing dosage requirements, or can provide compounds suitable for characterizing biological samples.

[0060] A wavy line terminating a regular bond (opposite to two atoms within the connecting structure) indicates the point of bonding with the structure, for example:

[0061] The Z portion is bonded via the Y portion, which terminates at a wavy line. When letter symbols are used to depict the substituent portion, a dash is used to indicate the point of bonding with the indicated substrate; for example, -ZY indicates that the Y portion is bonded via the Z portion.

[0062] When any variable or part is represented in a range, such as (-CH2-) 1-4 It includes the two extreme values ​​of the specified range (i.e., 1 and 4 in the example) and all integer values ​​in between (i.e., 2 and 3 in the example).

[0063] Unless otherwise specified at the point of use, the term "halogen" includes fluorine, chlorine, bromine, and iodine. As used herein, the term "subject" (or "patient") refers to an animal, preferably a mammal, and particularly a human or a non-human animal including livestock and domestic animals, including but not limited to cattle, horses, sheep, pigs, goats, rabbits, cats, dogs, and other mammals requiring treatment.

[0064] In some implementations, the subject is preferably a human. As used herein, “administer” and its variations (e.g., “administer” of a compound) means to provide a subject in need of treatment with the compound or a pharmaceutically acceptable salt thereof.

[0065] In this invention, "-C(O)-" represents a carbonyl group, i.e.

[0066] In this invention, "none" means "absent". For example, when R1' is selected as none, "-NR1R1'R1" is "-NR1R1". When R1" is selected as none, "-NR1R1'R1" is "-NR1R1'".

[0067] In this invention, R1' and R1" cannot both be "absent". That is, when R1' is selected as absent, R1" is selected as hydrogen or methyl. When R1" is selected as absent, R1' is selected as hydrogen or methyl.

[0068] In this invention, n1 can be selected from any integer from 1 to 14, such as 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14. n1 can also be selected from the following ranges: 1-13, 1-12, 1-11, 1-10, 1-9, 1-8, 1-7, 1-6, 1-5, 1-4, 1-3, 1-2, 2-14, 2-13, 2-12, 2-11, 2-10, 2-9, 2-8, 2-7, 2-6, 2-5, 2-4, 2-3, 3-14, 3-13, 3-12, 3-11, 3-10, 3-9, 3-8, 3-7, 3-6, 3-5, 3-4, 4-14, 4-13, 4-12, 4-11, 4-10, 4-9, 4-8, 4-7, 4-6, 4-5, 5-14, 5- 13, 5-12, 5-11, 5-10, 5-9, 5-8, 5-7, 5-6, 6-14, 6-13, 6-12, 6-11, 6-10, 6-9, 6-8, 6-7, 7-14, 7-13, 7-12, 7-11, 7-10, 7-9, 7-8, 8-14, 8-13, 8-12, 8-11, 8-10, 8-9, 9-14, 9-13, 9-12, 9-11, 9-10, 10-14, 10-13, 10-12, 10-11, 11-14, 11-13, 11-12, 12-14, 12-13, 13-14.

[0069] In this invention, n2 can be selected from any integer from 1 to 5, such as 1, 2, 3, 4, 5. n2 can also be selected from the following range: 1-4, 1-3, 1-2, 2-5, 2-4, 2-3, 3-5, 3-4, 4-5.

[0070] In this invention, "one key" refers to a connection key. For example, in -ZYW, when Y is selected as a one key, it becomes -ZW.

[0071] In this invention, "C1-C5 straight-chain alkanes" include at least -CH3, -CH2CH3, -CH2CH2CH3, -CH2CH2CH2CH3, and -CH2CH2CH2CH2CH3.

[0072] In this invention, "C1-C3 alkyl groups" include at least -CH3, -CH2CH3, -CH2CH2CH3, and -CH2(CH3)2.

[0073] In this invention, "C3-C5 cycloalkyl groups" include at least

[0074] In this invention, At least can include

[0075] In this invention, unless otherwise specified, the structure or group represented by A is connected from left to right in the general formula of the compound, as follows: Right now:

[0076] The compound of the present invention shall also include the following structure:

[0077] The compounds of the present invention shall further include the following structures:

[0078] Among them, “E” - "It is a pharmaceutically acceptable anion."

[0079] The term “salt” as used herein and its use in the phrase “pharmaceutically acceptable salt” includes any of the following: acid salts formed with inorganic and / or organic acids, base salts formed with inorganic and / or organic bases, zwitterions, and quaternary ammonium complexes.

[0080] Salts of the compounds of the present invention can be formed by methods known to those skilled in the art, such as by reacting the compounds of the present invention with a certain amount (e.g., a certain equivalent) of an acid or base in a medium such as a salt precipitation medium or an aqueous medium, followed by freeze-drying.

[0081] The compounds of the present invention contain a three-coordinate nitrogen atom, such as a primary, secondary, or tertiary amine moiety, wherein, as is known, the lone pair of electrons present on the nitrogen atom can be protonated with a suitable acid or alkylated with a suitable reagent (e.g., an alkyl bromide) under appropriate reaction conditions to provide a four-coordinate charged nitrogen stabilized by an anion (e.g., a halide ion or a conjugate base) produced in the process. Therefore, the compounds of the present invention can be prepared in the form of a free base or isolated as a quaternary complex or a salt complex. In some cases, it is possible for a suitable acidic proton to be close to the basic nitrogen of a zwitterionic complex. When used herein, the salts of the compounds of the present invention are included within the scope of the compounds of the present invention described herein, whether they are acid salts formed with inorganic and / or organic acids, basic salts formed with inorganic and / or organic bases, salts formed including zwitterionic salts (e.g., where the compound contains a basic moiety, such as, but not limited to, a nitrogen atom, such as an amine, pyridine, or imidazole; and an acidic moiety, such as, but not limited to, both carboxylic acids) and quaternary ammonium complexes.

[0082] Examples of pharmaceutically acceptable acid salts include, but are not limited to, acetates (including trifluoroacetate), adipates, alginates, ascorbic acid salts, aspartate salts, benzoates, benzenesulfonates, hydrogen sulfates, borates, butyrates, citrates, camphorates, camphorsulfonates, cyclopentanepropionates, diglucuronates, dodecyl sulfates, ethanesulfonates, fumarates, glucohepate, glycerophosphates, hemisulfates, heptarates, hexanoates, hydrochlorides, hydrobromide, hydroiodates, 2-hydroxyethanesulfonate, lactates, maleates, methanesulfonates, methyl sulfates, 2-naphthalenesulfonates, nicotinates, nitrates, oxalates, dihydroxynaphthalate, pectates, persulfates, 3-phenylpropionate, phosphates, picrates, neopentanoates, propionates, salicylates, succinates, sulfates, sulfonates (such as those mentioned herein), tartrates, thiocyanates, toluenesulfonates (also known as tosylates, undecanoates, etc.).

[0083] Examples of pharmaceutically acceptable alkali salts include, but are not limited to, ammonium salts, alkali metal salts (such as sodium, lithium, and potassium salts), alkaline earth metal salts (such as calcium and magnesium salts, aluminum salts, and zinc salts), salts containing organic bases (e.g., organic amines) (such as benzathines, diethylamine, dicyclohexylamine, hydrabamines (formed from N,N-bis(dehydrorosinyl)ethylenediamine), N-methyl-D-glucosamine, N-methyl-D-glucosamide, tert-butylamine, piperazine, phenylcyclohexylamine, choline, and tromethamine), and salts containing amino acids (such as arginine and lysine). Basic nitrogen-containing groups can be converted into ammonium ions or quaternized with agents such as: lower alkyl halides (e.g., methyl, ethyl, propyl and butyl chlorides, bromides and iodides), dialkyl sulfates (e.g., dimethyl, diethyl, dibutyl and dipentyl sulfates), long-chain halides (e.g., decyl, dodecyl, tetradecyl and octadecyl chlorides, bromides and iodides), aralkyl halides (e.g., benzyl and phenethyl bromides) and other agents.

[0084] "Pharmaceutically acceptable anions" refers to anions that are suitable for forming pharmaceutically acceptable salts.

[0085] Other examples of pharmaceutically acceptable salts that can be used with this invention include, but are not limited to, fluorides, chlorides, bromides, iodides, and anions.

[0086] This invention provides pharmaceutical compositions comprising one or more compounds of this invention. As used herein, the term "pharmaceutical composition" comprises at least one pharmaceutically active compound and at least one excipient.

[0087] An excipient is any component that adapts a composition to a particular route of administration or facilitates the processing of the composition into a dosage form without exerting an active pharmaceutical effect itself. Typically, a composition contains more than one excipient, depending on the route of administration and the characteristics of the active agent being applied. Examples of excipients that impart properties to a composition that make it easier to handle or process include, but are not limited to, lubricants or compression aids in powdered pharmaceuticals intended to be formulated into tablets, and emulsion stabilizers in a composition (where the active agent is present in the form of an emulsion). Examples of excipients that adapt a composition to a desired route of administration include, for example, but not limited to, absorption enhancers that promote absorption from the gastrointestinal tract for oral administration; and penetration enhancers, such as those for transdermal or transmucosal administration, for example, those for adhesive skin “patches” or compositions for buccal administration.

[0088] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In these solid dosage forms, the active compound is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or with the following components: (a) fillers or compatibilizers, such as starch, lactose, sucrose, glucose, mannitol, and silica; (b) binders, such as hydroxymethyl cellulose, alginate, gelatin, polyvinylpyrrolidone, sucrose, and gum arabic; (c) humectants, such as glycerin; (d) disintegrants, such as agar, calcium carbonate, potato starch or cassava starch, alginate, certain complex silicates, and sodium carbonate; (e) slowing agents, such as paraffin; (f) absorption accelerators, such as quaternary ammonium compounds; (g) wetting agents, such as cetyl alcohol and glyceryl monostearate; (h) adsorbents, such as kaolin; and (i) lubricants, such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium dodecyl sulfate, or mixtures thereof. Buffers may also be included in capsules, tablets, and pills.

[0089] Solid dosage forms such as tablets, sugar pills, capsules, pellets, and granules can be prepared using coatings and shells, such as casings and other materials known in the art. They may contain opacifying agents, and the release of the active compound or compound from such compositions can be delayed in a portion of the digestive tract. Examples of encapsulating components that can be used are polymeric substances and waxes. If necessary, the active compound may also be formed into microcapsules with one or more of the excipients described above.

[0090] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, or tinctures. In addition to the active compound, liquid dosage forms may contain inert diluents conventionally used in the art, such as water or other solvents, solubilizers, and emulsifiers, such as ethanol, isopropanol, ethyl carbonate, ethyl acetate, propylene glycol, 1,3-butanediol, dimethylformamide, and oils, particularly cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil, and sesame oil, or mixtures of these substances.

[0091] In addition to these inert diluents, the composition may also contain auxiliaries such as wetting agents, emulsifiers and suspending agents, sweeteners, flavoring agents and fragrances.

[0092] In addition to the active compound, the suspension may contain suspending agents such as ethoxylated isooctadecyl alcohol, polyoxyethylene sorbitol and dehydrated sorbitol esters, microcrystalline cellulose, aluminum methoxide and agar, or mixtures of these substances.

[0093] Compositions for parenteral injection may comprise physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions, or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Suitable aqueous and non-aqueous carriers, diluents, solvents, or excipients include water, ethanol, polyols, and suitable mixtures thereof.

[0094] Dosage forms of the compounds of the present invention for topical administration include ointments, powders, patches, sprays, and inhalers. The active ingredient is mixed under sterile conditions with a physiologically acceptable carrier and any preservatives, buffers, or propellants that may be necessary.

[0095] The compounds of this invention can also be used in injectable formulations. The injectable formulation is selected from liquid injections (water injections), sterile powders for injection (powder injections), or tablets for injection (referring to molded or machine-compressed tablets made by aseptic methods, dissolved in water for injection before use, for subcutaneous or intramuscular injection).

[0096] As used herein, the term "inhibition," or "antagonism," refers to a substance that provides an effect against PCSK9 in the affected tissue, inhibiting, counteracting, neutralizing, or reducing one or more functions of PCSK9 in the affected tissue. Inhibition or antagonism of one or more of the PCSK9-associated functional properties can be readily determined according to methods known in the art (see, for example, Barak and Webb, 1981, J. Cell Biol. 90:595-604; Stephan & Yurachek, 1993, J. Lipid Res. 34:325-330; and McNamara et al., 2006, Clinica Chimica Acta, 369:158-167.) and those methods described herein. Inhibition or antagonism will achieve a reduction in PCSK9 activity relative to activity observed in the absence of an antagonist or, for example, relative to activity observed when a control antagonist with unrelated specificity is present.

[0097] The present invention provides a method for inhibiting or antagonizing the activity of PCSK9 in a subject, the method comprising administering to the subject a therapeutically effective dose of the compound of the present invention.

[0098] The term "treatment method" refers to a process of action that results in a change in at least one symptom of a disease state that may be preventative or therapeutic in nature. In some embodiments, the present invention relates to a treatment method for a condition associated with and / or attributed to PCSK9 activity, or a condition in which PCSK9 function is contraindicated for a particular individual, the method comprising administering to the individual a therapeutically effective amount of a PCSK9 antagonist compound of formula I or a pharmaceutically acceptable salt thereof. In some embodiments, the condition may be atherosclerosis, hypercholesterolemia, coronary artery disease, metabolic syndrome, acute coronary syndrome, or related cardiovascular diseases and cardiovascular metabolic conditions, or may be a disease state or condition in which PCSK9 activity is contraindicated.

