Phthalazinone compounds, methods of making, pharmaceutical compositions, and uses thereof

Abstract

The application discloses a phthalazinone compound as shown in formula I and a preparation method, a pharmaceutical composition and an application thereof. The compound has excellent inhibitory activity on polyadenylic diphosphate ribopolymerase and nicotinamide phosphate ribose transferase, has significant inhibitory activity on tumors with BRAC1 / 2 mutation and tumors without BRAC1 / 2 mutation, has small toxic side effects, and has good safety.

Description

Technical Field

[0001] This invention relates to an phthalazinone compound, its preparation method, pharmaceutical composition, and application, and more particularly to an phthalazinone compound having dual inhibitory activities of poly(ADP-ribose) polymerase inhibitor and nicotinamide phosphoribosyltransferase inhibitor, its preparation method, pharmaceutical composition, and application. Background Technology

[0002] Poly(ADP-ribose) polymerase (PARP) catalyzes the ADP-ribose transfer of nicotinamide adenine dinucleotide to its receptor protein, thereby catalyzing poly(ADP-ribosylation) of the protein. Therefore, PARP plays a crucial role in single-stranded DNA repair. The PARP protein family consists of 18 members, whose catalytic domains exhibit high similarity. With the participation of PARP, broken single-stranded DNA can be repaired, allowing cells to survive. When PARP is inhibited by its inhibitors, the unrepaired single-stranded DNA leads to double-stranded DNA breaks. Under normal circumstances, homologous recombination repair mechanisms can repair broken double-stranded DNA, allowing cells to continue to survive. However, when homologous recombination repair is deficient in cells, i.e., when enzymes involved in the HR repair pathway, such as BRAC1 / 2, are lacking, cells die due to the inability to repair DNA damage. This is the synthetic lethal mechanism of PARP inhibitors and HRD.

[0003] NAD + Nicotinamide is a cofactor for many redox enzymes in the body, thus participating in energy metabolism processes, and is also a substrate for key enzymes in many signal transduction processes. The salvage synthesis pathway using nicotinamide as a starting material involves the body's replenishment of NAD. + The most important and direct source is NAD+, and nicotinamide phosphoribosyltransferase is the key rate-limiting enzyme catalyzing this process. Tumor cells have a much higher proliferation and energy demand than normal cells; therefore, tumor cells have a high reliance on NAD+. + It is more dependent on NAMPT and more sensitive to changes in NAMPT activity. In addition to participating in energy metabolism processes, NAMPT plays a key role in cancer stem cells, epithelial-mesenchymal transition and metastasis, DNA repair, signal transduction, and tumor immune escape.

[0004] Currently, olaparib, rucaparib, niraparib, tapolaparib, fluzoparib, and pamiparib, as small molecule PARP inhibitors, are used to treat ovarian, breast, pancreatic, and prostate cancers carrying BRAC mutations. Although PARP inhibitors have significantly improved the prognosis of various HRD-positive malignancies, their indications are narrow, and the patient population that benefits is limited. They are only effective against HRD-positive tumors that can induce synthetic lethality, thus limiting their clinical application. Furthermore, a significant proportion of HRD-positive tumors exhibit innate resistance to PARP inhibitors or develop acquired resistance after a period of treatment. In addition, FK866 and CHS828, as small molecule inhibitors of NAMPT, had their further clinical trials terminated due to poor therapeutic efficacy and dose-dependent toxicity. Summary of the Invention

[0005] Objectives of the Invention: The first objective of this invention is to provide a phthalazinone compound capable of inhibiting the activity of poly(ADP-ribose) polymerase and nicotinamide phosphoribosyltransferase; the second objective is to provide a method for preparing the compound; the third objective is to provide a pharmaceutical composition containing the compound; and the fourth objective is to provide a pharmaceutical application of the compound and the pharmaceutical composition thereof.

[0006] Technical solution: This invention relates to phthalazinone compounds of Formula I or their pharmaceutically acceptable salts, enantiomers, diastereomers, cis-trans isomers, tautomers, trans-blocking isomers, conformational isomers, metabolites, prodrugs, solvates, deuterated derivatives, and hydrates.

[0007]

[0008] In Formula I:

[0009] X is N or C;

[0010] L1 is a single bond or

[0011] Y is C 6-10 Aryl or 5- to 10-membered heteroaryl containing 1 to 3 N, O, or S heteroatoms;

[0012] L2 is -(CH2) n -, where n is 0, 1, 2, or 3;

[0013] R1 is a 9- to 10-membered bicyclic heteroaryl group containing one N atom and one to two N, O, and S heteroatoms.

[0014] The compounds described in this invention can exist in specific stereoisomer forms. The term "isomer" refers to isomers with the same structure but different spatial arrangements of atoms. These include cis and trans (or Z and E) isomers, (-)- and (+)- isomers, (R)- and (S)- enantiomers, diastereomers, (D)- and (L)- isomers, tautomers, blocked isomers, conformational isomers, and mixtures thereof (such as racemic mixtures and mixtures of diastereomers). Substituents in the compounds disclosed herein may contain additional asymmetric atoms. All such stereoisomers and mixtures thereof are included within the scope of this disclosure. Optically active (-)- and (+)- isomers, (R)- and (S)- enantiomers, and (D)- and (L)- isomers can be prepared by chiral synthesis, chiral reagents, or other conventional techniques. This disclosure discloses an isomer of a compound, which can be prepared by asymmetric synthesis or with chiral auxiliaries, or, when the molecule contains a basic functional group (such as an amino group) or an acidic functional group (such as a carboxyl group), by forming a salt of the diastereomer with a suitable optically active acid or base, followed by diastereomer resolution using conventional methods known in the art to obtain the pure isomer. Furthermore, the separation of enantiomers and diastereomers is typically performed by chromatography.

[0015] The term "pharmaceutically acceptable salt" refers to a salt of the compounds of this invention, prepared by reacting a compound having specific substituents discovered in this invention with a relatively non-toxic acid or base. When the compounds of this invention contain relatively acidic functional groups, base addition salts can be obtained by contacting the free form of such compounds with a sufficient amount of base in a pure solution or a suitable inert solvent. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amine, or magnesium salts or similar salts. When the compounds of this invention contain relatively basic functional groups, acid addition salts can be obtained by contacting the free form of such compounds with a sufficient amount of acid in a pure solution or a suitable inert solvent. Examples of pharmaceutically acceptable acid addition salts include inorganic acid salts, such as hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid (forming carbonates or bicarbonates), phosphoric acid (forming phosphates, monohydrogen phosphates, dihydrogen phosphates, sulfuric acid (forming sulfates or bisulfates), hydroiodic acid, phosphorous acid, etc.); and organic acid salts, such as acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, octanoic acid, fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, etc. Acids such as citric acid, tartaric acid, and methanesulfonic acid; organic acid salts also include salts of organic acids such as amino acids (e.g., arginine), glucuronic acid, etc. Certain specific compounds of the present invention contain basic and acidic functional groups, thus allowing them to be converted into any base or acid addition salt. Preferably, the salt is contacted with a base or acid in a conventional manner, and then the parent compound is separated, thereby regenerating the free form of the compound. The free form of the compound differs from its various salt forms in certain physical properties, such as different solubilities in polar solvents.

