A compound binding to crbn protein and degrader of the same

Abstract

The application belongs to the technical field of pharmaceutical chemistry, and particularly relates to a compound capable of combining with CRBN protein and a composition thereof, and further relates to a protein degradation agent based on CRBN protein and application thereof. The application provides a series of novel compounds capable of combining with CRBN protein, and a protein degradation agent based on the compounds. The CRBN small-molecule ligand of the application has high-efficiency CRBN combining activity, and can be used for E3 ubiquitin ligase-mediated protein degradation; the PROTAC degradation agent exhibits excellent targeted protein degradation activity, and is suitable for various cancer treatments. The compound of the application breaks through the limitation of traditional small-molecule inhibitors, and can promote personalized treatment and expand the application of precision medicine.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical chemistry technology, specifically relating to a compound and its composition that can bind to CRBN protein, and also to a protein degrading agent based on CRBN protein and its application. Background Technology

[0002] CRBN (Cereblon) is a brain-related protein with ionic protease activity. Its gene is located on the short arm of chromosome 3, region 26, and its molecular weight is 51 kDa. Initial studies showed that CRBN was associated with human autosomal recessive non-integrative mental developmental delay (ARNSMR). It was named based on its role in brain development and the presence of an ATP-dependent tyrosine kinase region (Jo S, Lee KH, Song S, et al. Identification and functional characterization of cereblon as a binding protein for large-conductance calcium-activated potassium channel in rat brain. J Neurochem, 2005; 94(5):1212-1224.). CRBN is the substrate receptor protein of the CRL4 CRBN isoform of the E3 ubiquitin ligase complex. It, along with DDB1 (Damage-specific DNA Binding protein 1), zinc finger domain protein RBX1, and Cullin4 (Cul4) scaffold protein, forms the DDB1–Cul4–Rbx1–CRBN E3 ubiquitin ligase complex, which determines the substrate specificity of the CRL4 E3 ubiquitin ligase. It is widely expressed in the cytoplasm, nucleus, and peripheral membranes of the prostate, liver, pancreas, placenta, kidney, lung, peripheral blood leukocytes, and brain. E3 ubiquitin ligases specifically recognize substrate proteins and induce polyubiquitination of these proteins. This leads to the degradation of ubiquitinated substrate proteins via the ubiquitin-proteasome pathway, thereby affecting various physiological activities such as cellular energy metabolism, membrane potential regulation, and transcription factor degradation.

[0003] In 2010, Ito et al. first confirmed that the direct target of thalidomide is a substrate receiver protein of E3 ubiquitin ligase—CRBN (Ito T, Ando H, Suzuki T, et al. Identification of a primary target of thalidomide teratogenicity[y]. Science, 2010, 327(5971):1345-50.). After thalidomide and other immunomodulators bind to CRBN, they activate the activity of E3 ubiquitin ligase, which recruits the substrate protein IKZF1 / IKZF3. Subsequently, the E3 ubiquitin ligase, the substrate protein IKZF1 / IKZF3, and the immunomodulator form a stable ternary complex, thereby transferring ubiquitin to the substrate protein IKZF1 / IKZF3, leading to the degradation of IKZF1 / IKZF3 by the 26S proteasome. IKZF1 and IKZF3 are essential transcriptional regulators for the proliferation and development of B cells and T cells [John LB, Ward AC. The ikaros gene family: transcriptional regulators of hematopoiesis and immunity[J]. Molecular immunology, 2011, 48(9):1272-1278. and Dijon M, Bardin F, Murati A, et al. The role of ikaros in human erythroid differentiation[J]. Blood, 2008, 111(3):1138-1146]. Under normal circumstances, IKZF3 inhibits the gene encoding IL-2 in T cells and stimulates the expression of IRF4. Therefore, the degradation of IKZF1 and IKZF3 directly reduces the expression of transcription factors such as IRF4 and Myc, thereby exerting cytotoxic effects on myeloma cells. On the other hand, the degradation of IKZF1 and IKZF3 increases the expression of IL-2 in T cells, thereby activating the immune response of T cells and inhibiting the function of B cells, achieving tumor killing and tumor proliferation inhibition. At the same time, it reduces the expression of tumor necrosis factor TNF and promotes the secretion of the anti-inflammatory factor IL-10 by human peripheral blood mononuclear cells. This imbalance leads to apoptosis of multiple myeloma cells. Based on this, in recent years, protein degradation-targeting chimeras (PROTACs) designed using the chemical structures of amine immunomodulators such as thalidomide, lenalidomide, and pomalidomide have emerged in large numbers.