[0099] The treatment methods according to the invention comprise administering to an individual a therapeutically (or preventively) effective amount of the PCSK9-specific antagonist of the invention. The terms "therapeutically effective" or "preventively effective" refer to the amount used, meaning the amount necessary at the intended dose for the desired duration of duration to achieve the desired therapeutic and / or preventive effect. The desired effect may, for example, be the relief, reduction, decrease, or cessation of at least one symptom associated with the treated condition. As those skilled in the art will understand, these amounts will vary depending on various factors, including but not limited to disease state, age, sex, and individual weight, as well as the ability of the PCSK9-specific antagonist to induce the desired effect in the individual. Responses may be documented by in vitro assays, in vivo non-human animal studies, and / or further supported by clinical trials.

[0100] The term "effective amount" or "effective dose" refers to the amount of an active compound sufficient to antagonize PCSK9 and thereby induce the sought response (i.e., a therapeutic response induced in the treatment or management of conditions associated with or affected by PCSK9 function, including but not limited to atherosclerosis, hypercholesterolemia, coronary heart disease, metabolic syndrome, acute coronary syndrome, and related cardiovascular diseases and cardiovascular metabolic conditions in animals or humans).

[0101] The compounds of the present invention can be prepared using readily available starting materials, reagents, and conventional synthetic procedures according to the following reaction schemes and examples or modifications thereof. Known variants may also be utilized in these reactions. Other methods for preparing the compounds of the present invention will be apparent to those skilled in the art in light of the following reaction schemes and examples.

[0102] The following are compound abbreviations: EA: Ethyl acetate; Cbz-Cl: Benzyl chloroformate; THF: Tetrahydrofuran; DCC: N,N'-Dicyclohexylcarbodiimide; DMAP: 4-Dimethylaminopyridine; EtOH: Ethanol; DMF: N,N-Dimethylformamide; DCM: Dichloromethane; TMSOTf: Trimethylsilyl trifluoromethanesulfonate; t-BuOH: Tert-Butanol; BOC: Tert-Butoxycarbonyl; dppb: 1,4-Bis(diphenylphosphine)butane. Ni: Raney nickel; Fmoc-Cl: 9-fluorenylmethylchloroformate; MTBE: Methyl tert-butyl ether; PE: Petroleum ether; DIPEA: N,N-diisopropylethylamine; HATU: 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate; TMSI: Trimethyliodosilane; TFA: Trifluoroacetic acid; LiHMDS: Lithium hexamethyldisilamide; Xantphos: 4,5-bis(diphenylphosphine)-9,9-dimethyloxanthracene; Pd(dppf)Cl2: 1,1'-bis(diphenylphosphine)ferrocene palladium dichloride; Me3N: Trimethylamine

[0103] Example 1: Synthesis of Key Fragments

[0104] 1. Synthesis of Pro fragments

[0105] 1) Preparation of NA2

[0106] Trans-3-hydroxy-L-proline (100 g, 762.6 mmol) was dissolved in 800 mL of 1,4-dioxane. A solution of sodium bicarbonate (141 g, 1677.7 mmol) in 1200 mL of water was added at 0 °C, followed by dropwise addition of benzyl chloroformate (195.14 g, 1143.9 mmol). The reaction was carried out at room temperature for 12 h. The reaction was monitored by LC / MS until completion. The pH was adjusted to 3 by adding dilute hydrochloric acid at 0 °C. The mixture was extracted three times with 400 mL of EA, dried over anhydrous Na₂SO₄, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 184.4 g of off-white solid NA₂, with a yield of 91%.

[0107] 2) Preparation of NA5

[0108] Intermediate NA2 (10 g, 37.7 mmol) was dissolved in 100 mL THF, and DCC (8.54 g, 41.5 mmol) and 10% DMAP (460 mg, 3.77 mmol) were added. After stirring for 0.5 h, EtOH (3.47 g, 75.4 mmol) was added, and the reaction was allowed to proceed until the starting NA2 was completely converted. Then, 100 mL EA was added to dilute the reaction solution, followed by washing with water and saturated brine. The solution was dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 6.73 g of white solid NA5, with a yield of 68%.

[0109] 3) Preparation of NA6

[0110] NA5 (50.0 g, 0.170 mol) was dissolved in 400 mL of DMF, and Cs2CO3 (83.3 g, 0.255 mol) and sodium iodide (1.27 g, 8.5 mmol) were added. The mixture was stirred at room temperature for 5 min, and then tert-butyl bromoacetate (33.2 g, 0.170 mol) was added. The reaction was carried out at 60 °C for 12 h. After the reaction of NA5 was completed under LC / MS control, the reaction solution was filtered. Then, 200 mL of EA and 400 mL of water were added to the filtrate, and the mixture was extracted with a small amount of EA. The organic phases were combined, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 54.1 g of pale yellow oily NA6, with a yield of 78%.

[0111] 4) Preparation of NA7

[0112] NA6 (15.76 g, 24.09 mmol) was dissolved in 100 mL of DCM. 2,6-Dimethylpyridine (8.04 g, 75.00 mmol) was added at 0 °C, followed by the dropwise addition of TMSOTf (15.16 g, 68.18 mmol). The reaction was carried out at room temperature for 2 h. The reaction was monitored by LC / MS. Upon completion of the reaction, the reaction solution was washed twice with 20 mL of water. The organic phase was dried over anhydrous Na2SO4, filtered, and the filtrate was rotary dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to obtain 9.86 g of a yellow oily substance, NA7, with a yield of 62%.

[0113] 2. Synthesis of Trp fragments:

[0114] 1) Preparation of intermediate Trp-1

[0115] The starting material 5-FN-Boc-tryptophan (120 g, 373 mmol) was dissolved in 1200 mL of t-BuOH, 5% DMAP (6.1 g, 18.6 mmol) was added, and Boc₂O (90 g, 447.6 mmol) was added dropwise. The reaction was carried out at 35 °C until the starting material was completely converted. The crude product was obtained by rotary drying to remove tert-BuOH and used directly in the next reaction without purification. LC / MS: [MH] - :377.3.

[0116] 2) Preparation of intermediate Trp-2

[0117] Crude Trp-1 was dissolved in 1000 mL of THF, and allyl methyl carbonate (51.9 g, 447.6 mmol), K₂CO₃ (102.9 g, 746 mmol), 20% dppb (31.78 g, 74.6 mmol), and 10% Pd(CH₃CN)₂Cl₂ (9.66 g, 37.3 mmol) were added. The reaction was carried out at 35 °C under nitrogen protection until the starting material was completely converted. The reaction solution was then added dropwise to 1000 mL of water, extracted three times with 500 mL of EA, and the organic phases were combined, washed twice with saturated brine, dried over anhydrous Na₂SO₄, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 98 g of pale yellow solid Trp-2, with a two-step yield of 63%.

[0118] 3) Preparation of intermediate Trp-3

[0119] The intermediate Trp-2 (98 g, 234 mmol) was dissolved in 1000 mL of DCM, cooled to 0 °C, and p-TsOH·H₂O (210 g, 1.17 mol) was added. The reaction was controlled by LC / MS until the starting material was completely converted. The reaction solution was then dried by rotary evaporation and redissolved in 1000 mL of THF / water (v / v = 2 / 1). Boc₂O (70 g, 351 mmol) was added, and the reaction was continued until the starting material was completely converted. The reaction solution was then dried by rotary evaporation under reduced pressure to remove THF. The crude product was purified by preparative chromatography to obtain 79 g of pale yellow solid Trp-3, with a yield of 93%.

[0120] 3. Synthesis of Phe fragments:

[0121] 1) Preparation of NA3

[0122] The starting material, Boc-L-3-cyanophenylalanine (50.0 g, 0.172 mol), was dissolved in 300 mL of methanol and 50 mL of concentrated ammonia water. Raney-Ni (10.1 g, 0.172 mol) was added, followed by three purgings with hydrogen gas. The reaction was carried out at room temperature for 12 h. After the reaction was completed under LC / MS control, most of the methanol was removed by rotary evaporation. The pH was adjusted to 4 by adding 5% phosphoric acid aqueous solution, and the mixture was stirred for 30 min and then allowed to stand at -20 °C for 12 h. A large amount of crude solid precipitated. The crude product was purified by preparative chromatography to obtain 32.9 g of white solid NA3, with a yield of 65%.

[0123] 2) Preparation of NA4

[0124] NA3 (32.9 g, 0.112 mol) was dissolved in 300 mL of water and 150 mL of ethanol. NaHCO3 (23.5 g, 0.280 mol) was added at room temperature. After this, the temperature was lowered to 2-8 °C, and Fmoc-Cl (30.3 g, 0.117 mol) was added in portions. The temperature was slowly increased, and the reaction was carried out under nitrogen protection for 12 h. The reaction of NA3 was monitored to ensure complete reaction by LC / MS. A portion of the bulk solvent was removed by rotary evaporation, and the aqueous phase was extracted three times with 300 mL of a mixed solution (PE / MTBE, v / v = 1:1). The lower aqueous phase was collected, and the system was cooled to 10-15 °C. The pH was adjusted to 5 with 1 M acetic acid, and then the aqueous phase was extracted three times with 100 mL of EA. The organic phases were combined, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to obtain 42.5 g of white solid NA4, with a yield of 73%.

[0125] 4. Synthesis of NA fragments:

[0126] 1) Preparation of intermediate NA8

[0127] Intermediate NA7 (100 g, 0.28 mol) was dissolved in 800 mL of ethanol, and 10% Pd / C was added. The reaction was carried out under hydrogen balloon conditions until the starting material was completely converted. The reaction solution was filtered and rotary dried to obtain crude intermediate NA8, which could be used directly in the next reaction without purification. LC / MS: [M+H] + :218.1.

[0128] 2) Preparation of intermediate NA9

[0129] Intermediate Trp-2 (78 g, 0.21 mol) was dissolved in 780 mL of DMF, and DIPEA (95 mL, 0.55 mol) was added. The mixture was cooled to 0 °C, and HATU (85.4 g, 0.22 mol) was added. The reaction proceeded until the starting material was completely converted. Intermediate NA8 was then added to the reaction mixture, and the mixture was allowed to rise naturally to room temperature until the starting material was completely converted. The reaction mixture was then added dropwise to 1 M acetic acid solution at 0 °C, extracted three times with 500 mL of EA, washed with saturated brine, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to obtain 43.2 g of a pale yellow oily substance NA9, with a yield of 36%.

[0130] 3) Preparation of intermediate NA10

[0131] The intermediate NA4 (39.6 g, 76.9 mmol) was dissolved in 400 mL of DMF, DIPEA (80 mL, 461.4 mmol) was added, the temperature was lowered to 0 °C, HATU (31.3 g, 80.7 mmol) was added, and the reaction was continued until the starting material was completely converted.

[0132] Intermediate NA9 (43.2 g, 76.9 mmol) was dissolved in 300 mL of CH3CN, cooled to 0 °C, and TMSI (38.4 g, 192.2 mmol) was added dropwise. The reaction proceeded until the starting material was completely converted, and then added dropwise to the above reaction solution. The reaction proceeded until the starting material was completely converted. Then, the reaction solution was added dropwise to 1 M acetic acid solution at 0 °C, extracted three times with 200 mL of EA, washed with saturated brine, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified to obtain 40.2 g of a pale yellow oily substance NA10, with a yield of 54.3%.

[0133] 4) Preparation of intermediate NA11

[0134] Intermediate NA10 (40.2 g, 41.8 mmol) was dissolved in 500 mL of CH3CN, cooled to 0 °C, and piperidine (17.8 g, 209 mmol) was added dropwise. The reaction proceeded until the starting material was completely converted. The reaction solution was then dried by rotary evaporation to remove the solvent and residual piperidine. The crude reaction solution was pulped with PE and filtered before being used directly in the next reaction step. LC / MS: [M+H] + :738.4.

[0135] 5) Preparation of intermediate NA12

[0136] The crude NA11 was dissolved in 2000 mL of DCM, and DIPEA (58 mL, 334.4 mmol) was added. The mixture was cooled to 0 °C, and HATU (81.1 g, 209 mmol) was added. The reaction proceeded until the starting material was completely converted. The reaction solution was added dropwise to an ice-water solution, extracted with EA, washed with saturated brine, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to obtain 15.4 g of white solid NA12, with a two-step yield of 51%.

[0137] 6) Preparation of intermediate NA

[0138] Intermediate NA12 (15.4 g, 21.3 mmol) was dissolved in 800 mL of THF / MeOH / H2O (v / v / v = 6 / 3 / 2), and then 91.6 mL of 1 M LiOH aqueous solution was added dropwise at 0 °C. After reacting for 36 h and the starting material was completely converted, the pH was adjusted to 6 with 1 M acetic acid, and then extracted three times with 200 mL of EA. The extract was dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to obtain 12.6 g of white solid NA, with a yield of 85%.

[0139] 5. Synthesis of NC fragments:

[0140] 1) Preparation of intermediate NC1

[0141] S1 (10.0 g, 33.9 mmol) was dissolved in 100 mL of DMF. DIPEA (6.48 mL, 37.29 mmol) and HATU (14.17 g, 7.3 mmol) were added at 0 °C, and the mixture was stirred in an ice bath for 1 h. Then, 2-methyl-L-proline methyl ester hydrochloride (6.068 g, 33.9 mmol) was added, and the mixture was stirred in an ice bath for 10 min. The mixture was then stirred at room temperature for 12 h. The reaction was stopped when S1 was completely controlled by LC / MS. The mixture was extracted three times with 30 mL of EA. The organic phase was dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 10.88 g of yellow oily substance NC1, with a yield of 77%.

[0142] 2) Preparation of intermediate NC2

[0143] Dissolve NC1 (10.88 g, 25.9 mmol) in 100 mL of DCM. Add 25 mL of TFA at 0 °C and stir continuously in an ice bath until LC / MS shows that NC1 has completely reacted. Adjust the pH to 7-8 with DIPEA in an ice bath, and then proceed directly to the next step of the reaction. LC / MS: [M+H] + :321.4.