[0016] The pharmaceutically acceptable salts of the present invention can be synthesized from parent compounds containing acid radicals or bases by conventional chemical methods. Generally, such salts are prepared by reacting these compounds in free acid or base form with a stoichiometric amount of a suitable base or acid in water or an organic solvent or a mixture of both. Non-aqueous media such as ethers, ethyl acetate, ethanol, isopropanol, or acetonitrile are generally preferred.

[0017] The term "metabolite" refers to a pharmaceutically active product produced in vivo by the metabolism of a compound of Formula I or a salt thereof. Such products can be generated, for example, by oxidation, reduction, hydrolysis, amidation, deamidation, esterification, deesterification, glucuronidation, enzymatic cleavage, etc., of the administered compound. Therefore, this invention includes metabolites of the compounds of this invention, including compounds produced by methods that expose the compounds of this invention to mammals for a sufficient period of time to obtain their metabolites.

[0018] The identification of metabolites is typically performed by preparing a radiolabeled isotope of the compound of the invention, administering it parenterally to an animal, such as a rat, mouse, guinea pig, monkey, or human, at a detectable dose (e.g., greater than about 0.5 mg / kg), allowing sufficient time for metabolism to occur (typically about 30 seconds to 30 hours), and isolating the metabolites from urine, blood, or other biological samples. These products are readily isolated because they are labeled (others are isolated using antibodies capable of binding to antigenic epitopes present in the metabolites). The metabolite structure is determined in a conventional manner, for example, by MS, LC / MS, or NMR analysis. Typically, the analysis of metabolites is performed using methods known to those skilled in the art for routine drug metabolism studies. The metabolite products can be used for assays of therapeutic doses of the compound of the invention, provided they are not otherwise undetectable in vivo. The compounds of the invention may contain atomic isotopes in non-natural proportions on one or more atoms constituting the compound. For example, the compound may be radiolabeled with radioisotopes, such as tritium ( 3 H), Iodine-125 125 I) or C-14 14 C). All isotopic variations of the compounds of the present invention, regardless of radioactivity, are included within the scope of the present invention.

[0019] In addition to the salt form, the compounds provided by this invention also exist in prodrug form. The prodrugs of the compounds described herein readily undergo chemical changes under physiological conditions to transform into the compounds of this invention. Any compound that can be converted in vivo to provide a bioactive substance (i.e., the compound shown in Formula I) is a prodrug within the scope and spirit of this invention. For example, compounds containing a carboxyl group can form physiologically hydrolyzable esters, which act as prodrugs by hydrolysis in vivo to yield the compound shown in Formula I itself. The prodrugs are preferably administered orally because hydrolysis in many cases occurs primarily under the influence of digestive enzymes. Parenteral administration may be used when the ester itself is active or when hydrolysis occurs in the bloodstream.

[0020] The term "deuterated compound" refers to a molecule in which any hydrogen atom can be replaced by its isotope deuterium. In the compounds involved in this invention, any hydrogen atom can be replaced by a deuterium atom.

[0021] The term "aryl" refers to an aromatic group having a specified number of carbon atoms, preferably a monocyclic, bicyclic, or tricyclic aromatic group. When it is bicyclic or tricyclic, each ring satisfies Hückel's rule. The present invention's C... 6-10 The aryl group refers to an aromatic group containing 6 to 10 carbon atoms, such as phenyl.

[0022] The term "heteroaryl" refers to a 9- to 10-membered bicyclic heteroaryl group containing one nitrogen atom and one or two other heteroatoms selected from nitrogen, oxygen, and sulfur. This includes bicyclic groups formed by the fusion of an unsaturated monocyclic ring containing a heteroatom and an aryl group, bicyclic groups formed by the fusion of two unsaturated monocyclic rings containing heteroatoms, and bicyclic groups formed by the fusion of a saturated monocyclic ring containing a heteroatom and a heteroaryl group or aryl group. When the "heteroaryl" is a bicyclic group formed by the fusion of a saturated monocyclic ring containing a heteroatom and a heteroaryl group, it is linked to other segments or groups in the compound shown in Formula I through the saturated monocyclic ring containing the heterocycle. The 9- to 10-membered bicyclic groups include, but are not limited to, imidazopyridine, imidazopyrimidine, pyrazolopyridine, thienopyridine, furanopyridine, tetrahydropyrrolopyridine, or tetrahydropyrrolophenyl.

[0023] Those skilled in the art will understand that, according to conventions used in the art, the structural formulas of the groups described in this invention are... This refers to the connection of the corresponding group to other segments or groups in the compound shown in Formula I through this site.

[0024] Preferably, in formula I:

[0025] Y is phenyl or pyridinyl;

[0026] n is 0, 1, or 2.

[0027] Preferably, in formula I:

[0028] Y represents phenyl,

[0029] n is 0 or 1.

[0030] Preferably, in formula I:

[0031] R1 is pyridinium imidazolyl, pyridinium pyrazolyl, pyridinium thiophene, pyridinium furanyl, pyridinium tetrahydropyrrole, or phenyl tetrahydropyrrole.

[0032] Preferably, in formula I:

[0033] R1 is

[0034]

[0035] Preferably, the compounds of the present invention represented by Formula I are selected from any one of the following compounds:

[0036]

[0037] NAMPT inhibitors inhibit NAD +The generation of NAMPT inhibitors inhibits the substrate supply of PARP and enhances the competitive inhibition of the PARP catalytic domain by PARP inhibitors. Therefore, NAMPT inhibitors can increase DNA damage, thereby increasing the antitumor activity of PARP inhibitors. NAMPT activity can reduce BRAC1 / 2 expression, reduce RAD51 expression, and induce HRD. NAMPT inhibitors and PARP1 inhibitors exhibit synergistic antitumor effects in BRAC wild-type triple-negative breast cancer and Ewing sarcoma. However, combination therapy has drawbacks such as potential drug-drug interactions, unclear pharmacokinetic properties, and poor patient compliance. Dual-target or multi-target drugs can reduce toxicity, increase efficacy, and avoid the drawbacks of combination therapy. Therefore, developing dual-target inhibitors of PARP and NAMPT can improve the narrow indications of PARP inhibitors, reduce the significant toxic side effects of NAMPT inhibitors, and address drug resistance issues.

[0038] To address the limitations of poly(ADP-ribose) polymerase inhibitors and the dose-dependent nature of nicotinamide phosphoribosyltransferase inhibitors, this invention designs a dual inhibitor of phthalazinone-type poly(ADP-ribose) polymerase and nicotinamide phosphoribosyltransferase. This inhibitor demonstrates significant therapeutic effects on both tumors carrying and not carrying BRAC1 / 2 mutations, with minimal toxic side effects.

[0039] The present invention provides a method for preparing phthalazinone compounds of Formula I or their pharmaceutically acceptable salts, enantiomers, diastereomers, cis-trans isomers, tautomers, trans-blocking isomers, conformational isomers, metabolites, prodrugs, solvates, deuterated derivatives, and hydrates, comprising the following steps:

[0040] Compound II and compound III undergo a condensation reaction in the presence of a base and a condensing agent to yield compound I:

[0041]

[0042] Among them, X, Y, L1, L2, and R1 are as described above;

[0043] The solvents mentioned in the above reactions include, but are not limited to, N,N-dimethylformamide, N,N-dimethylacetamide, dimethyl sulfoxide, dichloromethane, N-methylpyrrolidone, or mixtures thereof;

[0044] The base mentioned in the above reaction is an organic base, including but not limited to triethylamine, pyridine, and N,N-diisopropylethylamine (DIPEA); preferably triethylamine or N,N-diisopropylethylamine.

[0045] The condensing agents mentioned in the above reactions include, but are not limited to, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride, N,N'-dicyclohexylcarbodiimide (DCC), N,N'-diisopropylcarbodiimide (DIC), O-benzotriazole-N,N,N',N'-tetramethylurea tetrafluoroborate (TBTU), 1-hydroxybenzotriazole (HOBt), 1-hydroxy-7-azobenzotriazole (HOAt), and O -Benztriazole-N,N,N',N'-Tetramethylurea hexafluorophosphate (HBTU), 2-(7-azobenzotriazole)-N,N,N',N'-Tetramethylurea hexafluorophosphate (HATU), benzotriazole-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate (BOP) or benzotriazole-1-yl-oxytripyrrolylphosphonium hexafluorophosphate (PyBOP); preferably EDCI, HATU, BOP or PyBOP.

[0046] The present invention relates to the use of phthalazinone compounds of Formula I or pharmaceutically acceptable salts thereof, enantiomers, diastereomers, cis-trans isomers, tautomers, transisomers, conformational isomers, metabolites, prodrugs, solvates, deuterates, and hydrates in the preparation of pharmaceuticals for poly(ADP-ribose) polymerase inhibitors and / or nicotinamide phosphoribosyltransferase inhibitors.

[0047] The present invention relates to the use of phthalazinone compounds of Formula I or pharmaceutically acceptable salts thereof, enantiomers, diastereomers, cis-trans isomers, tautomers, transisomers, conformational isomers, metabolites, prodrugs, solvates, and hydrates in the preparation of medicaments for the treatment and / or prevention of cancer.

[0048] Preferably, the drug is one or more of the following: bone cancer, lung cancer, gastric cancer, colorectal cancer, endometrial cancer, breast cancer, ovarian cancer, glioma, prostate cancer, pancreatic cancer, liver cancer, brain cancer, bladder cancer, cervical cancer, testicular cancer, kidney cancer, head and neck cancer, thyroid cancer, esophageal cancer, lymphoma, leukemia, or skin cancer.

[0049] The pharmaceutical compositions of the present invention contain phthalazinone compounds as shown in Formula I or pharmaceutically acceptable salts thereof, enantiomers, diastereomers, cis-trans isomers, tautomers, transisomers, conformational isomers, metabolites, prodrugs, solvates, deuterates, hydrates, and pharmaceutically acceptable carriers or excipients.

[0050] Preferably, in the pharmaceutical composition, the amount of the phthalazinone compound or its pharmaceutically acceptable salt, enantiomer, diastereomer, cis-trans isomer, tautomer, transisomer, conformational isomer, metabolite, solvate, deuterated product, or hydrate is a therapeutically effective amount.

[0051] The pharmaceutical composition described in this invention is used in the preparation of pharmaceuticals for poly(adenosine diphosphate ribose) polymerase inhibitors and / or nicotinamide phosphoribosyltransferase inhibitors.

[0052] The pharmaceutical composition described in this invention is used in the preparation of medicaments for treating and / or preventing cancer.

[0053] Preferably, the cancer drug is one or more of the following: bone cancer, lung cancer, stomach cancer, colorectal cancer, endometrial cancer, breast cancer, ovarian cancer, glioma, prostate cancer, pancreatic cancer, liver cancer, brain cancer, bladder cancer, cervical cancer, testicular cancer, kidney cancer, head and neck cancer, thyroid cancer, esophageal cancer, lymphoma, leukemia, or skin cancer.

[0054] The pharmaceutically acceptable carrier can be an excipient widely used in the pharmaceutical manufacturing industry. Excipients primarily serve to provide a safe, stable, and functional pharmaceutical composition, and may also provide methods to allow the active ingredient to dissolve at a desired rate after administration to a subject, or to promote the effective absorption of the active ingredient after administration to a subject. The pharmaceutical excipient can be an inert filler, or it may provide a function such as stabilizing the overall pH of the composition or preventing the degradation of the active ingredient. The pharmaceutical excipient may include one or more of the following: binders, suspending agents, emulsifiers, diluents, fillers, granulators, adhesives, disintegrants, lubricants, anti-adhesion agents, flow aids, wetting agents, gelling agents, absorption delay agents, dissolution inhibitors, enhancers, adsorbents, buffers, chelating agents, preservatives, colorants, flavoring agents, and sweeteners.

[0055] The pharmaceutical compositions of the present invention can be prepared using any method known to those skilled in the art, based on the disclosure. For example, conventional mixing, dissolving, granulation, emulsification, grinding, encapsulation, embedding, or lyophilization processes.

[0056] The pharmaceutical compositions of this invention can be administered in any form, including by injection (intravenous), mucosal, oral (solid and liquid formulations), inhalation, ocular, rectal, topical, or parenteral (infusion, injection, implantation, subcutaneous, intravenous, intra-arterial, intramuscular) administration. The pharmaceutical compositions of this invention can also be controlled-release or sustained-release dosage forms (e.g., liposomes or microspheres). Examples of solid oral formulations include, but are not limited to, powders, capsules, tablets, soft capsules, and tablets. Examples of liquid formulations for oral or mucosal administration include, but are not limited to, suspensions, emulsions, elixirs, and solutions. Examples of topical formulations include, but are not limited to, emulsions, gels, ointments, creams, patches, pastes, foams, lotions, drops, or serum preparations. Examples of parenteral formulations include, but are not limited to, solutions for injection, dry powder formulations that can be dissolved or suspended in a pharmaceutically acceptable carrier, suspensions for injection, and emulsions for injection. Examples of other suitable formulations of the pharmaceutical composition include, but are not limited to, eye drops and other ophthalmic preparations; aerosols, such as nasal sprays or inhalers; liquid dosage forms suitable for parenteral administration; suppositories; and tablets.

[0057] Without violating common sense in the field, the above-mentioned preferred conditions can be combined arbitrarily to obtain various preferred embodiments of the present invention.

[0058] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages:

[0059] The compounds designed in this invention exhibit excellent inhibitory activity against both poly(ADP-ribose) polymerase and nicotinamide phosphoribosyltransferase, and also show significant inhibitory activity against both BRAC1 / 2-mutated and non-BRAC1 / 2-mutated tumors, with minimal toxicity and good safety. Detailed Implementation

[0060] The technical solution of the present invention will be further described below with reference to the embodiments.

[0061] Example 1: Synthesis of compound S1

[0062]

[0063] Step 1: Synthesis of Intermediate 7

[0064]

[0065] a) Compound 1 (3.5 g, 11.25 mmol) was dissolved in acetonitrile (34 mL). 4-Methoxythiophenol (2.58 g, 13.73 mmol) and K₂CO₃ (2.33 g, 16.88 mmol) were added to the reaction system. The reaction was carried out at room temperature for 6 hours. After complete reaction, K₂CO₃ was removed by filtration. The mixture was concentrated under reduced pressure and purified by column chromatography to obtain intermediate 2 (2.95 g, 71%). MS (ESI, m / z): 372 (M + +1).