[0144] 3) Preparation of intermediate NC3

[0145] [Corrected according to Rule 91 04.03.2025] N-Boc-tert-butoxycarbonyl-O-tert-butyl-L-threonine (7.124 g, 25.9 mmol) was dissolved in 70 mL of DCM. DIPEA (5 mL, 28.5 mmol) and HATU (11.06 g, 28.5 mmol) were added at 0 °C. The mixture was stirred for 1 h in an ice bath. Then, it was added to the pH-adjusted NC2 solution. The mixture was stirred at 0 °C for 10 min and then brought to room temperature. The mixture was stirred for 4 h in room temperature. 100 mL of water was added to the reaction solution. The mixture was extracted three times with 100 mL of DCM. The organic phases were combined and dried over anhydrous Na2SO4. The solution was filtered and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 10.42 g of yellow oily substance NC3, with a yield of 70%.

[0146] 4) Preparation of intermediate NC

[0147] NC3 (10.42 g, 18.06 mmol) was dissolved in 660 mL of THF / MeOH / H2O (v / v / v = 6:3:2). An aqueous solution of LiOH (3.79 g, 90.3 mmol) was added at 0 °C. After the reaction of the starting material NC3 was complete, the pH was adjusted with 1 M acetic acid solution. The organic phase was then dried over anhydrous Na2SO4, filtered, and the filtrate was rotary dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to obtain 6.09 g of white solid NC, with a yield of 60%.

[0148] Table 1. Structural identification data of some compounds in the key fragment synthesis pathway.

[0149] Example 2 Synthesis of intermediate BD

[0150] 1. Synthesis of intermediate BD02:

[0151] 1) Preparation of intermediate ND-OTos

[0152] TsCl (2.9 g, 383.6 mmol) and DMAP (4.29 g, 34.8 mmol) were dissolved in 300 mL of DCM. Triethylamine (52.8 g, 523.2 mmol) and 4-penten-1-ol (30 g, 348.8 mmol) were added separately at 0 °C. The mixture was then heated to room temperature and stirred for 3 h. The reaction of 4-penten-1-ol was monitored by LC / MS until complete. The mixture was extracted three times with 150 mL of DCM. The organic phases were combined and washed twice with 500 mL of saturated brine. The organic phase was then dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 83 g of yellow oily ND-OTos, with a yield of 99%.

[0153] 2) Preparation of intermediate ND3

[0154] ND-OTos (52.37 g, 217.2 mmol) and ND2 (60 g, 239 mmol) were dissolved in 800 mL of CH3CN. K2CO3 (89.96 g, 651.6 mmol) was added at 0 °C, and the mixture was then heated to 85 °C and stirred for 12 h until the ND-OTos reaction was complete as controlled by LC / MS. The K2CO3 was filtered off, and the mixture was washed with 500 mL of EA. The filtrate was dried under reduced pressure by rotary drying to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 35 g of a yellow oily substance, ND3, with a yield of 50%.

[0155] 3) Preparation of intermediate BD01-1

[0156] Intermediate ND3 (13 g, 40.8 mmol) and methyl 4-carboxyphenylacetate (8.72 g, 53 mmol) were dissolved in 100 mL of DMF. DIPEA (21.3 mL, 122.4 mmol) was added at 0 °C, and the mixture was stirred for 15 min. Then, HATU (18.6 g, 49 mmol) was added, and the mixture was slowly brought to room temperature and reacted for another 5 h. LC / MS analysis showed that intermediate ND3 had completely reacted. 200 mL of water and 200 mL of EA were added to the reaction mixture, and the mixture was extracted three times with 100 mL of EA. The organic phases were combined, washed twice with saturated brine, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 16.8 g of a yellow oily substance, BD01-1, with a yield of 83%.

[0157] 4) Preparation of intermediate BD01-2

[0158] BD01-1 (12.5 g, 25.2 mmol) was dissolved in 100 mL of DCM, and 20 mL of TFA was added dropwise at 0 °C. The mixture was stirred for 2 h. LC / MS monitoring showed that BD01-1 reacted completely. The reaction solution was dried under reduced pressure to obtain crude BD01-2, which could be used directly in the next reaction without purification. LC / MS: [M+H] + :395.2.

[0159] 5) Preparation of intermediate BD01

[0160] The crude product BD01-2 was dissolved in 160 mL of THF / H2O (v / v = 1:1), and an aqueous solution of LiOH (10.6 g, 252 mmol) was added at 0 °C. The mixture was slowly brought back to room temperature and the reaction continued for 5 h. After the reaction was confirmed to be complete by LC / MS, the pH was adjusted to 5-6 with 1 M acetic acid. Then, THF was removed by vortexing under reduced pressure. The remaining aqueous phase was purified by preparative chromatography to obtain 8.6 g of white solid BD01, with a yield of 86%.

[0161] 2. Synthesis of intermediate BDO2:

[0162] 1) Preparation of intermediate BD02-1

[0163] Methyl 4-carboxyphenylacetate (40.0 g, 206.2 mmol) was dissolved in 200 mL of dry THF solution, purged three times with nitrogen, and 1 M LiHMDS (412.4 mL, 412.4 mmol) was slowly added dropwise at 0 °C. The reaction was carried out at 0 °C for 2 h, followed by the slow addition of CH3I (12.44 mL, 195.8 mmol). The mixture was then gradually brought to room temperature and reacted for 12 h at room temperature. 0.1 M acetic acid aqueous solution was then added sequentially to the reaction mixture, and the mixture was extracted three times with 100 mL of EA. The organic phases were combined, dried over anhydrous Na2SO4, filtered, and the filtrate was rotary dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to obtain 9.86 g of a pale yellow oily substance, BD02-1, with a yield of 23%.

[0164] 2) Preparation of intermediate BD02-2

[0165] For the synthesis of BD02-2, please refer to the synthesis method of BD01-1. The crude product was purified by silica gel column chromatography to obtain 14.2 g of a yellow oily substance, BD02-2, with a yield of 87%.

[0166] 3) Preparation of intermediate BD02-3

[0167] For the synthesis of BD02-3, please refer to the synthesis method of BD01-2. The crude product was obtained by rotary drying of the organic solvent under reduced pressure. The crude product can be used directly in the next reaction without purification. LC / MS: [M+H] + :409.4.

[0168] 4) Preparation of intermediate BDO2

[0169] For the synthesis of BD02, please refer to the synthesis method of BD01. THF was removed under reduced pressure, and the remaining aqueous phase was purified by preparative chromatography to obtain 10.4 g of white solid BD02, with a yield of 66%.

[0170] 3. Synthesis of intermediate BD03:

[0171] 1) Preparation of intermediate BD03-1

[0172] Methyl 4-carboxyphenylacetate (20.0 g, 103.1 mmol) was dissolved in 100 mL of dry THF. The solution was purged with nitrogen three times. 1 M LiHMDS (464.0 mL, 464.0 mmol) was slowly added dropwise at 0 °C. After reacting for 1 h, CH3I (17.97 mL, 288.7 mmol) was slowly added dropwise to the reaction solution. The mixture was then gradually brought back to room temperature and reacted for 12 h at room temperature. The reaction was monitored by LC / MS. After the reactants had completely reacted, 0.1 M acetic acid aqueous solution was added sequentially to the reaction system. The mixture was extracted three times with 50 mL of EA. The organic phases were combined, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to obtain 8.0 g of a pale yellow oil, BD03-1, with a yield of 35%.

[0173] 2) Preparation of intermediate BD03-2

[0174] For the synthesis of BD03-2, please refer to the synthesis method of BD01-1. Purification of the crude product by silica gel column chromatography yielded 9.41 g of a yellow oily substance, BD03-2, in 80% yield.

[0175] 3) Preparation of intermediate BD03-3

[0176] For the synthesis of BD03-3, please refer to the synthesis method of BD01-2. Rotate to dry the reaction solution; the crude product can be used directly in the next reaction without purification. LC / MS: [M+H] + :423.4.

[0177] 4) Preparation of intermediate BDO3

[0178] For the synthesis of BD03, please refer to the synthesis method of BD01. THF was removed under reduced pressure, and the remaining aqueous phase was purified by preparative chromatography to give 7.72 g of a white solid, with a yield of 84%.

[0179] 4. Synthesis of intermediate BD04:

[0180] 1) Preparation of intermediate BD04-1

[0181] Pd(OAc)₂ (850 mg, 3.75 mmol) and Xantphos (6.5 g, 11.26 mmol) were dissolved in 95 mL of DMF, purged three times with nitrogen, and then formic acid (19.6 mL, 525.85 mmol), S1 (19 g, 75.12 mmol), and Et₃N (20.8 mL, 150.24 mmol) were added sequentially. A 100 mL solution of DCC (31 g, 150.24 mmol) in DMF was added dropwise at 0 °C. The reaction temperature was then slowly increased to 80 °C. After 18 h, the amount of BD₀₄⁻¹ was very small in the LC / MS control. The reaction solution was cooled to room temperature, purged with nitrogen, and Pd(OAc)₂ (850 mg, 3.75 mmol), formic acid (9.8 mL, 262.92 mmol), and Et₃N (10.4 mL, 75.12 mmol) were added. DCC was added dropwise in an ice bath. The reaction continued at 80℃ for 24 hours. LC / MS analysis showed that the reaction of starting material S1 was complete. The reaction solution was diluted with 50 mL of EA, filtered, and the filter cake was washed with 30 mL of EA. Water was added to 100 mL of the filtrate, and the mixture was extracted twice more with 50 mL of EA. The organic phases were combined, washed twice with brine, and then dried over anhydrous Na₂SO₄. The mixture was filtered, and the filtrate was dried under reduced pressure using a rotary dryer to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 7.3 g of white solid BDO₄⁻, with a yield of 44%.

[0182] 2) Preparation of intermediate BD04-2

[0183] For the synthesis of BD04-2, please refer to the synthesis method of BD01-1. The crude product was purified by silica gel column chromatography to obtain 11.4 g of white solid BD04-2, with a yield of 66%.

[0184] 3) Preparation of intermediate BD04-3

[0185] For the synthesis of BD04-3, please refer to the synthesis method of BD01-2. The organic solvent was evaporated under reduced pressure to obtain the crude product, which can be used directly in the next reaction without purification. LC / MS: [M+H] + :421.3.

[0186] 4) Preparation of intermediate BD04

[0187] For the synthesis of BD04, please refer to the synthesis method of BD01. THF was removed under reduced pressure, and the remaining aqueous phase was purified by preparative chromatography to give 6.2 g of a white solid, with a yield of 72%.

[0188] 5. Synthesis of intermediate BD16:

[0189] 1) Preparation of intermediate BD16-1

[0190] SM1 (27 g, 121.59 mmol), Xantphos (7 g, 12.15 mmol), and K2CO3 (67 g, 406.37 mmol) were dissolved in 270 mL of 1,4-dioxane. The mixture was purged with nitrogen three times. Then, Pd(PPh3)4 (7 g, 6.07 mmol) and CuI (1.15 g, 6.07 mmol) were added, and the mixture was purged with nitrogen three more times. The reaction was carried out in an oil bath at 80 °C for 12 h. After the reaction was complete, the mixture was diluted with water to 800 mL, extracted three times with 150 mL of EA, and the organic phases were combined. The organic phases were then dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to obtain 10.3 g of a yellow oily substance BD16-1, with a yield of 28%.

[0191] 2) Preparation of intermediate BD16-2

[0192] BD16-1 (7.59 g, 25.3 mmol) was dissolved in DCM (40 mL), and TFA (8 mL, 104.5 mmol) was added under ice bath conditions. The mixture was then reacted at room temperature for 2 h until BD16-1 was completely reacted as indicated by LC / MS. The reaction was then stopped, and DCM and TFA were removed by vortexing under reduced pressure to obtain 6.1 g of crude yellow oil, BD16-2. The crude product was used directly in the next reaction without further purification.

[0193] 3) Preparation of intermediate BD16-3

[0194] For the synthesis of BD16-3, please refer to the synthesis method of BD01-1. The crude product was purified by silica gel column chromatography to obtain 8.71 g of a yellow oily substance, BD16-3, in 64% yield.

[0195] 4) Preparation of intermediate BD16-4

[0196] For the synthesis of BD16-4, please refer to the synthesis method of BD01-2. Rotary drying of the reaction solvent yielded 7.52 g of a yellow oily substance, BD16-4. The product was used directly in the next reaction without purification. LC / MS: [M+H] + :445.2.

[0197] 5) Preparation of intermediate BD16

[0198] For the synthesis of BD16, please refer to the synthesis method of BD02. THF was removed by vortexing under reduced pressure, and the remaining aqueous phase was purified by preparative chromatography to obtain 6.9 g of white solid BD16, with a yield of 97%.

[0199] 6. Synthesis of intermediate BD10:

[0200] 1) Preparation of intermediate BD10-1

[0201] 3-Iodobenzoate (15.0 g, 57.24 mmol) was dissolved in 150 mL of DMF, and tert-butyl acrylate (36.68 g, 286.2 mmol), palladium acetate (1.3 g, 5.724 mmol), triethylamine (17.38 g, 171.72 mmol), and tetrabutylammonium iodide (2.11 g, 5.724 mmol) were added. The reaction was carried out at 120 °C for 3 h. The reaction was confirmed by LC / MS. The reaction solution was filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 12.91 g of a yellow oily substance, with a yield of 86%.