[0066] b) Intermediate 2 (2.95 g, 7.95 mmol) was dissolved in DCM (25 mL), and 3-chloroperoxybenzoic acid (6.86 g, 39.75 mmol) was added at 0 °C. The mixture was transferred to room temperature and reacted for 2 hours. After the reaction was complete, the reaction was cooled to 0 °C, and quenched by slowly adding saturated sodium thiosulfate solution (80 mL). The mixture was extracted with dichloromethane (30 mL × 3), and the organic phases were combined. The mixture was washed twice with 1 M NaOH aqueous solution and twice with saturated sodium chloride solution. The mixture was dried over anhydrous Na2SO4, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain intermediate 3 (2.78 g, 87%). MS (ESI, m / z): 404 (M + +1).

[0067] c) Intermediate 3 (2.78 g, 6.90 mmol) was dissolved in DMF (28 mL). Tetra(triphenylphosphine)palladium (797 mg, 0.69 mmol) and zinc cyanide (956 mg, 8.14 mmol) were added to the reaction system. The reaction was carried out at 85 °C for 7 hours. After the reaction was complete, the mixture was diluted with water, filtered, and the filter cake was purified by column chromatography to obtain intermediate 4 (1.5 g, 62%). MS (ESI, m / z): 351 (M + +1).

[0068] d) Intermediate 4 (1.5 g, 4.29 mmol) was dissolved in MeOH / THF (13 mL / 13 mL). Nickel chloride hexahydrate (1.02 g, 4.29 mmol) was added to the reaction system, and sodium borohydride (243 mg, 6.44 mmol) was slowly added at 0 °C. The temperature was raised to 60 °C and the reaction was carried out for 12 hours. After the reaction was complete, the mixture was filtered through diatomaceous earth. The filtrate was cooled to 0 °C, and sodium borohydride was quenched with 2 M HCl aqueous solution. The mixture was extracted with ethyl acetate (20 mL × 3). The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous Na₂SO₄, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain intermediate 5 (600 mg, 40%). MS (ESI, m / z): 355 (M + +1).

[0069] e) Intermediate 5 (120 mg, 0.34 mmol) was dissolved in DMF (1 mL). Imidazolo[1,2-A]pyridine-7-carboxylic acid (55 mg, 0.34 mmol), BOP (225 mg, 0.51 mmol), and DIPEA (131 mg, 1.02 mmol) were added to the reaction system. The reaction was carried out at room temperature for 16 hours. After the reaction was complete, water was added for dilution, and the mixture was extracted with ethyl acetate (10 mL × 3). The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous Na2SO4, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain intermediate 6 (140 mg, 83%). MS (ESI, m / z): 499 (M + +1).

[0070] f) Intermediate 6 (140 mg, 0.28 mmol) was dissolved in DCM (1 mL), and TFA (0.5 mL) was added to the reaction system. The reaction was carried out at room temperature for 1 hour. After the reaction was complete, dichloromethane was removed by vacuum distillation. The solution was diluted with water, and the pH was adjusted to 8-10 at 0 °C with 2 M NaOH aqueous solution. The solution was extracted with dichloromethane, and the organic phases were combined, dried over anhydrous Na2SO4, filtered to remove salts, and concentrated under reduced pressure to obtain intermediate 7 (45 mg, 41%). MS (ESI, m / z): 399 (M + +1).

[0071] Step 2: Synthesis of compound S1

[0072]

[0073] a) Intermediate 7 (45 mg, 0.11 mmol) and compound 8 (21 mg, 0.11 mmol) were dissolved in DMF (1 mL). BOP (75 mg, 0.17 mmol) and DIPEA (44 mg, 0.34 mmol) were added to the reaction system. The reaction was carried out at room temperature for 12 hours. After the reaction was complete, the mixture was diluted with water and extracted with ethyl acetate (10 mL × 3). The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous Na2SO4, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain S1 (41 mg, 53%).

[0074] S1: 1HNMR (400MHz, DMSO-d6) δ12.58(s,1H),9.27(t,J=5.9Hz,1H),9.18(s,1H),8.26(d,J=7.6Hz,1H),8.09(s,1H),7.97( d,J=7.9Hz,1H),7.88(t,J=7.3Hz,1H),7.83(dd,J=8.1,2.7Hz,3H),7.73-7.61(m,5H),7.45-7.37(m,1H),7.33(s,1H) ,7.20(t,J=9.0Hz,1H),4.63(d,J=5.8Hz,2H),4.55(d,J=13.2Hz,1H),4.32(s,2H),3.56(t,J=12.1Hz,1H),3.01(t,J= 13.0Hz,1H),2.75(t,J=12.6Hz,1H),1.93(d,J=12.9Hz,1H),1.77(s,1H),1.41(d,J=12.9Hz,2H).MS(ESI,m / z):679(M + +1).

[0075] Example 2: Synthesis of compound S2

[0076]

[0077] The synthesis method is the same as in Example 1, except that the corresponding raw materials can be replaced to obtain compound S2.

[0078] S2: 1HNMR (400MHz, DMSO-d6) δ13.93(s,1H),12.60(s,1H),9.37(t,J=6.0Hz,1H),9.04(d,J=2.1Hz,1H),8.78(d,J=2.1Hz,1H),8.31(s ,1H),8.30-8.21(m,1H),7.96(d,J=7.9Hz,1H),7.91-7.78(m,4H),7.65(d,J=8.0Hz,2H),7.44-7.37(m,1H),7.33(s,1H),7.20(t ,J=9.0Hz,1H),4.65(d,J=5.8Hz,2H),4.55(d,J=13.0Hz,1H),4.32(s,2H),3.56(t,J=23.1Hz,1H),3.01(t,J=26.7,12.5Hz,1H), 3.01(d,J=25.7Hz,1H),2.75(t,J=12.5Hz,1H),1.93(d,J=12.6Hz,1H),1.77(s,1H),1.39(d,J=23.7Hz,2H).MS(ESI,m / z):680(M + +1).

[0079] Example 3: Synthesis of compound S3

[0080]

[0081] Step 1: Synthesis of Intermediate 10

[0082]

[0083] a) Intermediate 5 (612 mg, 1.72 mmol), N,N'-disuccinimidyl carbonate (530 mg, 2.07 mmol), and pyridine (136 mg, 1.72 mmol) were dissolved together in anhydrous acetonitrile (8.6 mL), and a solution of 2,3-dihydro-1H-pyrrolo[3,4-C]pyridine was added dropwise at 0 °C. The 2,3-dihydro-1H-pyrrolo[3,4-C]pyridine solution was prepared by dissolving 2,3-dihydro-1H-pyrrolo[3,4-C]pyridine (270 mg, 1.72 mmol) and DIPEA (446 mg, 3.45 mmol) together in anhydrous acetonitrile / DMF (0.95 mL / 0.48 mL). The two solutions were mixed and reacted at room temperature for 12 hours. After complete reaction, the mixture was diluted with water, extracted with ethyl acetate (15 mL × 3), and the organic phases were combined. The mixture was washed twice with saturated sodium chloride solution, dried over anhydrous Na₂SO₄, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain intermediate 9 (450 mg, 52%). MS (ESI, m / z): 501 (M+ +1).