[0202] 2) Preparation of intermediate BD10-2

[0203] BD10-1 (10.1 g, 28.88 mmol) was dissolved in 100 mL of methanol, and 2.0 g of 10% Pd / C hydrogen was added to replace the hydrogen three times. The reaction was carried out at room temperature for 12 h. The reaction was confirmed to be complete by LC / MS. The reaction solution was filtered, and the filtrate was dried under reduced pressure by rotary drying to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 9.26 g of yellow oil, with a yield of 91%.

[0204] 3) Preparation of intermediate BD10-3

[0205] BD10-2 (8.0 g, 30.29 mmol) was dissolved in a mixed solvent of 75 mL acetonitrile and 3 mL water. LiBr (15.78 g, 181.74 mmol) and DIPEA (23.49 g, 181.74 mmol) were added, and the mixture was reacted at 85 °C for 3 h. The reaction mixture was checked by LC / MS to confirm complete reaction. The pH of the reaction solution was then adjusted to 5-6 with 1 M acetic acid aqueous solution. The mixture was extracted three times with 30 mL EA, dried over anhydrous Na2SO4, and the organic solvent was removed by rotary drying. The crude product was purified by silica gel column chromatography to obtain 6.7 g of a yellow oily substance, with a yield of 88%.

[0206] 4) Preparation of intermediate BD10-4

[0207] For the synthesis of BD10-4, please refer to the synthesis method of BD01-1. The organic solvent was dried by rotary drying, and the crude product was purified by silica gel column chromatography to obtain 10 g of a yellow oily substance, BD10-4, with a yield of 83%.

[0208] 5) Preparation of intermediate BD10

[0209] For the synthesis of BD10, please refer to the synthesis method of BD01. THF was removed under reduced pressure, and the remaining aqueous phase was purified by preparative chromatography to obtain 6.79 g of the yellow oily substance BD10, with a yield of 96%.

[0210] 7. Synthesis of intermediate BD12:

[0211] 1) Preparation of intermediate BD12-3

[0212] BD10-2 (49 g, 185.4 mmol) was dissolved in 500 mL of DCM, and 100 mL of TFA was added in an ice bath. The reaction was incubated at room temperature for 2 h. The reaction was checked by LC / MS to confirm completion. The reaction solution was dried by rotary evaporation and proceeded directly to the next step of the reaction. LC / MS: [M+H] + : 209.1.

[0213] 2) Preparation of intermediate BD12-4

[0214] The crude BD12-3 was dissolved in 500 mL of LDM, and DIPEA (65.35 g, 505.65 mmol) and HATU (76.9 g, 202.26 mmol) were added in an ice bath. After reacting for 30 min, ND3 (53.68 g, 168.55 mmol) was added, and the reaction was carried out at room temperature for 3 h. The reaction was stopped by LC / MS. The reaction solution was added dropwise to water, and the mixture was extracted three times with 300 mL of EA. The organic phase was washed twice with a semi-saturated ammonium chloride aqueous solution and twice with saturated brine. The organic phases were combined, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 87 g of yellow oil, with a yield of 92%.

[0215] 3) Preparation of intermediate BD12-5

[0216] BD12-4 (86.5 g, 168.55 mmol) was dissolved in 800 mL of LCM, and 160 mL of TFA was added in an ice bath. The reaction was incubated at room temperature for 2 h. The reaction was checked by LC / MS to confirm completion. The reaction solution was dried by rotary evaporation and proceeded directly to the next step of the reaction. LC / MS: [M+H] + : 409.4.

[0217] 4) Preparation of intermediate BD12

[0218] For the synthesis of BD12, please refer to the synthesis method of BD01. THF was rotary evaporated under reduced pressure, and the remaining aqueous phase was purified by preparative chromatography to obtain 57.1 g of a yellow oily substance, BD12, in 85% yield.

[0219] 8. Synthesis of intermediate BD21:

[0220] 1) Preparation of intermediate BD21-1

[0221] S1 (37.54 g, 173.8 mmol) was dissolved in 220 mL of THF. 60% NaH (5.55 g, 138.8 mmol) was added in portions at 0 °C, and the reaction was carried out at 0 °C for 2 h. Then, S2 (22.0 g, 115.7 mmol) was added dropwise, and the reaction was slowly brought back to room temperature for 12 h. The reaction of S1 was monitored by LC / MS until complete. The pH was adjusted to 6-7 by adding 0.1 M acetic acid solution. The mixture was extracted three times with 150 mL of EA. The organic phases were combined, dried over anhydrous Na2SO4, filtered, and the filtrate was rotary dried under reduced pressure to obtain 19.78 g of crude yellow oil, BD21-1. The crude product was used directly in the next reaction without further purification. LC / MS: [M+H] + :387.2.

[0222] 2) Preparation of intermediate BD21-2

[0223] The crude BD21-1 (16.50 g, 42.7 mmol) was dissolved in 300 mL of THF, and 300 mL of NaOH (2.66 g, 63.8 mmol) aqueous solution was added at 0 °C. The reaction was slowly brought back to room temperature. After 12 h, the reaction of BD21-1 was completed under LC / MS control. 0.1 M acetic acid solution, water, and EA were added to the reaction solution to adjust the pH to 6. Extraction was performed three times with 150 mL of EA. The organic phases were combined, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 10.37 g of transparent oily BD21-2, with a yield of 66%.

[0224] 3) Preparation of intermediate BD21-3

[0225] For the synthesis of BD21-3, please refer to the synthesis method of BD01-1. The crude product was purified by silica gel column chromatography to obtain 11.43 g of a yellow oily substance, BD21-3, in 82% yield.

[0226] 4) Preparation of intermediate BD21

[0227] BD21-3 (11.28 g, 16.8 mmol) was dissolved in DCM (100 mL), and TFA (30 mL, 391.8 mmol) was added at 0 °C. The mixture was then stirred at room temperature for 24 h until the reaction was complete as controlled by LC / MS. The reaction was stopped, and DCM and TFA were removed by vortexing under reduced pressure. Excess sodium bicarbonate aqueous solution was added to adjust the pH to 8-9. The aqueous phase was purified by preparative chromatography to give 4.25 g of white solid BD21, with a yield of 61%.

[0228] 9. Synthesis of intermediate BD07:

[0229] 1) Preparation of intermediate BD07-1

[0230] 15.0 g (58.3 mmol) of tert-butyl 2-bromobenzoate was dissolved in 100 mL of 1,4-dioxane and 100 mL of water. K₂CO₃ (16.1 g, 116.6 mmol), Pd(dppf)Cl₂ (4.26 g, 5.83 mmol), and pinacol 3-methoxycarbonylphenylboronic acid (16 g, 61.2 mmol) were added, and the mixture was purged with nitrogen four times. The reaction was carried out at 80 °C for 3 h. The reaction was monitored by LC / MS until complete. Water and EA were added to the reaction system, followed by extraction three times with 100 mL of EA. The organic phases were combined, washed twice with saturated brine, dried over anhydrous Na₂SO₄, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 18 g of a yellow oily substance BD07-1, with a yield of 98%.

[0231] 2) Preparation of intermediate BD07-2

[0232] Intermediate BD07-1 (18 g, 57.7 mmol) was dissolved in 180 mL of DCM. TMSI (17.3 g, 86.5 mmol) was added at 0 °C, and the mixture was stirred at room temperature for 3 h until the reaction of intermediate BD07-1 was complete as indicated by LC / MS. The reaction was quenched with water at 0 °C, dried under vacuum, and the product was removed from the DCM. The mixture was then extracted three times with 50 mL of EA. The organic phases were combined, washed twice with saturated brine, dried over anhydrous Na₂SO₄, and concentrated to obtain 16.8 g of a yellow oily crude product BD07-2, which was used directly in the next reaction without further purification. LC / MS: [MH] - : 255.0.

[0233] 3) Preparation of intermediate BD07-3

[0234] Intermediate ND3 (14 g, 43.9 mmol) and intermediate BD07-2 (16.8 g, 65.85 mmol) were dissolved in 200 mL of DMF. DIPEA (30.5 mL, 175.6 mmol) was added at 0 °C, and the mixture was stirred in an ice bath for 15 min. Then, HATU (25 g, 65.85 mmol) was added, and the mixture was slowly allowed to return to room temperature for 5 h. LC / MS monitoring of ND3 showed complete reaction. The mixture was then extracted three times with 500 mL of water, 500 mL of EA, and 300 mL of EA. The organic phases were combined, washed twice with saturated brine, dried over anhydrous Na2SO4, and the organic solvent was removed by rotary drying. The crude product was purified by silica gel column chromatography to obtain 24 g of a yellow oily intermediate BD07-3, with a yield of 98%. LC / MS: [M+H] + : 557.4.

[0235] 4) Preparation of intermediate BD07-4

[0236] Intermediate BD07-3 (12 g, 21.58 mmol) was dissolved in 100 mL of DCM. 20 mL of TFA was added dropwise at 0 °C, and stirring continued for 2 h. LC / MS monitoring showed complete reaction of BD07-3. The organic solvent was dried by rotation to obtain crude BD07-4. LC / MS: [M+H] + : 457.4.

[0237] 5) Preparation of intermediate BD07

[0238] The crude BD07-4 was dissolved in 160 mL of THF / H2O (v / v = 1:1), and an aqueous solution of LiOH (9.06 g, 215.8 mmol) was added at 0 °C. The mixture was slowly brought back to room temperature and reacted for 5 h. The reaction was confirmed to be complete by LC / MS. The THF was dried by rotary evaporation, and the remaining aqueous phase was purified by preparative chromatography to obtain 7.5 g of white solid BD07, with a yield of 78%.

[0239] Synthesis of intermediate E

[0240] 1. Synthesis of intermediate NE2C:

[0241] 1) Preparation of intermediate NE2C-1

[0242] 10 g (51.28 mmol) of tert-butyl 2-bromoacetate was dissolved in 100 mL of acetonitrile. 2 M trimethylamine methanol solution (76.92 mL, 153.85 mmol) was added at room temperature, and the reaction was carried out at 50 °C for 12 h. After complete reaction by LC / MS, the methanol and acetonitrile solvent in the reaction system were dried by rotary evaporation to obtain the crude product. The crude product did not require purification and was directly used in the next reaction. LC / MS: [M] + :174.17.

[0243] 2) Preparation of intermediate NE2C

[0244] The crude NE2C-2 was dissolved in DCM, and 4M 1,4-dioxane hydrochloric acid solution (64 mL, 256.41 mmol) was added at room temperature. After stirring at room temperature, the reaction was monitored by LC / MS until the NE2C-2 was completely reacted. The crude product was then dried by rotary drying of DCM. The crude product was purified by preparative chromatography to obtain 8.53 g of white solid NE2C. The two-step yield was 84%.

[0245] 2. Synthesis of intermediate NE4C:

[0246] 1) Preparation of intermediate NE4C-1

[0247] For the synthesis of NE4C-1, please refer to the synthesis method of NE2C-1, except that tert-butyl 4-bromopropionate is used instead of tert-butyl 2-bromoacetate. LC / MS: [M] + :202.16.

[0248] 2) Preparation of intermediate NE4C

[0249] For the synthesis of NE4C, please refer to the synthesis method of NE2C, except that NE4C-1 is used instead of NE2C-1. The crude product was purified by preparative chromatography to obtain 7.12 g of white solid NE4C, with a two-step yield of 70%.

[0250] 3. Synthesis of intermediate NE5C:

[0251] 1) Preparation of intermediate NE5C-1

[0252] For the synthesis of NE5C-1, please refer to the synthesis method of NE2C-1, except that tert-butyl 3-bromopentanoate is used instead of tert-butyl 2-bromoacetate. LC / MS: [M] + :216.18.

[0253] 2) Preparation of intermediate NE5C

[0254] For the synthesis of NE5C, please refer to the synthesis method of NE2C, except that NE5C-1 is used instead of NE2C-1. The crude product was purified by preparative chromatography to obtain 7.53 g of white solid NE5C, with a two-step yield of 74%.

[0255] 4. Synthesis of intermediate NE6C:

[0256] 1) Preparation of intermediate NE6C-1

[0257] For the synthesis of NE6C-1, please refer to the synthesis method of NE2C-1, except that tert-butyl 3-bromohexanoate is used instead of tert-butyl 2-bromoacetate. LC / MS: [M] + :230.2.

[0258] 2) Preparation of intermediate NE6C

[0259] For the synthesis of NE6C, please refer to the synthesis method of NE2C, except that NE6C-1 is used instead of NE2C-1. The crude product was purified by preparative chromatography to obtain 6.79 g of white solid NE6C, with a two-step yield of 67%.

[0260] 5. Synthesis of intermediate NE7C:

[0261] 1) Preparation of intermediate NE7C-1

[0262] For the synthesis of NE7C-1, please refer to the synthesis method of NE2C-1, except that tert-butyl 3-bromoheptanoate is used instead of tert-butyl 2-bromoacetate. LC / MS: [M] + :244.2.

[0263] 2) Preparation of intermediate NE7C

[0264] For the synthesis of NE7C, please refer to the synthesis method of NE2C, except that NE7C-1 is used instead of NE2C-1. The crude product was purified by preparative chromatography to obtain 7.09 g of white solid NE7C, with a two-step yield of 72%.

[0265] 6. Synthesis of intermediate NE15C:

[0266] 1) Preparation of intermediate NE15C-1

[0267] For the synthesis of NE15C-1, please refer to the synthesis method of NE2C-1, except that tert-butyl 3-bromopentadecanoate is used instead of tert-butyl 2-bromoacetate. LC / MS: [M] + :356.3.

[0268] 2) Preparation of intermediate NE15C

[0269] For the synthesis of NE15C, please refer to the synthesis method of NE2C, except that NE15C-1 is used instead of NE2C-1. The crude product was purified by preparative chromatography to obtain 8.31 g of white solid NC15C, with a two-step yield of 82%.