[0084] b) Intermediate 9 (450 mg, 0.90 mmol) was dissolved in DCM (2 mL), and TFA (1 mL) was added to the reaction system. The reaction was carried out at room temperature for 1 hour. After the reaction was complete, dichloromethane was removed by vacuum distillation. The solution was diluted with water, and the pH was adjusted to 8-10 at 0 °C with 2 M NaOH aqueous solution. The solution was extracted with dichloromethane, and the organic phases were combined, dried over anhydrous Na2SO4, filtered to remove salts, and concentrated under reduced pressure to obtain intermediate 10 (295 mg, 82%). MS (ESI, m / z): 401 (M + +1).

[0085] Step 2: Synthesis of compound S3

[0086]

[0087] a) Intermediate 10 (75 mg, 0.19 mmol) and compound 8 (56 mg, 0.19 mmol) were dissolved in DMF (1 mL). BOP (124 mg, 0.28 mmol) and DIPEA (73 mg, 0.56 mmol) were added to the reaction system. The reaction was carried out at room temperature for 12 hours. After the reaction was complete, the mixture was diluted with water and extracted with ethyl acetate (10 mL × 3). The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous Na2SO4, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain S3 (57 mg, 45%).

[0088] S3: 1 H NMR (400MHz, DMSO-d6) δ12.60(s,1H),8.59(s,1H),8.48(d,J=5.0Hz,1H),8.29-8.23(m,1H),7.97(d,J=7.9Hz,1H) ,7.91-7.75(m,4H),7.61(d,J=8.1Hz,2H),7.41(d,J=5.4Hz,2H),7.33(s,1H),7.27-7.13(m,2H),4.69(d,J=7.8Hz, 4H),4.55(d,J=13.3Hz,1H),4.42(d,J=5.8Hz,2H),4.32(s,2H),3.55(t,J=12.2Hz,1H),3.41(s,1H),3.01(t,J=12 .9Hz,1H),2.75(t,J=12.7Hz,1H),1.93(d,J=12.8Hz,1H),1.77(s,1H),1.41(d,J=24.3Hz,2H).MS(ESI,m / z):681(M + +1).

[0089] Example 4: Synthesis of compound S4

[0090]

[0091] The synthesis method is the same as in Example 1, except that the corresponding raw materials can be replaced to obtain compound S4.

[0092] S4: 1 H NMR (300MHz, DMSO-d6) δ12.59(s,1H),12.16(s,1H),9.42(t,J=6.1Hz,1H),8.80(s,1H),8.25(dd,J=7.3,1.7H z,1H),8.13(d,J=5.6Hz,1H),8.00-7.92(m,1H),7.92-7.78(m,4H),7.69-7.58(m,3H),7.46-7.35(m,1H),7.3 2(s,1H),7.27-7.15(m,2H),4.65(d,J=5.9Hz,2H),4.54(d,J=13.2Hz,1H),4.31(s,2H),3.55(s,1H),3.00(s, 1H),2.74(t,J=12.4Hz,1H),1.92(d,J=12.7Hz,1H),1.76(s,1H),1.37(d,J=15.1Hz,2H).MS(ESI,m / z):679(M + +1).

[0093] Example 5: Synthesis of compound S5

[0094]

[0095] The synthesis method is the same as in Example 1, except that the corresponding raw materials can be replaced to obtain compound S5.

[0096] S5: 1HNMR (300MHz, DMSO-d6) δ12.60(s,1H),12.16(s,1H),9.42(t,J=6.1Hz,1H),8.81(s,1H),8.24(dd,J=7.3,1.7 Hz,1H),8.13(d,J=5.6Hz,1H),7.99-7.90(m,1H),7.94-7.79(m,4H),7.67-7.56(m,3H),7.48-7.35(m,1H),7. 30(s,1H),7.29-7.14(m,2H),4.65(d,J=5.9Hz,2H),4.55(d,J=13.2Hz,1H),4.30(s,2H),3.56(s,1H),3.02(s ,1H),2.76(t,J=12.4Hz,1H),1.93(d,J=12.7Hz,1H),1.77(s,1H),1.39(d,J=15.1Hz,2H).MS(ESI,m / z):679(M + +1).

[0097] Example 6: Synthesis of compound S6

[0098]

[0099] The synthesis method is the same as in Example 1, except that the corresponding raw materials can be replaced to obtain compound S6.

[0100] S6: 1 H NMR (400MHz, DMSO-d6) δ12.60(s,1H),9.67(t,J=6.0Hz,1H),9.33(s,1H),8.54(d,J=5.4Hz,1H),8.26(d,J=7. 8Hz,1H),8.21(s,1H),7.96(t,J=6.0Hz,2H),7.93-7.77(m,4H),7.65(d,J=8.1Hz,2H),7.40(t,J=6.5Hz,1H),7 .32(s,1H),7.20(t,J=9.0Hz,1H),4.64(d,J=5.9Hz,2H),4.55(d,J=13.3Hz,1H),4.31(s,2H),3.57(s,1H),3. 01(s,1H),2.75(t,J=12.7Hz,1H),1.93(d,J=12.9Hz,1H),1.77(s,1H),1.56-1.28(m,2H).MS(ESI,m / z):696(M + +1).

[0101] Example 7: Synthesis of compound S7

[0102]

[0103] The synthesis method is the same as in Example 1, except that the corresponding raw materials can be replaced to obtain compound S7.

[0104] S7: 1 HNMR (400MHz, DMSO-d6) δ12.59(s,1H),9.70(t,J=6.0Hz,1H),9.38(s,1H),8.53(d,J=5.4Hz,1H),8.27(d,J=7 .8Hz,1H),8.24(s,1H),7.98(t,J=6.0Hz,2H),7.95-7.79(m,4H),7.67(d,J=8.1Hz,2H),7.43(t,J=6.5Hz,1H), 7.32(s,1H),7.22(t,J=9.0Hz,1H),4.67(d,J=5.9Hz,2H),4.55(d,J=13.3Hz,1H),4.32(s,2H),3.56(s,1H),3. 02(s,1H),2.74(t,J=12.7Hz,1H),1.93(d,J=12.9Hz,1H),1.76(s,1H),1.57-1.30(m,2H).MS(ESI,m / z):680(M + +1).

[0105] Example 8: Synthesis of compound S8

[0106]

[0107] The synthesis method is the same as in Example 1, except that the corresponding raw materials can be replaced to obtain compound S8.

[0108] S8: 1HNMR (400MHz, DMSO-d6) δ12.60(s,1H),9.37(t,J=6.0Hz,1H),9.06(d,J=2.1Hz,1H),8.79(d,J=2.1Hz,1H),8.32(s,1H),8.28 -8.20(m,2H),7.98(d,J=7.9Hz,1H),7.89-7.77(m,4H),7.64(d,J=8.0Hz,2H),7.45-7.38(m,1H),7.35(s,1H),7.21(t,J=9.0 Hz,1H),4.63(d,J=5.8Hz,2H),4.57(d,J=13.0Hz,1H),4.32(s,2H),3.58(t,J=23.1Hz,1H),3.01(t,J=26.7,12.5Hz,1H),3.0 1(d,J=25.7Hz,1H),2.75(t,J=12.5Hz,1H),1.93(d,J=12.6Hz,1H),1.78(s,1H),1.35(d,J=23.7Hz,2H).MS(ESI,m / z):680(M + +1).