[0270] 7. Synthesis of intermediate E1O:

[0271] 1) Preparation of intermediate E1O-1

[0272] SM1 (9.0 g, 67.59 mmol) was dissolved in 180 mL of THF, followed by the addition of K2CO3 (46.6 g, 337.96 mmol) and then CH3I (21 mL, 337.96 mmol). The tube was sealed and the reaction was allowed to proceed overnight at room temperature. The reaction was monitored by LC / MS until complete. The reaction solution was filtered, the filter cake was washed with THF, and the filtrate was dried by rotary evaporation to obtain 22 g of crude product E1O-1. LC / MS: [M] + : 190.07.

[0273] 2) Preparation of intermediate E1O

[0274] Crude E1O-1 (22 g, 69.36 mmol) was dissolved in 200 mL of H2O, and NaOH (4.16 g, 69.36 mmol) was added at 0 °C. The reaction was allowed to proceed for 10 min at room temperature. The reaction was monitored by LC / MS until complete. The reaction solution was purified to yield 8.20 g of white solid E1O, with a yield of 57%.

[0275] 8. Synthesis of intermediate E2O:

[0276] 1) Preparation of intermediate E2O-1

[0277] For the synthesis of E2O-1, please refer to the synthesis method of E1O-1, except that SM2 is used instead of SM1. LC / MS: [M] + :276.2.

[0278] 2) Preparation of intermediate E2O

[0279] For the synthesis of E2O, please refer to the synthesis method of E1O, except that E2O-1 is used instead of E1O-1. The crude product was purified by preparative chromatography to obtain 7.68 g of white solid E2O, with a two-step yield of 57%.

[0280] 9. Synthesis of intermediate E3O:

[0281] 1) Preparation of intermediate E3O-1

[0282] For the synthesis of E3O-1, please refer to the synthesis method of E1O-1, where SM3 is used instead of SM1. LC / MS: [M] + :320.1.

[0283] 2) Preparation of intermediate E3O

[0284] For the synthesis of E3O, please refer to the synthesis method of E1O, except that E3O-1 is used instead of E3O-1. The crude product was purified by preparative chromatography to obtain 8.66 g of white solid E2O, with a two-step yield of 68%.

[0285] Table 2. Structural identification data in intermediate synthesis pathways.

[0286] Example 3 Synthesis of the target molecule

[0287] 1. Synthesis of target molecule 2401

[0288] 1) Preparation of intermediate T2401-1

[0289] Intermediate NC (4.13 g, 7.3 mmol) was dissolved in 40 mL of EA. DIPEA (3.82 mL, 21.9 mmol) and HATU (3.04 g, 8.0 mmol) were added at 0 °C. After reacting for 1 h, intermediate BD01 (3.5 g, 9.1 mmol) was dissolved in 30 mL of DMF and added dropwise to the above reaction system. The reaction was carried out at 0 °C for 4 h. Then, 50 mL of water and 50 mL of EA were added to the reaction solution, and the pH was adjusted to 6 with 0.1 M acetic acid solution. The mixture was extracted three times with 30 mL of EA. The organic phases were combined, washed twice with 0.1 M acetic acid solution, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to obtain 5.4 g of white solid T2401-1, with a yield of 79%.

[0290] 2) Preparation of intermediate T2401-2

[0291] NB (1.97 g, 5.8 mmol) was dissolved in 20 mL of DCM, and p-TsOH·H2O (3.32 g, 17.5 mmol) was added at 0 °C. The reaction was carried out at 0 °C for 4 h. LC / MS showed that NB reacted completely. The organic solvent was dried by rotation, and the crude product was dissolved in 20 mL of DCM.

[0292] T2401-1 (3.6 g, 3.9 mmol) was dissolved in 36 mL of DMF. DIPEA (2.7 mL, 15.6 mmol) was added at 0 °C, followed by HATU (1.67 g, 4.3 mmol). The reaction was carried out at 0 °C for 1 h, and LC / MS monitoring confirmed complete reaction of T2401-1. This solution was then added dropwise to the above reaction system, and the reaction was carried out at 0 °C for 4 h. 50 mL of water and 50 mL of EA were added to the reaction system, and the pH was adjusted to 6 using 0.1 M acetic acid solution. The mixture was extracted three times with 50 mL of EA, and the organic phases were combined. The organic phases were washed twice with 50 mL of 0.1 M acetic acid solution, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to obtain 2.45 g of white solid T2401-2, with a yield of 53%.

[0293] 3) Preparation of intermediate T2401-3

[0294] T2401-2 (2.4 g, 2.1 mmol) was dissolved in 240 mL of DCM. p-TsOH·H2O (2.38 g, 12.6 mmol) was added at 0 °C, and the mixture was stirred at room temperature for 24 h until the reaction was complete as controlled by LC / MS. Excess sodium bicarbonate aqueous solution was added dropwise at 0 °C to pH = 8. The DCM was then removed under reduced pressure. The remaining aqueous phase was purified by preparative chromatography to yield 1.88 g of white solid T2401-3, with a yield of 90%.

[0295] 4) Preparation of intermediate T2401-4

[0296] Intermediate NA (0.9 g, 1.3 mmol) was dissolved in 9 mL of DMF. DIPEA (0.68 mL, 3.9 mmol) was added at 0 °C, followed by HATU (0.543 g, 1.4 mmol). The reaction was carried out at 0 °C for 1 h, and LC / MS analysis confirmed complete reaction of intermediate NA. Then, T2401-3 (1.48 g, 1.5 mmol) was dissolved in 15 mL of DMF and added dropwise to the reaction system. The reaction was carried out at 0 °C for 12 h. After the reaction was complete, 20 mL of water, 20 mL of EA, and 0.1 M acetic acid solution were added to the reaction system to adjust the pH to 6. The mixture was extracted three times with 20 mL of EA. The organic phases were combined, washed twice with 0.1 M acetic acid solution, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 1.5 g of pale yellow solid T2401-4, with a yield of 69%.

[0297] 5) Preparation of intermediate T2401-5

[0298] T2401-4 (1.4 g, 0.8 mmol) was dissolved in 140 mL of DCM, and p-TsOH·H2O (0.798 g, 4 mmol) was added. The mixture was stirred at room temperature for 12 h until the reaction of T2401-4 was complete as indicated by LC / MS. The DCM was removed under reduced pressure, leaving a large amount of solid. Excess water was added in an ice bath, and the mixture was sonicated, resulting in the precipitation of a large amount of solid. The solid was filtered, dissolved in 20 mL of THF, and dried over anhydrous Na2SO4. The THF was then dried by rotation to obtain 2 g of crude, pale yellow solid T24011-5, which could be used directly in the next reaction without further purification. LC / MS: [M+H] + :1564.6.

[0299] 6) Preparation of intermediate T2401-6

[0300] The crude T2401-5 was dissolved in 250 mL of DCM. DIPEA (1.33 mL, 4.8 mmol) was added at 0 °C, followed by 60 mL of HATU (1.24 g, 3.2 mmol) in DCM. The mixture was stirred overnight at 0 °C. LC / MS analysis confirmed complete reaction of T2401-5. 30 mL of water was added to the system, followed by extraction once with 30 mL of DCM, and then twice with 20 mL of EA. The organic phases were combined, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 0.6 g of yellow solid T2401-5, with a yield of 46%.

[0301] 7) Preparation of intermediate T2401-7

[0302] T2401-6 (0.5 g, 0.3 mmol) was dissolved in 200 mL of DCM and 6.6 mL of glacial acetic acid. Janssen catalyst-1B (23.7 mg, 0.03 mmol) was added, and the mixture was purged with nitrogen four times. The reaction was carried out at 55 °C for 12 h. The reaction was monitored by LC / MS until T2401-6 was completely reacted. The pH was adjusted to 8 by adding saturated sodium bicarbonate solution. The aqueous phase was extracted three times with 30 mL of DCM. The organic phases were combined, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to give 0.33 g of pale red solid T2401-7, with a yield of 67%. LC / MS: [M+H] + :1518.1.

[0303] 8) Preparation of intermediate T2401-8

[0304] T2401-7 (100 mg, 0.065 mmol) was dissolved in 10 mL of THF, and 10 mg of 10% Pd / C was added. The mixture was purged with hydrogen four times, and the reaction was carried out at room temperature for 12 h. LC / MS analysis showed that T2401-7 was essentially complete. Pd / C was filtered off, and the mixture was washed three times with 5 mL of THF. The organic solvent was then dried by rotation to obtain T2401-8, which could be used directly for the next reaction without further purification. LC / MS: [M+H] + :1386.2.

[0305] 9) Synthesis of target molecule 2401

[0306] Intermediate NE6C (50 mg, 0.197 mmol) was dissolved in 5 mL of DMF. DIPEA (0.045 mL, 0.262 mmol) was added at 0 °C, followed by AOP (102 mg, 0.229 mmol). The reaction was carried out at room temperature for 1 h. After 1 h of reaction, 5 mL of the above T2401-8 DMF solution was added, and the reaction was carried out at room temperature for 2 h. LC / MS analysis showed that the reaction of T2401-8 was complete. The reaction solution was purified by preparative chromatography to obtain 23.1 mg of white solid T2401, with a yield of 22.8%.

[0307] 2. Synthesis of target molecule 2450

[0308] For the synthesis of 2450, please refer to the synthesis method of 2401, except that E2O is used instead of NE6C. The reaction solution was purified by preparative chromatography to obtain 40 mg of white solid T2450, with a yield of 38%.

[0309] 3. Synthesis of target molecule 2451

[0310] For the synthesis of 2451, please refer to the synthesis method of 2401, except that NE5C is used instead of NE6C. The reaction solution was purified by preparative chromatography to obtain 90.2 mg of white solid 2451, with a yield of 74%.

[0311] 4. Synthesis of target molecule 2455

[0312] [Correction based on Rule 91, 04.03.2025] For the synthesis of 2455, please refer to the synthesis method of 2401, where NE2C is used instead of NE6C. The reaction solution was purified by preparative chromatography to obtain 86.7 mg of white solid 2455, with a yield of 54%.

[0313] 5. Synthesis of target molecule 2457

[0314] For the synthesis of 2457, please refer to the synthesis method of 2401, except that NE4C is used instead of NE6C. The reaction solution was purified by preparative chromatography to obtain 127.5 mg of white solid 2457, with a yield of 77%.

[0315] 6. Synthesis of target molecule 2460

[0316] For the synthesis of 2460, please refer to the synthesis method of 2401, except that NE7C is used instead of NE6C. The reaction solution was purified by preparative chromatography to obtain 76.6 mg of white solid 2460, with a yield of 68%.

[0317] 7. Synthesis of target molecule 2461

[0318] The synthesis of 2461 is similar to that of 2401, except that NE15C is used instead of NE6C. The reaction solution was purified to yield 72.4 mg of a white solid, 2461, in 50% yield.

[0319] 8. Synthesis of target molecule 2462

[0320] T2451 (110 mg, 0.072 mmol) was dissolved in 3 mL of TFA, and SCF2 reagent (99 mg, 0.360 mmol) was added at 0 °C. The reaction was then carried out at 30 °C for 16 h. LC / MS analysis showed that the reaction of T2451 was complete. The reaction solution was purified by preparative chromatography to give 47.2 mg of white solid T2462, with a yield of 40%.

[0321] 9. Synthesis of target molecule 2463

[0322] 2451 (100 mg, 0.065 mmol) was dissolved in 3 mL of TFA, and SCF3 reagent (96.2 mg, 0.325 mmol) was added at 0 °C. The reaction was then carried out at 30 °C for 16 h. LC / MS analysis showed that the reaction was complete. The reaction solution was purified by preparative chromatography to give 72.7 mg of white solid 2463, with a yield of 68%.

[0323] 10. Synthesis of target molecules 2402A and 2402B

[0324] 1) Preparation of intermediate T2402-1

[0325] The synthesis of T2402-1 is described by referring to the synthesis method of T2401-1, except that BD02 is used instead of BD01. The crude product was purified by preparative chromatography to obtain 16.5 g of T2402-1, with a yield of 84%.

[0326] 2) Preparation of intermediate T2402-2

[0327] The synthesis of T2402-2 is described by referring to the synthesis method of T2401-2, except that T2402-1 is used instead of T2401-1. The crude product was purified by preparative chromatography to obtain 11.16 g of T2402-2, with a yield of 67%.

[0328] 3) Preparation of intermediate T2402-3

[0329] The synthesis of T2402-3 is described by referring to the synthesis method of T2401-3, except that T2402-2 is used instead of T2401-2. The crude product was purified by preparative chromatography to obtain T2402-3, totaling 8.6 g, with a yield of 90%.

[0330] 4) Preparation of intermediate T2402-4

[0331] The synthesis of T2402-4 is described by referring to the synthesis method of T2401-4, except that T2402-3 is used instead of T2401-3. The crude product was purified by silica gel column chromatography to obtain 10.31 g of T2402-4, with a yield of 78%.

[0332] 5) Preparation of intermediate T2402-5

[0333] For the synthesis of T2402-5, please refer to the synthesis method of T2401-5, except that T2402-4 is used instead of T2401-4. LC / MS: [M+H] + :1577.7.

[0334] 6) Preparation of intermediate T2402-6

[0335] The synthesis of T2402-6 is described by referring to the synthesis method of T2401-6, except that T2402-5 is used instead of T2401-5. The crude product was purified by preparative chromatographic analysis to obtain two isomers: 1.6 g of T2402-6A (18% yield) and 1.3 g of T2402-6B (14% yield). LC / MS: [M+H] + :1559.7.

[0336] 7) Preparation of intermediate T2402-7

[0337] T2402-7A: The synthesis of T2402-7A is similar to that of T2401-7, except that T2402-6A is used instead of T2401-6. The crude product was purified by preparative chromatography to obtain 1.07 g of a white solid, T2402-7A, in 70% yield. LC / MS: [M+H] + :1531.7.

[0338] T2402-7B: The synthesis of T2402-7B is similar to that of T2401-7, except that T2402-6B is used instead of T2401-6. The crude product was purified by preparative chromatographic analysis to obtain 760 mg of white solid T2402-7B, with a yield of 60%. LC / MS: [M+H] + :1531.7.

[0339] 8) Preparation of intermediate T2402-8

[0340] T2402-8A: For the synthesis of T2402-8A, please refer to the synthesis method of T2401-8, where T2402-7A is used instead of T2401-7. LC / MS: [M+H] + :1399.7.

[0341] T2402-8B: For the synthesis of T2402-8B, please refer to the synthesis method of T2401-8, where T2402-7B is used instead of T2401-7. LC / MS: [M+H] + :1399.7.

[0342] 9) Synthesis of target molecule 2402A

[0343] For the synthesis of 2402A, please refer to the synthesis method of 2401. The reaction solution was purified by preparative chromatography to obtain 32 mg of white solid 2402A, with a yield of 31%.

[0344] 10) Synthesis of target molecule 2402B

[0345] For the synthesis of 2402B, please refer to the synthesis method of 2401. The reaction solution was purified by preparative chromatography to obtain 28 mg of white solid 2402B, with a yield of 27%.

[0346] 11. Synthesis of the target molecule 204B

[0347] For the synthesis of 204B, please refer to the synthesis method of 2401, where NE4C is used instead of NE6C. The reaction solution was purified to obtain 22 mg of white solid 204B, with a yield of 22%.

[0348] 12. Synthesis of target molecule 205B

[0349] For the synthesis of 205B, please refer to the synthesis method of 2401, where NE5C is used instead of NE6C. The reaction solution was purified by preparative chromatography to obtain 83 mg of white solid 205B, with a yield of 81%.

[0350] 13. Synthesis of target molecule 205A

[0351] For the synthesis of 205A, please refer to the synthesis method of 2401, where NE5C is used instead of NE6C. The reaction solution was purified by preparative chromatography to obtain 82 mg of white solid 205A, with a yield of 80%.

[0352] 14. Synthesis of the target molecule 211B

[0353] For the synthesis of 211B, please refer to the synthesis method of 2401, where E1O is used instead of NE6C. The reaction solution was sent for preparative purification to obtain 50 mg of white solid 211B, with a yield of 48%.

[0354] 15. Synthesis of the target molecule 212B

[0355] For the synthesis of 212B, please refer to the synthesis method of 2401, where E2O is used instead of NE6C. The reaction solution was sent for preparative purification to obtain 53 mg of colored solid 212B, with a yield of 50%.

[0356] 16. Synthesis of target molecule 212A

[0357] For the synthesis of 212A, please refer to the synthesis method of 2401, where E2O is used instead of NE6C. The reaction solution was purified to obtain 16 mg of white solid 212A, with a yield of 15%.

[0358] 17. Synthesis of the target molecule 213B

[0359] For the synthesis of 213B, please refer to the synthesis method of 2401, where E3O is used instead of NE6C. The reaction solution was purified to obtain 78 mg of white solid 213B, with a yield of 72%.

[0360] 18. Synthesis of target molecule 2403

[0361] 1) Preparation of intermediate T2403-1

[0362] The synthesis of T2403-1 is described by referring to the synthesis method of T2401-1, except that BD03 is used instead of BD01. The crude product was prepared and purified by chromatographic chromatography to obtain 11.32 g of white solid T2403-1, with a yield of 85%.

[0363] 2) Preparation of intermediate T2403-2

[0364] The synthesis of T2403-2 is described by referring to the synthesis method of T2401-2, except that T2403-1 is used instead of T2401-1. The crude product was prepared and purified by chromatographic chromatography to obtain 4.42 g of white solid T2403-2, with a yield of 52%.

[0365] 3) Preparation of intermediate T2403-3

[0366] The synthesis of T2403-3 is described by referring to the synthesis method of T2401-3, except that T2403-2 is used instead of T2401-2. After preparative chromatographic purification, 2.56 g of white solid T2403-3 was obtained, with a yield of 70%.

[0367] 4) Preparation of intermediate T2403-4

[0368] The synthesis of T2403-4 is described by referring to the synthesis method of T2401-4, except that T2403-2 is used instead of T2401-2. The crude product was purified by silica gel column chromatography to obtain 2.48 g of white solid T2403-4, with a yield of 92%.

[0369] 5) Preparation of intermediate T2403-5

[0370] For the synthesis of T2403-5, please refer to the synthesis method of T2401-5, except that T2403-4 is used instead of T2401-4. LC / MS: [M+H] + :1591.7.

[0371] 6) Preparation of intermediate T2403-6

[0372] The synthesis of T2403-6 is described by referring to the synthesis method of T2401-6, except that T2403-5 is used instead of T2401-5. The crude product was purified by preparative chromatography to obtain 1.32 g of white solid T2403-6, with a yield of approximately 60%.

[0373] 7) Preparation of intermediate T2403-7

[0374] The synthesis of T2403-7 is described by referring to the synthesis method of T2401-7, except that T2403-6 is used instead of T2401-6. The crude product was purified by preparative chromatography to obtain 0.61 g of a pale yellow solid, T2403-7, in 50% yield. LC / MS: [M+H] + :1545.5.

[0375] 8) Preparation of intermediate T2403-8

[0376] For the synthesis of T2403-8, please refer to the synthesis method of T2401-8, except that T2403-7 is used instead of T2401-7. LC / MS: [M+H] + :1413.6.

[0377] 9) Synthesis of target molecule 2403

[0378] For the synthesis of 2403, please refer to the synthesis method of 2401. For the synthesis of T2403-6, please refer to the synthesis method of T2401-6, except that T2403-8 is used instead of T2401-8. The reaction solution was purified by preparative chromatography to obtain 208 mg of white solid 2403, with a yield of 72%.

[0379] 19. Synthesis of target molecule 2430

[0380] The prepared T2403-8 was dissolved in DMF (2.5 mL), and Na2CO3 (21 mg, 0.18 mmol) was added at 0 °C, followed by CH3I (0.03 mL, 0.52 mmol). The reaction was carried out at 40 °C for 2 h. The reaction of T2403-8 was monitored by LC / MS until complete. The solid sodium carbonate was removed by filtration, and the solid was washed twice with 5 mL of acetonitrile. The organic phases were combined, and the organic solvent was dried by rotary evaporation. The crude product was purified by preparative chromatography to give 68 mg of white solid 2430, with a yield of 87%.

[0381] 20. Synthesis of target molecule 2404

[0382] 1) Preparation of intermediate T2404-1

[0383] The synthesis of T2404-1 is described by referring to the synthesis method of T2401-1, except that BD04 is used instead of BD01. The crude product was purified to obtain 12.9 g of white solid T2404-1, with a yield of 95%.

[0384] 2) Preparation of intermediate T2404-2

[0385] [Correction based on Rule 91, 04.03.2025] For the synthesis of T2404-2, please refer to the synthesis method of T2401-2, where T2404-1 is used instead of T2401-1. The crude product was purified by silica gel column chromatography to obtain 14.1 g of T2404-2, with a yield of 89%.

[0386] 3) Preparation of intermediate T2404-3

[0387] The synthesis of T2404-3 is described by referring to the synthesis method of T2401-3, except that T2404-2 is used instead of T2401-2. The crude product was purified by preparative chromatographic analysis to obtain 8.67 g of white solid T2404-3, with a yield of 72%.

[0388] 4) Preparation of intermediate T2404-4

[0389] The synthesis of T2404-4 is described by referring to the synthesis method of T2401-4, except that T2404-3 is used instead of T2401-3. The crude product was purified by silica gel column chromatography to obtain 7.57 g of white solid T2404-4, with a yield of 77%.

[0390] 5) Preparation of intermediate T2404-5

[0391] For the synthesis of T2404-5, please refer to the synthesis method of T2401-5, except that T2404-4 is used instead of T2401-4. LC / MS: [M+H] + :1589.8.

[0392] 6) Preparation of intermediate T2404-6

[0393] The synthesis of T2404-6 is based on the same method as T2401-6, except that T2404-5 is used instead of T2401-5. The crude product was purified by silica gel column chromatography to obtain 3.59 g of white solid T2404-6, with a yield of 52%.

[0394] 7) Preparation of intermediate T2404-7

[0395] The synthesis of T2404-7 is based on the same method as T2401-7, except that T2404-6 is used instead of T2401-6. The crude product was purified by preparative chromatography to obtain 1.7 g of white solid T2404-7, with a yield of 55%. LC / MS: [M+H] + : 1543.9.

[0396] 8) Preparation of intermediate T2404-8

[0397] For the synthesis of T2404-8, please refer to the synthesis method of T2401-8, except that T2404-7 is used instead of T2401-7. LC / MS: [M+H] + :1411.9.

[0398] 9) Synthesis of target molecule 2404

[0399] For the synthesis of 2404, please refer to the synthesis method of 2401, except that NE5C is used instead of NE6C. The reaction solution was purified by preparative chromatography to obtain 410 mg of white solid 2404, with a yield of 75%.

[0400] 21. Synthesis of target molecule 2416

[0401] 1) Preparation of intermediate T2416-1

[0402] For the synthesis of T2416-1, please refer to the synthesis method of T2401-1, except that BD16 is used instead of BD01. The crude product was purified by silica gel column chromatography to obtain 8.3 g of white solid T2416-1, with a yield of 60%.

[0403] 2) Preparation of intermediate T2416-2

[0404] Preparation of intermediate NB-Me

[0405] S1 (20 g, 59.2 mmol) was dissolved in 200 mL of DCM. DIPEA (30.86 mL, 177.5 mmol) was added at 0 °C, followed by CH3I (7.37 mL, 118.3 mmol). The mixture was gradually brought back to room temperature and reacted for 12 h. After 12 h, the reaction was complete as controlled by LC / MS. 100 mL of water was added to the reaction system, and the mixture was extracted three times with 100 mL of DCM. The organic phases were combined, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 10.35 g of NB-Me, with a yield of 50%. LC / MS: [M+H] + :353.2.

[0406] NB-Me (10.30 g, 29.3 mmol) was dissolved in 180 mL of DCM. 36 mL of TFA was added at 0 °C, and the mixture was gradually brought to room temperature before reacting for another 3 h. LC / MS analysis showed that NB-Me reacted completely. The DCM and TFA were then directly removed by vortexing to obtain a yellow oil. This oil was used directly in the next reaction without purification. The yellow oil was dissolved in 100 mL of DMF, and 45 mL of DMF solution containing DIPEA (6.10 mL, 35.1 mmol), T2416-1 (11.25 g, 11.7 mmol), and finally HATU (8.89 g, 23.4 mmol) was added. The mixture was then reacted at room temperature for 48 h. The reaction of T2416-1 was controlled to be complete by LC / MS. 50 mL of water and 50 mL of EA were added to the reaction system, and the mixture was extracted three times with 50 mL of EA. The organic phases were combined and washed twice with saturated saline solution. The organic phases were then dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 11.49 g of T2416-2, with a yield of 82%.

[0407] 3) Preparation of intermediate T2416-3

[0408] T2416-2 (11.12 g, 9.3 mmol) was dissolved in 350 mL of DCM. TsOH·H2O (10.61 g, 55.8 mmol) was added at 0 °C, and the mixture was stirred at room temperature for 24 h until the reaction was complete as indicated by LC / MS. The mixture was then dried under vacuum using a rotary dryer, removing the DCM. A large amount of solid remained. Excess water was added in an ice bath, and the mixture was sonicated, resulting in the precipitation of a large amount of solid. The solid was filtered, the liquid phase was discarded, and the solid was dissolved in THF and dried over anhydrous Na2SO4. The mixture was then dried under vacuum using a rotary dryer, removing the THF, to obtain crude T2416-2-Me, which could be used directly in the next reaction without further purification. The crude product was dissolved in 150 mL of THF, and LiOH·H2O (3.91 g, 93 mmol) was dissolved in 150 mL of water. This solution was added dropwise to the reaction system at 0 °C, and the mixture was stirred at room temperature for 5 h until the reaction was complete as indicated by LC / MS. The reaction was stopped, the pH of the solution was adjusted to 6 with 0.1M phosphoric acid solution, THF was removed under vacuum, and the remaining aqueous phase was purified by preparative chromatography to obtain 6.47 g of white solid T2416-3, with a yield of 68%.

[0409] 4) Preparation of intermediate T2416-4

[0410] The synthesis of T2416-4 is described by referring to the synthesis method of T2401-4, except that T2416-3 is used instead of T2401-3. The crude product was purified by silica gel column chromatography to obtain 5.58 g of white solid T2416-4, with a yield of 76%.

[0411] 5) Preparation of intermediate T2416-5

[0412] For the synthesis of T2416-5, please refer to the synthesis method of T2401-5, except that T2416-4 is used instead of T2401-4. LC / MS: [M+H] + :1599.7.

[0413] 6) Preparation of intermediate T2416-6

[0414] The synthesis of T2416-6 is described by referring to the synthesis method of T2401-6, except that T2416-5 is used instead of T2401-5. The crude product was purified by preparative chromatography to obtain 2.62 g of white solid T2416-6, with a yield of 53%.

[0415] 7) Preparation of intermediate T2416-7

[0416] The synthesis of T2416-7 is based on the same method as T2401-7, except that T2416-6 is used instead of T2401-6. The crude product was purified by preparative chromatography to obtain 1.5 g of a pale yellow solid, T2416-7, in 60% yield. LC / MS: [M+H] +:1553.6.