[0109] Example 9: Synthesis of compound S9

[0110]

[0111] Step 1: Synthesis of Intermediate 15

[0112]

[0113] a) Compound 11 (201 mg, 1.0 mmol) was dissolved in anhydrous DCM (4 mL). 1-tert-butyloxycarbonylpiperazine (220 mg, 1.18 mmol) and TEA (304 mg, 3 mmol) were added to the reaction system. The mixture was reacted at room temperature for 1 hour. After complete reaction, water was added for dilution, followed by extraction with dichloromethane. The organic phases were combined, dried over anhydrous Na₂SO₄, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain intermediate 12 (250 mg, 71%). MS (ESI, m / z): 352 (M + +1).

[0114] b) Intermediate 12 (1.0 g, 2.85 mmol) and nickel chloride hexahydrate (675 mg, 2.85 mmol) were dissolved in MeOH / THF (2 mL / 4 mL). Sodium borohydride (161 mg, 4.27 mmol) was added in portions at 0 °C, and the reaction was carried out at 60 °C for 12 hours. After the reaction was complete, the mixture was filtered through diatomaceous earth. The filtrate was cooled to 0 °C, and sodium borohydride was quenched with 2 M HCl aqueous solution. The mixture was extracted with ethyl acetate (20 mL × 3). The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous Na₂SO₄, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain intermediate 13 (500 mg, 49%). MS (ESI, m / z): 356 (M + +1).

[0115] c) Intermediate 13 (500 mg, 1.41 mmol) was dissolved in DMF (4 mL). Imidazolo[1,2-A]pyridine-7-carboxylic acid (229 mg, 1.41 mmol), BOP (935 mg, 2.12 mmol), and DIPEA (698 mg, 4.23 mmol) were added to the reaction system. The reaction was carried out at room temperature for 10 hours. After the reaction was complete, water was added for dilution, and the mixture was extracted with ethyl acetate (10 mL × 3). The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous Na2SO4, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain intermediate 14 (540 mg, 77%). MS (ESI, m / z): 500 (M + +1).

[0116] d) Intermediate 14 (540 mg, 1.08 mmol) was dissolved in DCM (6 mL), and TFA (3 mL) was added to the reaction system. The reaction was carried out at room temperature for 1 hour. After the reaction was complete, dichloromethane was removed by vacuum distillation. The solution was diluted with water, and the pH was adjusted to 8-10 at 0 °C with 2 M NaOH aqueous solution. The solution was extracted with dichloromethane, and the organic phases were combined, dried over anhydrous Na2SO4, filtered to remove salts, and concentrated under reduced pressure to obtain intermediate 15 (410 mg, 95%). MS (ESI, m / z): 400 (M + +1).

[0117] Step 2: Synthesis of compound S9

[0118]

[0119] a) Intermediate 15 (617 mg, 1.29 mmol) and compound 8 (241 mg, 1.29 mmol) were dissolved in DMF (7 mL). BOP (856 mg, 1.94 mmol) and DIPEA (44 mg, 0.34 mmol) were added to the reaction system. The reaction was carried out at room temperature for 12 hours. After the reaction was complete, the mixture was diluted with water and extracted with ethyl acetate (20 mL × 3). The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous Na2SO4, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain S9 (550 mg, 52%).

[0120] S9: 1 H NMR (400MHz, DMSO-d6) δ12.57(s,1H),9.24(s,1H),9.18(s,H),8.26(d,J=7 .4Hz,1H),8.08(s,1H),7.94-7.81(m,3H),7.74-7.61(m,7H),7.40(s,1H),7 .29(d,J=5.8Hz,1H),7.18(t,J=9.0Hz,1H),4.62(d,J=5.8Hz,2H),4.29(s,2 H),3.69(s,2H),3.25(s,2H),2.96(s,2H),2.82(s,2H).MS(ESI,m / z):680(M + +1).

[0121] Example 10: Synthesis of compound S10

[0122]

[0123] The synthesis method is the same as in Example 3. Compound S10 can be obtained by simply replacing intermediate 5 with intermediate 13.

[0124] S10: 1 H NMR (400MHz, DMSO-d6) δ12.60 (s, 1H), 8.56 (s, 1H), 8.48 (d, J = 5.0Hz, 1H), 8.29-8.2 4(m,1H),7.96(d,J=7.9Hz,1H),7.90-7.74(m,4H),7.63(d,J=8.1Hz,2H),7.42(d,J =5.4Hz,2H),7.33(s,1H),7.28-7.12(m,2H),4.74(s,4H),4.63(d,J=6.8Hz,2H),4. 35(s,2H),3.77(s,2H),3.28(s,2H),3.11(s,2H),2.92(s,2H).MS(ESI,m / z):682(M + +1).

[0125] Example 11: Synthesis of compound S11

[0126]

[0127] Step 1: Synthesis of Intermediate 18

[0128]

[0129] a) Intermediate 16 (200 mg, 0.72 mmol), N,N'-disuccinimidyl carbonate (221 mg, 0.86 mmol), and pyridine (57 mg, 0.72 mmol) were dissolved together in anhydrous acetonitrile (3.6 mL), and a solution of 2,3-dihydro-1H-pyrrolo[3,4-C]pyridine was added dropwise at 0 °C. The 2,3-dihydro-1H-pyrrolo[3,4-C]pyridine solution was prepared by dissolving 2,3-dihydro-1H-pyrrolo[3,4-C]pyridine (86 mg, 0.72 mmol) and DIPEA (186 mg, 1.44 mmol) together in anhydrous acetonitrile / DMF (0.4 mL / 0.2 mL). The two solutions were mixed and reacted at room temperature for 12 hours. After complete reaction, the mixture was diluted with water, extracted with ethyl acetate (10 mL × 3), and the organic phases were combined. The mixture was washed twice with saturated sodium chloride solution, dried over anhydrous Na₂SO₄, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain intermediate 17 (152 mg, 50%). MS (ESI, m / z): 423 (M + +1).

[0130] b) Intermediate 17 (152 mg, 0.36 mmol) was dissolved in DCM (2 mL), and TFA (1 mL) was added to the reaction system. The reaction was carried out at room temperature for 1 hour. After the reaction was complete, dichloromethane was removed by vacuum distillation. The solution was diluted with water, and the pH was adjusted to 8-10 at 0 °C with 2 M NaOH aqueous solution. The solution was extracted with dichloromethane, and the organic phases were combined, dried over anhydrous Na2SO4, filtered to remove salts, and concentrated under reduced pressure to obtain intermediate 18 (105 mg, 90%). MS (ESI, m / z): 323 (M + +1).

[0131] Step 2: Synthesis of compound S11

[0132]

[0133] a) Intermediate 21 (127 mg, 0.29 mmol) and compound 8 (54 mg, 0.29 mmol) were dissolved in DMF (1 mL). BOP (194 mg, 0.44 mmol) and DIPEA (113 mg, 0.88 mmol) were added to the reaction system. The reaction was carried out at room temperature for 12 hours. After the reaction was complete, the mixture was diluted with water and extracted with ethyl acetate (10 mL × 3). The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous Na2SO4, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain S11 (109 mg, 46%).

[0134] S11: 1 H NMR(400MHz,DMSO-d6)δ12.60(s,1H),8.29-8.24(m,1H),8.16(s,1H),8.03-7.96(m,1H),7.93-7.81(m,2H),7.49-7.22(m, 9H),6.92-6.84(m,2H),4.74(s,4H),4.34(s,2H),3.76(s,2H),3.29(s,2H),3.11(s,2H),2.95(s,2H).MS(ESI,m / z):603(M + +1).

[0135] Example 12: Synthesis of compound S12

[0136] Step 1: Synthesis of intermediate 21

[0137]

[0138] a) Intermediate 19 (476 mg, 1.72 mmol), N,N'-disuccinimidyl carbonate (530 mg, 2.07 mmol), and pyridine (136 mg, 1.72 mmol) were dissolved together in anhydrous acetonitrile (8.6 mL), and a solution of 2,3-dihydro-1H-pyrrolo[3,4-C]pyridine was added dropwise at 0 °C. The 2,3-dihydro-1H-pyrrolo[3,4-C]pyridine solution was prepared by dissolving 2,3-dihydro-1H-pyrrolo[3,4-C]pyridine (270 mg, 1.72 mmol) and DIPEA (446 mg, 3.45 mmol) together in anhydrous acetonitrile / DMF (0.95 mL / 0.48 mL). The two solutions were mixed and reacted at room temperature for 12 hours. After complete reaction, the mixture was diluted with water, extracted with ethyl acetate (15 mL × 3), and the organic phases were combined. The mixture was washed twice with saturated sodium chloride solution, dried over anhydrous Na₂SO₄, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to give intermediate 20 (510 mg, 55%). MS (ESI, m / z): 423 (M + +1).

[0139] b) Intermediate 20 (510 mg, 1.2 mmol) was dissolved in DCM (2 mL), and TFA (1 mL) was added to the reaction system. The reaction was carried out at room temperature for 1 hour. After the reaction was complete, dichloromethane was removed by vacuum distillation. The solution was diluted with water, and the pH was adjusted to 8-10 at 0 °C with 2 M NaOH aqueous solution. The solution was extracted with dichloromethane, and the organic phases were combined, dried over anhydrous Na2SO4, filtered to remove salts, and concentrated under reduced pressure to obtain intermediate 21 (350 mg, 90%). MS (ESI, m / z): 323 (M + +1).

[0140] Step 2: Synthesis of compound S12

[0141]

[0142] a) Intermediate 21 (120 mg, 0.37 mmol) and compound 8 (90 mg, 0.30 mmol) were dissolved in DMF (1 mL). BOP (200 mg, 0.45 mmol) and DIPEA (232 mg, 1.80 mmol) were added to the reaction system. The reaction was carried out at room temperature for 12 hours. After the reaction was complete, the mixture was diluted with water and extracted with ethyl acetate (20 mL × 3). The organic phases were combined, washed twice with saturated sodium chloride solution, dried over anhydrous Na2SO4, filtered to remove salt, concentrated under reduced pressure, and purified by column chromatography to obtain S12 (108 mg, 48%).

[0143] S12: 1 H NMR (300MHz, DMSO-d6) δ12.62(s,1H),8.61(s,1H),8.50(d,J=5.0Hz,1H),8.39(s,1H),8.26(dd,J=7.6,1.6 Hz,1H),7.98(d,J=7.8Hz,1H),7.93-7.77(m,2H),7.49(d,J=8.2Hz,2H),7.42(dd,J=8.9,5.1Hz,3H),7.24(t ,J=9.0Hz,1H),7.13(d,J=8.5Hz,2H),4.80(d,J=5.4Hz,4H),4.62(d,J=12.8Hz,1H),4.35(s,2H),3.43(s,1H ),3.12(t,J=12.6Hz,1H),2.90-2.67(m,2H),1.86(d,J=12.8Hz,1H),1.73-1.36(m,3H).MS(ESI,m / z):603(M + +1).

[0144] Example 13: Synthesis of compound S13

[0145]

[0146] The synthesis method is the same as in Example 12. Compound S13 can be obtained simply by changing the corresponding raw materials.

[0147] S13: 1 H NMR (400MHz, DMSO-d6) δ12.61(s,1H),8.64(d,J=2.6Hz,1H),8.54(s,1H),8.26(d, J=7.8Hz,1H),7.99(d,J=7.9Hz,1H),7.96-7.78(m,3H),7.45-7.29(m,6H),7.28-7. 15(m,2H),4.78(s,4H),4.60(d,J=12.9Hz,1H),4.35(s,2H),3.20-3.09(m,2H),2.8 9(d,J=12.1Hz,2H),1.93(d,J=13.0Hz,1H),1.80-1.44(m,3H).MS(ESI,m / z):603(M + +1).

[0148] Example 14: Synthesis of compound S14

[0149]

[0150] The synthesis method is the same as in Example 12. Compound S14 can be obtained simply by changing the corresponding raw materials.

[0151] S14: 1 H NMR (400MHz, DMSO-d6) δ12.61(s,1H),8.96(s,1H),8.25(d,J=7.8Hz,1H),8.15(d,J=2.3Hz,1H),7.97 (d,J=7.9Hz,1H),7.91-7.76(m,3H),7.65(d,J=8.8Hz,1H),7.46-7.39(m,2H),7.39-7.27(m,4H),7.23 (t,J=9.0Hz,1H),4.80(s,4H),4.63(d,J=12.9Hz,1H),4.34(s,2H),3.42(s,1H),3.13(t,J=12.7Hz,1H ),2.82(q,J=12.2Hz,2H),1.96-1.75(m,1H),1.58(d,J=12.4Hz,2H),1.45(s,1H).MS(ESI,m / z):603(M + +1).

[0152] Example 15: Synthesis of compound S15

[0153]

[0154] The synthesis method is the same as in Example 11. Compound S15 can be obtained simply by changing the corresponding raw materials.

[0155] S15: 1 H NMR(400MHz,DMSO-d6)δ12.61(s,1H),8.29-8.26(m,1H),8.15(s,1H),8.03-7.97(m,1H),7.93-7.83(m,2H),7.46-7.23(m,9H),6 .93-6.86(m,2H),4.73(s,4H),4.34(s,2H),3.72(s,2H),3.43(s,1H),3.29(s,2H),3.10(s,2H),2.96(s,2H).MS(ESI,m / z):602(M + +1).

[0156] Example 16: Evaluation of PARP and NAMPT enzyme inhibitory activities

[0157] 1. Experimental Methods

[0158] PARP1 inhibitory activity was assessed using a homogeneous PARP assay kit. The assay consisted of 1 ng / μL PARP-1, 20 ng / μL activated DNA (Sigma), and 500 nM NAD dissolved in the following reaction buffer: 100 mM Tris-HCl (pH 8.0), 100 mM NaCl, and 20 mM MgCl2. At room temperature, PARP-1 protein (1 ng / μL) was pre-incubated with different concentrations of the test compound for 30 min, followed by the addition of 500 nM NAD to initiate the parylation reaction. Remaining unreacted NAD was incubated with a circulating assay solution (1% ethanol, 25 μM resazurin, 0.30 U / mL alcohol dehydrogenase, and 0.25 U / mL myocardial flavin). The NAD concentration was proportional to the fluorescence signal at Ex 540 nm / Em 590 nm. The IC50 of the compounds was assessed by evaluating the rate of NAD reduction under different concentrations of the compounds. 50 The value.