[0417] 8) Preparation of intermediate T2416-8

[0418] For the synthesis of T2416-8, please refer to the synthesis method of T2401-8, except that T2416-7 is used instead of T2401-7. LC / MS: [M+H] + :1421.6.

[0419] 9) Synthesis of target molecule 2416

[0420] For the synthesis of 2416, please refer to the synthesis method of 2401, except that NE5C is used instead of NE6C. The reaction solution was sent for preparative purification to obtain 180 mg of white solid 2416, with a yield of 80%.

[0421] 22. Synthesis of target molecule 2410

[0422] Preparation of intermediate T2410-1

[0423] The synthesis of T2410-1 is described by referring to the synthesis method of T2401-1, except that BD10 is used instead of BD01. The crude product was prepared and purified by chromatographic chromatography to obtain 13.3 g of white solid T2410-1, with a yield of 99%.

[0424] 1) Preparation of intermediate T2410-2

[0425] The synthesis of T2410-2 is described by referring to the synthesis method of T2401-2, except that T2410-1 is used instead of T2401-1. The crude product was purified by preparative chromatography to obtain 2.8 g of white solid T2410-2, with a yield of 39%.

[0426] 2) Preparation of intermediate T2410-3

[0427] The synthesis of T2410-3 is described by referring to the synthesis method of T2401-3, except that T2410-2 is used instead of T2401-2. The crude product was purified by preparative chromatography to obtain 1.8 g of white solid T2410-3, with a yield of 74%.

[0428] 3) Preparation of intermediate T2410-4

[0429] The synthesis of T2410-4 is described by referring to the synthesis method of T2401-4, except that T2410-3 is used instead of T2401-3. The crude product was purified by silica gel column chromatography to obtain 1.78 g of pale yellow solid T2410-4, with a yield of 73%.

[0430] 4) Preparation of intermediate T2410-5

[0431] For the synthesis of T2410-5, please refer to the synthesis method of T2401-5, except that T2410-4 is used instead of T2401-4. LC / MS: [M+H] + :1578.7.

[0432] 5) Preparation of intermediate T2410-6

[0433] [Correction based on Rule 91, 04.03.2025] For the synthesis of T2410-6, please refer to the synthesis method of T2401-6, where T2410-5 is used instead of T2401-5. The crude product was purified by silica gel column chromatography to obtain 0.72 g of yellow solid T2410-6, with a yield of 46%.

[0434] 6) Preparation of intermediate T2410-7

[0435] The synthesis of T2410-7 is based on the same method as T2401-7, except that T2410-6 is used instead of T2401-6. The crude product was purified by silica gel column chromatography to obtain 0.2 g of a pale red solid, which was then added to T2401-7 in 31% yield. LC / MS: [M+H] + :1531.6.

[0436] 7) Preparation of intermediate T2410-8

[0437] For the synthesis of T2410-8, please refer to the synthesis method of T2401-8, except that T2410-7 is used instead of T2401-7. LC / MS: [M+H] + : 1399.7.

[0438] 8) Synthesis of target molecule 2410

[0439] [Correction based on Rule 91, 04.03.2025] For the synthesis of 2410, please refer to the synthesis method of 2401. The reaction solution was purified by preparative chromatography to obtain 68.8 mg of white solid 2410, with a yield of 68.8%.

[0440] 23. Synthesis of target molecule 2452

[0441] The synthesis of 2452 is described by referring to the synthesis method of 2401, except that NE5C is used instead of NE6C. The reaction solution was purified by preparative chromatography to obtain 39.6 mg of a white solid, 2452, in 49% yield.

[0442] 24. Synthesis of target molecule 2412

[0443] 1) Preparation of intermediate T2412-1

[0444] For the synthesis of T2412-1, please refer to the synthesis method of T2401-1, except that BD12 is used instead of BD01. The crude product was purified by silica gel column chromatography to obtain 18.5 g of yellow solid T2412-1, with a yield of 91%.

[0445] 2) Preparation of intermediate T2412-2

[0446] The synthesis of T2412-2 is described by referring to the synthesis method of T2401-2, except that T2412-1 is used instead of T2401-1. The crude product was purified by preparative chromatography to obtain 2.9 g of white solid T2412-2, with a yield of 33%.

[0447] 3) Preparation of intermediate T2412-3

[0448] The synthesis of T2412-3 is described by referring to the synthesis method of T2401-3, except that T2412-2 is used instead of T2401-2. The crude product was purified by preparative chromatography to obtain 1.7 g of white solid T2412-3, with a yield of 68%.

[0449] 4) Preparation of intermediate T2412-4

[0450] The synthesis of T2412-4 is described by referring to the synthesis method of T2401-4, except that T2412-3 is used instead of T2401-3. The crude product was purified by silica gel column chromatography to obtain 1.62 g of pale yellow solid T2412-4, with a yield of 74%.

[0451] 5) Preparation of intermediate T2412-5

[0452] For the synthesis of T2412-5, please refer to the synthesis method of T2401-5, except that T2412-4 is used instead of T2401-4. LC / MS: [M+H] + :1577.8.

[0453] 6) Preparation of intermediate T2412-6

[0454] [Correction based on Rule 91, 04.03.2025] For the synthesis of T2412-6, please refer to the synthesis method of T2401-6, where T2412-5 is used instead of T2401-5. The crude product was purified by silica gel column chromatography to obtain 0.67 g of pale yellow solid T2412-6, with a yield of 47%.

[0455] 7) Preparation of intermediate T2412-7

[0456] The synthesis of T2412-7 is described by referring to the synthesis method of T2401-7, except that T2412-6 is used instead of T2401-6. The crude product was purified by silica gel column chromatography to obtain 0.26 g of pale red solid T2412-7, with a yield of 44%. LC / MS: [M+H] +:1532.7.

[0457] 8) Preparation of intermediate T2412-8

[0458] For the synthesis of T2412-8, please refer to the synthesis method of T2401-8, except that T2412-7 is used instead of T2401-7. LC / MS: [M+H] + :1400.7.

[0459] 9) Synthesis of target molecule 2412

[0460] The synthesis of 2412 is based on the same method as 2401, except that T2412-8 is used instead of T2401-8. The reaction mixture was purified by preparative chromatography to yield 85.3 mg of a white solid, 2412, in 84% yield.

[0461] 25. Synthesis of target molecule 2453

[0462] For the synthesis of 2453, please refer to the synthesis method of 2401. The reaction solution was purified by preparative chromatography to obtain 35.1 mg of white solid 2453, with a yield of 35%.

[0463] 26. Synthesis of target molecule 2454

[0464] For the synthesis of 2454, please refer to the synthesis method of 2401. After preparative chromatographic purification, 12.7 mg of white solid 2454 was obtained, with a yield of 24%.

[0465] 27. Synthesis of target molecule 2421

[0466] 1) Preparation of intermediate T2421-1

[0467] The synthesis of T2421-1 is described by referring to the synthesis method of T2401-1, except that BD21 is used instead of BD01. The crude product was purified to obtain 4.66 g of white solid T2421-1, with a yield of 60%.

[0468] 2) Preparation of intermediate T2421-2

[0469] The synthesis of T2421-2 is described by referring to the synthesis method of T2401-2, except that T2421-1 is used instead of T2401-1. The crude product was purified to obtain 5.5 g of white solid T2421-2, with a yield of 95%.

[0470] 3) Preparation of intermediate T2421-3

[0471] The synthesis of T2421-3 is described by referring to the synthesis method of T2401-3, except that T2421-2 is used instead of T2401-2. The remaining aqueous phase was purified by preparative chromatography to obtain 2.97 g of white solid T2421-3, with a yield of 61%.

[0472] 4) Preparation of intermediate T2421-4

[0473] The synthesis of T2421-4 is described by referring to the synthesis method of T2401-4, except that T2421-3 is used instead of T2401-3. The crude product was purified by silica gel column chromatography to obtain 2.99 g of white solid T2421-4, with a yield of 80%.

[0474] 5) Preparation of intermediate T2421-5

[0475] For the synthesis of T2421-5, please refer to the synthesis method of T2401-5, except that T2421-4 is used instead of T2401-4. LC / MS: [M+H] + :1599.7.

[0476] 6) Preparation of intermediate T2421-6

[0477] The synthesis of T2421-6 is described by referring to the synthesis method of T2401-6, except that T2421-5 is used instead of T2401-5. The crude product was purified by silica gel column chromatography to obtain 1.67 g of pale yellow solid T2421-6, with a yield of 64%.

[0478] 7) Preparation of intermediate T2421-7

[0479] The synthesis of T2421-7 is similar to that of T2401-7, except that T2421-6 is used instead of T2401-6. The crude product was purified by preparative chromatography to give 1.05 g of white solid T2421-7, with a yield of 73%. LC / MS: [M+H] + :1553.7.

[0480] 8) Preparation of intermediate T2421-8

[0481] For the synthesis of T2421-8, please refer to the synthesis method of T2401-8, except that T2421-7 is used instead of T2401-7. LC / MS: [M+H] + :1421.7.

[0482] 9) Synthesis of target molecule 2421

[0483] For the synthesis of 2421, please refer to the synthesis method of 2401, except that NE5C1 is used instead of NE6C. The reaction solution was purified by preparative chromatography to obtain 65 mg of white solid 2421, with a yield of 65%.

[0484] 28. Synthesis of target molecule 2427

[0485] T2421-8 (100 mg, 0.071 mmol) was dissolved in 10 mL of MeOH, and selectfluor fluorinating reagent (55 mg, 0.155 mmol) was added. The mixture was purged with nitrogen three times and reacted at 65 °C for 10 h. The reaction was monitored by LC / MS until T2421-8 was completely reacted. The reaction was then stopped, and the organic solvent was dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to give 35 mg of white solid T2427-8, with a yield of 34%. LC / MS: [M+H] + :1457.7.

[0486] [Corrected according to Rule 91, 04.03.2025] NE5C (19 mg, 0.079 mmol) was dissolved in 1 mL of DMF. DIPEA (0.018 mL, 0.102 mmol) was added at 0 °C, followed by AOP (40 mg, 0.091 mmol). The reaction was carried out at room temperature for 1 h. Then, 1 mL of the above-mentioned DMF solution of T2427-8 (33 mg, 0.023 mmol) was added, and the reaction was carried out at room temperature for 2 h. LC / MS analysis showed that the reaction of T2427-8 was complete. The reaction solution was purified by preparative chromatography to obtain 22 mg of white solid 2427, with a yield of 60%.

[0487] 29. Synthesis of target molecule 2428

[0488] T2421-8 (100 mg, 0.071 mmol) was dissolved in 8 mL of TFA, and S1 (98 mg, 0.355 mmol) was added. The mixture was purged with nitrogen three times and reacted at 40 °C for 16 h. The reaction was monitored by LC / MS until T2421-8 was completely reacted, and then the TFA was removed under reduced pressure. Sodium bicarbonate aqueous solution was slowly added dropwise at 0 °C to adjust the pH to 8-9. Methanol was then added, at which point a solid precipitated. The solid was filtered, and most of the methanol was removed under reduced pressure. The crude product was purified by preparative chromatography to obtain 20 mg of white solid T2428-8, with a yield of 19%. LC / MS: [M+H] + :1503.5.

[0489] [Corrected according to Rule 91, 04.03.2025] NE5C (11 mg, 0.047 mmol) was dissolved in 0.5 mL of DMF. DIPEA (0.011 mL, 0.060 mmol) was added at 0 °C, followed by AOP (24 mg, 0.053 mmol). The reaction was carried out at room temperature for 1 h. After 1 h, 0.5 mL of the above-mentioned DMF solution of T2428-8 (20 mg, 0.014 mmol) was added, and the reaction was carried out at room temperature for 2 h. LC / MS showed that the reaction of T2428-8 was complete. The reaction solution was purified by preparative chromatography to obtain 10 mg of white solid 2428, with a yield of 46%.

[0490] 30. Synthesis of target molecule 2407

[0491] 1) Preparation of intermediate T2407-1

[0492] The synthesis of T2407-1 is described by referring to the synthesis method of T2401-1, except that BD07 is used instead of BD01. The crude product was prepared and purified by chromatographic chromatography to obtain 4.54 g of white solid T2407-1, with a yield of 72%.

[0493] 2) Preparation of intermediate T2407-2

[0494] The synthesis of T2407-2 is based on the same method as T2401-2, except that T2407-1 is used instead of T2401-1. The crude product was prepared and purified by chromatographic chromatography to obtain 3.5 g of white solid T2407-2, with a yield of 68%.

[0495] 3) Preparation of intermediate T2407-3

[0496] [Correction based on Rule 91, 04.03.2025] For the synthesis of T2407-3, please refer to the synthesis method of T2401-3, where T2407-2 is used instead of T2401-1. After preparative chromatographic purification, 2.2 g of white solid T2407-3 was obtained, with a yield of 74%.

[0497] 4) Preparation of intermediate T2407-4

[0498] Intermediate NA (340 mg, 0.49 mmol) was dissolved in 5 mL of DMF. DIPEA (0.26 mL, 1.47 mmol) was added at 0 °C, followed by HATU (210 mg, 0.54 mmol). The reaction was carried out at 0 °C for 1 h. Then, 5 mL of DM solution containing T2407-3 (600 mg, 0.56 mmol) was added dropwise to the reaction mixture, and the reaction was carried out at 0 °C for 4 h. Then, 10 mL of water, 10 mL of EA, and 0.1 M acetic acid solution were added to the reaction mixture to adjust the pH to 6. The mixture was extracted three times with 10 mL of EA. The combined organic phases were washed twice with 10 mL of 0.1 M acetic acid solution. The organic phase was then dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 550 mg of pale yellow solid T2407-4, with a yield of 64%. LC / MS: [M+H] + :1726.7.

[0499] 5) Preparation of intermediate T2407-5

[0500] T2407-4 (550 mg, 0.31 mmol) was dissolved in 30 mL of DCM, and p-TsOH·H2O (302 mg, 1.55 mmol) was added. The mixture was then stirred overnight at room temperature until the reaction of T2407-4 was complete as indicated by LC / MS. The DCM was removed by vacuum rotary drying, leaving a large amount of solid. Excess water was added in an ice bath, and the mixture was sonicated, resulting in the precipitation of a large amount of solid. The solid was filtered, the liquid phase was discarded, and the solid was dissolved in THF. After drying with anhydrous Na2SO4, the solid was rotary dried under reduced pressure to obtain 830 mg of T2407-5, a pale yellow crude product, which could be used directly in the next reaction without further purification. LC / MS: [M+H] + : 1626.6.

[0501] 6) Preparation of intermediate T2407-6

[0502] [Corrected according to Rule 91, 04.03.2025] The above-mentioned crude T2407-5 (830 mg, 0.51 mmol) was dissolved in 70 mL of DCM. DIPEA (0.7 mL, 4.08 mmol) was added at 0 °C, followed by HATU (792 mg, 2.04 mmol). The mixture was stirred overnight at 0 °C. LC / MS monitoring showed that T2407-5 had completely reacted. 10 mL of water was added to the system, and the mixture was extracted once with 10 mL of DCM, followed by two extractions with 10 mL of EA. The organic phases were combined, dried over anhydrous Na2SO4, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography to obtain 430 mg of yellow solid T2407-6, with a yield of 84%. LC / MS: [M+H] + :1608.6.

[0503] 7) Preparation of intermediate T2407-7

[0504] T2407-6 (430 mg, 0.26 mmol) was dissolved in 150 mL of DCM and 5 mL of acetic acid. Janssen catalyst-1B (19.6 mg, 0.026 mmol) was added, and the mixture was purged with nitrogen four times. The reaction was carried out at 55 °C for 12 h. The reaction was monitored by LC / MS until complete. Saturated sodium bicarbonate solution was added to adjust the pH to 8. The aqueous phase was extracted three times with 10 mL of DCM. The organic phases were combined, dried over anhydrous Na₂SO₄, filtered, and the filtrate was dried under reduced pressure to obtain the crude product. The crude product was purified by preparative chromatography to obtain 100 mg of pale red solid T2407-7, with a yield of 23%. LC / MS: [M+H] + :1580.4.

[0505] 8) Preparation of intermediate T2407-8

[0506] T2401-7 (100 mg, 0.063 mmol) was dissolved in 10 mL of THF, and 10 mg of 10% Pd / C was added. The mixture was purged with hydrogen four times, and the reaction was carried out overnight at room temperature. LC / MS showed that T2401-7 was essentially complete. Pd / C was filtered off, and the mixture was washed three times with THF. The organic phase was then dried over anhydrous Na2SO4, filtered, and the filtrate was rotary dried under reduced pressure to obtain crude T2407-8, which could be used directly for the next reaction without further purification. LC / MS: [M+H] + : 1446.5.

[0507] 9) Synthesis of target molecule 2407

[0508] Intermediate NE6C (48 mg, 0.189 mmol) and the crude product T2407-8 were dissolved in 5 mL of DMF. DIPEA (0.088 mL, 0.504 mmol) was added at 0 °C, followed by AOP (923 mg, 2.04 mmol). The reaction was carried out at room temperature for 2 h. LC / MS analysis showed that the reaction of T2407-8 was complete. The reaction solution was purified by preparative chromatography to obtain 25.1 mg of a white solid, 2407, with a yield of 22.8%. LC / MS: [M] + :1603.8.

[0509] Table 3. Structural identification data of some compounds in the target molecule synthesis pathway.

[0510] Experimental Example 1 Activity Test

[0511] 1. In vitro enzyme activity test

[0512] This experiment used the PCSK9-LDLR HTRF Assay to test the enzyme activity of the synthesized new compounds.

[0513] 1) The experimental steps are as follows:

[0514] i) Dilute the compound to be tested to the reference concentration, and use the diluted solution as the first point to dilute it by a factor of 3, for a total of 10 points.

[0515] ii) Transfer the diluted compound to a 384 analytical plate, with two replicates for each concentration. Centrifuge the 384 analytical plate at 1000 rpm.

[0516] iii) Add 2.5 μL of PCSK9 solution to each well and then centrifuge at 1000 r / min for 1 min.

[0517] iv) Incubate at 25°C for 10 min.

[0518] v) Add 2.5 μL of LDLR solution to each well, and then centrifuge at 1000 r / min for 1 min.

[0519] vi) Add 5 μL of streptavidin-TB&PAB ANTI human IGG-XL665 working solution to each well, and then centrifuge at 1000 r / min for 1 min.

[0520] vii) Incubate at 25°C for 60 min.

[0521] viii) The HTRF ratio (665 / 620) was detected using a BMG microplate reader for data analysis.

[0522] 2) The enzyme activity results of the new compounds are shown in Table 4:

[0523] Table 4. Enzyme activity test results of the new compounds

[0524] 2. Efficacy test of intraperitoneal injection in mice

[0525] Eight-week-old male C57 mice were divided into seven groups of five each. The control group received daily injections of Control (2.5% DMSO in PBS), while the experimental groups received daily intraperitoneal injections of the test compound (1 mg / kg / day). Blood samples were collected from the orbital sinus before administration and on day 7 after administration to measure serum LDL-C concentrations. The results are shown in Tables 5 and 6 below.

[0526] Table 5. Summary of blood lipid levels before and after drug administration

[0527] As shown in Table 5 and Figure 1, under the experimental conditions, after 7 days of intraperitoneal injection, the LDL-C values ​​of groups 2451, 2453, 205A and 2421 decreased by 32.63%, 35.56%, 32.63% and 35.03% respectively compared with the control group, and there was a significant difference compared with the MK-0616 group.

[0528] Table 6. Summary of blood lipid levels before and after drug administration

[0529] As shown in Table 6 and Figure 2, under the experimental conditions, after 7 days of intraperitoneal injection, the LDL-C values ​​of groups 2401, 2455, 2457, 2460, 2461, and 2462 decreased by 28.44%, 31.09%, 31.39%, 34.94%, 33.76%, and 31.56% respectively compared with the control group, and there was a significant difference compared with the MK-0616 group.

[0530] Experiment Example 2: Oral Dosing Experiment

[0531] Objective: To induce a hyperlipidemia model in golden hamsters by feeding them a high-fat diet, and to evaluate the therapeutic effect of the test substance on the hyperlipidemia model by detecting and monitoring the blood lipids LDL-C and TG in the animals.

[0532] Methods: Golden hamsters were fed a high-fat diet for 6 weeks to establish a hyperlipidemia model. At the end of the 6th week, blood was collected from the orbital sinus of all golden hamsters to monitor serum LDL-C and TG levels. Then, based on body weight and the pre-group monitoring of the four lipid parameters, the golden hamsters fed the high-fat diet were evenly divided into 5 groups of 8 hamsters each. Group 1 was the solvent control group (360 mg / kg sodium caprylate solution dissolved in 2% DMSO + 98% physiological saline). Groups 2-5 were the MK-0616 administration group, 2451 administration group, 205A administration group, and 2416 administration group, respectively. The drugs were administered orally at a dose of 20 mg / kg once daily for 14 consecutive days. The results are shown in Table 7 below.

[0533] Table 7. Blood lipid statistics for each group

[0534] Results: In the golden hamster hyperlipidemia model, on day 17 after the first administration, compared with the solvent control group, the low-density lipoprotein (LDL-C) levels in the MK-0616 administration group, 2451 administration group, 205A administration group, and 2416 administration group decreased by 20.2%, 42.0%, 21.7%, and 24.7%, respectively; and the triglyceride (TG) levels decreased by 40.8%, 54.4%, 42.1%, and 41.6%, respectively.

[0535] [Correction based on Rule 91, 04.03.2025] Compared with the solvent control group, the LDL-C and TG of animals in the 2451 administration group were significantly reduced (P<0.05), while the LDL-C and TG of animals in the other administration groups showed a decreasing trend, but no significant difference was observed; compared with the MK-0616 administration group, the LDL-C of animals in the 2451 administration group was significantly reduced (P<0.05).

[0536] Conclusion: In the golden hamster hyperlipidemia model, compared with the solvent control group, all drug administration groups could reduce LDL-C and TG in animals. The 2451 drug administration group showed the most significant reduction effect, which was significantly better than the solvent control group. The reduction effect on LDL-C was significantly better than that of the MK-0616 drug administration group.

Claims

1. The compound represented by Formula I or a pharmaceutically acceptable salt thereof: in, R1 is selected from -CH3, -C(O)-(CH2)n1-N + (CH3)3,-C(O)-(CH2CH2O)n2-CH2CH2N + (CH3)3, n1 is selected from integers from 1 to 14, and n2 is selected from integers from 1 to 5; R1' and R1” are independently selected from none, hydrogen, and -CH3, respectively; R2 is selected from hydrogen, halogen, -SCH3, -SCH2X, -SCHX2, -SCX3; R3 is selected from hydrogen or halogens; A is selected from: R4 and R5 are independently selected from single-chain, C1-C5 straight-chain alkyl and phenyl groups, respectively. R4 and R5 are not both single-chain, nor are they both phenyl groups. R 41 R 42 Each is independently selected from unsubstituted, hydrogen-containing, deuterium-containing, substituted or unsubstituted C1-C3 alkyl groups, halogens, or R. 41 R 42 The carbon atom attached to it forms a C3-C5 cycloalkyl group; R 51 R 52 Each is independently selected from unsubstituted, hydrogen-containing, deuterium-containing, substituted or unsubstituted C1-C3 alkyl groups, halogens, or R. 51 R 52 The carbon atom attached to it forms a C3-C5 cycloalkyl group; n3 is selected from 0 or integers from 1 to 4; n4 is selected from 0 or 1; In the substituted C1-C3 alkyl group, the substituent is selected from deuterium and halogen.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: When R1 is not -CH3, either R1' or R1" is selected as none, and the other is selected as hydrogen; or, when R1 is -CH3, both R1' and R1" are -CH3.

3. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: R1 is selected from -C(O)-(CH2)n1-N + (CH3)3 or -C(O)-(CH2CH2O)n2-CH2CH2N + (CH3)3.

4. The compound according to claim 1 or 3, or a pharmaceutically acceptable salt thereof, characterized in that: R2 is selected from hydrogen, -SCHX2, and -SCX3.

5. The compound according to claim 1 or 4, or a pharmaceutically acceptable salt thereof, characterized in that: R3 is selected from hydrogen.

6. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: When R4 is a one-key, R 41 R 42 It is none; or, when R5 is a one-key, R 51 R 52 It is none; Alternatively, when R4 is a straight-chain alkyl group, R 41 R 42 It is hydrogen, deuterium, substituted or unsubstituted C1-C3 alkyl, halogen, or R 41 R 42 Carbon atoms attached to the same carbon atom and bonded to it form a C3-C5 cycloalkyl group; or, when R5 is a straight-chain alkyl group, R... 51 R 52 It is hydrogen, deuterium, substituted or unsubstituted C1-C3 alkyl, halogen, or R 51 R 52 Carbon atoms attached to the same carbon atom and bonded to it form C3-C5 cycloalkyl groups; Or, when R4 is phenyl, R 41 R 42 It is hydrogen, deuterium, substituted or unsubstituted C1-C3 alkyl, or halogen; or, when R5 is phenyl, R 51 R 52 It is hydrogen, deuterium, substituted or unsubstituted C1-C3 alkyl, or halogen.

7. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: R4 and R5 are independently selected from mono-bonded, C1-C2 straight-chain alkyl, and phenyl groups, respectively; R4 and R5 are not both mono-bonded, nor are they both phenyl groups.

8. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: R 41 R 42 They are independently selected from none, H, D, -CH3, halogen, -CH2X, -CHX2, -CX3, -CH2D, -CHD2, and -CD3, respectively. R 51 R 52 Each of the following is independently selected from: none, H, D, -CH3, halogen, -CH2X, -CHX2, -CX3, -CH2D, -CHD2, -CD 3。 9. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: A is selected from one of the following structures: (a1) (a2) (a3) 10. The compound according to claim 1 or 9, or a pharmaceutically acceptable salt thereof, characterized in that: R 41 R 42 Each element is independently selected from hydrogen, methyl, or halogen; R 51 R 52 Each is independently selected from hydrogen.

11. The compound according to claim 1 or 9, or a pharmaceutically acceptable salt thereof, characterized in that: The halogen is selected from F.

12. The compound or a pharmaceutically acceptable salt thereof according to claim 1 or 9, characterized in that: The n3 is selected from 0, 1 or 2.

13. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: The compound's structural formula is selected from one of the following:

14. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, characterized in that: The compound is selected from one of the following:

15. A pharmaceutical composition, characterized in that, It includes the compound as described in any one of claims 1 to 14 or a pharmaceutically acceptable salt thereof.

16. The pharmaceutical composition according to claim 15, characterized in that, The pharmaceutical composition is selected from oral preparations and injectable preparations.

17. A method for inhibiting PCSK9, comprising administering to a patient an effective dose of the compound of any one of claims 1 to 14 or a pharmaceutically acceptable salt thereof.

18. [Correction 04.03.2025 according to Rule 91] The method according to claim 17, characterized in that, The patient was diagnosed with hypercholesterolemia.

19. A method of treating a disease by administering to a patient an effective dose of the compound of any one of claims 1 to 14 or a pharmaceutically acceptable salt thereof; said disease is selected from atherosclerosis, and / or hypercholesterolemia, and / or coronary heart disease, and / or metabolic syndrome, and / or acute coronary syndrome.

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