[0159] The NAMPT inhibitory activity assay was performed according to the operating procedures provided by CycLex NAMPT colorimetric assay kit (MBL International Corp.). The basic principle is as follows: NAM and PRPP react with NAMPT to generate NMN, while NMN and ATP react with NMNAT1 to generate NAD. NAD then reacts with alcohol dehydrogenase (ADH) to generate NADH, which is subsequently converted back to NAD by myocardial flavin enzyme. WST-1 forms an orange-yellow formazan in the NAD / NADH enzymatic cycle. The effect of the compound on NAMPT enzyme activity can be detected by measuring the change in absorbance at OD 450 nM.

[0160] The specific experimental procedures are as follows:

[0161] (1) Preparation of test compound solutions: All compounds were dissolved in DMSO with a stock solution concentration of 10 mM. The solutions were prepared to the required concentrations according to the test requirements. Starting from the initial concentration, the solutions were diluted 2 times in a gradient, and 8 concentration gradients were set. Each concentration was repeated three times.

[0162] (2) Add ddH2O (10 μL) and NAM (5 μL) to the 96-well detection plate;

[0163] (3) Add 5 μL of the test compound or DMSO to the 96-well detection plate;

[0164] (4) Prepare mixture I, comprising: 20×NAMPT test buffer (5 μL), PRPP (5 μL), ATP (5 μL), recombinant NMNAT1 (5 μL), ddH2O (35 μL), and NAMPT (5 μL), totaling 60 μL. Add mixture I to the 96-well detection plate;

[0165] (5) Place the 96-well plate in a 30°C incubator and incubate for 60 min;

[0166] (6) Prepare mixture I, including: WST-1 (5 μL), ADH (5 μL), myocardial flavin (5 μL), ethanol (5 μL), totaling 20 μL. After incubation, remove the 96-well plate and add mixture II to the 96-well plate;

[0167] (7) Monitor the absorbance of each well at OD 450nm dynamically within 30 min using an ELISA reader, and perform the test every 5 min.

[0168] (8) Select a time period in which absorbance and time change linearly, and calculate the reaction rate. Process using GraphPad Prism5 software. To compensate for the NAMPT inhibitory activity of DMSO, the values ​​were corrected using DMSO as a solvent control.

[0169] 2. Experimental Results

[0170] Table 1 Inhibitory activities of NAMPT and PARP1 (IC50) 50 (nM)

[0171]

[0172] As shown in Table 1, the compounds designed in this invention exhibit good inhibitory activity against NAMPT and PARP1 in cells, with an enzyme inhibition IC50 value of [missing value]. 50 The value reached the nanomolar concentration level.

[0173] Example 17: Evaluation of antitumor activity

[0174] 1. Experimental Methods

[0175] (1) Collect cells in the logarithmic growth phase, adjust the cell suspension concentration, and add 100 μL of cell suspension to each well of a 96-well plate; the number of cells per well is about 7000, and incubate overnight at 37°C with 5% CO2 until the cells are completely attached.

[0176] (2) Set up a concentration gradient of the compound of the present invention, with 3 replicates for each concentration gradient. Dilute the compound of the present invention to the corresponding culture medium to the required final concentration. Remove the original culture medium from the 96-well plate and add 100 μL of the prepared culture medium containing the required final concentration of the drug. Incubate at 37°C with 5% CO2. At the same time, set up a blank group (containing only 100 μL of culture medium, without cells, and subsequent treatment is the same as other wells) and a control group (containing cells and culture medium).

[0177] (3) Add 10 μL MTT solution (5 mg / ml) to each well after 68 hours of drug treatment and continue culturing for 4 hours (total drug treatment of cells for 72 hours);

[0178] (4) Remove the culture medium from the wells (if the cells are suspended, centrifuge at 2500 rpm for 5 min before removing the culture medium). Add 150 μL of dimethyl sulfoxide to each well and shake until the crystals are fully dissolved. Measure the absorbance at OD490 nm in each well using a microplate reader;

[0179] (5) Calculate the inhibition rate: Inhibition rate = 1 - (OD value of the drug-treated group - OD value of the blank group) / (OD value of the control group - OD value of the blank group) = (OD value of the control group - OD value of the drug-treated group) / (OD value of the control group - OD value of the blank group);

[0180] (6) IC was calculated using GraphPad Prism fitting. 50 value.

[0181] 2. Experimental Results

[0182] Table 2 Anti-cell proliferation activity (IC50) 50 (μM)

[0183] S1 0.11 0.23 0.50 0.42 1.82 ＞50 S2 0.22 0.18 0.29 0.53 0.99 ＞50 S3 0.10 0.12 0.25 0.19 1.07 ＞50 S4 0.33 0.21 0.54 0.88 2.33 ＞50 S5 0.22 0.18 0.21 0.18 0.65 ＞50 S7 0.18 0.11 0.32 0.039 0.056 ＞50 S8 0.32 0.22 0.22 0.45 0.46 ＞50 S12 0.043 0.033 0.034 0.012 0.028 ＞50 FK866 19.2 20.3 40.3 9.8 34.5 ＞50 Olapani 0.32 0.12 3.89 10.21 7.95 ＞50

[0184] As shown in Table 2, the compounds designed in this invention have good anti-proliferative activity against BRAC-mutant triple-negative breast cancer cells (MDA-MB-436 and HCC1937) and BRAC wild-type triple-negative breast cancer cells (MDA-MB-468, MCF-7, and MDA-MB-231); and have no cytotoxicity to normal cells MCF-10A, indicating that the compounds designed in this invention have good safety.

Claims

1. A phthalazinone compound as shown in Formula I, or a pharmaceutically acceptable salt thereof, ， In Formula I: X is N or C; L1 is a single bond or ; Y is C 6-10 Aryl or 5- to 10-membered heteroaryl containing 1 to 3 N, O, or S heteroatoms; L2 is -(CH2) n -, where n is 0, 1, 2, or 3; R1 is a 9- to 10-membered bicyclic heteroaryl group containing one N atom and one to two N, O, and S heteroatoms.

2. The phthalazinone compound or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, In Formula I: Y is phenyl or pyridyl; n is 0, 1, or 2.

3. The phthalazinone compound or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, In Formula I: Y represents phenyl, or ; n is 0 or 1.

4. The phthalazinone compound or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, In Formula I: R1 is imidazopyridyl, imidazopyrimidyl, pyrazolopyridyl, thienopyridyl, furanopyridyl, tetrahydropyrrolopyridyl, or tetrahydropyrrolophenyl.

5. The phthalazinone compound or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, In Formula I: R1 is .

6. The phthalazinone compound or a pharmaceutically acceptable salt thereof according to claim 1, characterized in that, Selected from any one of the following compounds: 。 7. A method for preparing a phthalazinone compound of claim 1 or a pharmaceutically acceptable salt thereof, characterized in that, Includes the following steps: Compound II and compound III undergo a condensation reaction in the presence of a base and a condensing agent to yield compound I: ， Wherein, X, Y, L1, L2, and R1 are as described in claim 1.

8. A pharmaceutical composition, characterized in that, The compound contains a therapeutically effective amount of the phthalazinone compound of claim 1 or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

9. The use of a phthalazinone compound of claim 1 or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition of claim 8, in the preparation of a medicament for treating triple-negative breast cancer.

10. The application according to claim 9, characterized in that, The drug treats triple-negative breast cancer by inhibiting poly(ADP-ribose) polymerase and / or nicotinamide phosphoribosyltransferase.

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