[0004] Traditional small molecule drugs bind to target proteins reversibly or irreversibly as a means of regulating specific biological activities. In contrast, PROTACs bind to their target proteins, subsequently leading to the degradation of those proteins. Theoretically, once this effect is achieved, PROTACs can repeat this process with another target protein. Unlike traditional small molecule inhibitors, PROTACs exhibit better degradation capabilities (DC50 and Dmax) than those achieved solely through binding affinity. PROTAC molecules typically consist of three parts: (1) a ligand portion that binds to the target protein; (2) a ligand portion that binds to the E3 ubiquitin ligase; and (3) a linker that connects (1) and (2). PROTAC technology is characterized by high efficiency and high selectivity. In application, PROTAC molecules simultaneously bind to the target protein and the E3 ubiquitin ligase, forming a ternary complex that ubiquitinates the target protein, which is then degraded via the proteasome. This PROTAC-mediated target protein degradation has the potential to treat certain diseases, such as cancer, inflammation, and autoimmune diseases. Regardless of which target protein binding fragment (1) is used in the PROTAC target protein ligand portion, the essential unit that must be present at the other end of the PROTAC molecule is the E3 ubiquitin ligase ligand (2) to label the target protein for degradation. Several effective E3 ubiquitin ligase ligand structures have been discovered and applied to PROTAC development. However, metabolic instability and off-target toxicity are common problems. Therefore, developing metabolically stable and selective E3 ubiquitin ligase ligands has significant application value.

[0005] Currently, over 100 proteins have been successfully degraded by PROTAC. These targets include (1) kinases, such as IRAK4, RIPK2, BCR-ABL, EGFR, HER2, c-Met, TBK1, CDK2 / 4 / 6 / 9, ALK, Akt, CK2, ERK1 / 2, FLT3, PI3K, BTK, Fak, etc.; (2) BET proteins, such as BRD2 / 4 / 6 / 9; (3) nuclear receptors, such as AR, ER, etc.; and (4) other proteins, such as MetAp-2, Bcl-xL, Sirt2, HDAC6, Pirin, SMAD3, ARNT, PCAF / GCN5, Tau, FRS2, etc. Even transcription factor regulatory proteins such as Pirin, epigenetic proteins such as PCAF / GCN5, and KRAS-G12C, which are considered "undruggable targets," are included. Therefore, PROTAC technology may become a potential method for addressing and treating diseases related to the above target proteins. Summary of the Invention

[0006] This invention prepares a series of compounds that can bind to CRBN proteins. These compounds can be used as structural units for E3 ubiquitin ligase ligands in PROTAC technology. PROTACs designed based on this series of compounds can use specific inhibitors of target proteins requiring degradation or other small molecule compounds as target protein ligands to achieve the degradation of target proteins dependent on CRBN-type E3 ubiquitin ligases, thus providing more therapeutic options for target protein-related diseases. Based on this, this invention has been completed.

[0007] In a first aspect, the present invention provides a compound, or a stereoisomer thereof, or a pharmaceutically acceptable salt thereof, or a deuterated compound thereof, or a tautomer thereof, or a polymorph thereof, or a solvate thereof, or an N-oxide thereof, or an isotopically labeled compound thereof, or a metabolite thereof, or a prodrug thereof, capable of binding to CRBN protein; wherein the compound comprises a fused ring skeleton consisting of two ring systems, A and B, and the G group is connected to any one of the 4, 5, or 6 positions of the A ring via a carbon-carbon single bond, and the structure of the compound capable of binding to CRBN protein is shown in Formula (I);

[0008]

[0009] In Equation I, the G substructure includes the following G 1 G 2 G 3 Three types, namely R configuration, S configuration, and racemic configuration:

[0010]

[0011] In Formula I, ring A is a benzene ring, selected from any of the following:

[0012]

[0013] R1 is selected from any one of H, C1-C3 alkyl, C1-C3 alkoxy or halogen;

[0014] R2 is selected from any one of H, C1-C3 alkyl, C1-C3 alkoxy or halogen;

[0015] R3 is selected from any one of H, C1-C3 alkyl, C1-C3 alkoxy or halogen;

[0016] R4 is selected from any one of H, C1-C3 alkyl, C1-C3 alkoxy or halogen;

[0017] In Formula I, ring B... Representing a covalent single bond or a covalent double bond, X1, X2 and X3 of the B ring are selected from either C or N;

[0018] The specific structure of the B ring is preferably selected from any of the following structures:

[0019]

[0020] Wherein, P1, P2, and P3 are selected from H, halogens, or any of the following structures:

[0021]

[0022] Furthermore, in Formula I, the halogens of the substituents R1, R2, R3, and R4 are selected from any one of F, Cl, and Br.

[0023] Furthermore, the pharmaceutically acceptable salt refers to the parent compound prepared by addition of a non-toxic acid or its base.

[0024] In some embodiments of the present invention, the specific structure of the compound capable of binding to the CRBN protein is selected from any of the following:

[0025]

[0026]

[0027] Secondly, the present invention provides a PROTAC degrading agent, the structure of which is as follows:

[0028] Wherein, the CRBN protein ligand is the compound described in the first aspect of the present invention; the structure of the PROTAC degrading agent is shown in formula (II):

[0029]

[0030] In Formula II, the occupying group The structure is none, or it can be selected from any of the following structures:

[0031]

[0032] When the occupying group When the structure is empty, the Linker's L 1 The CRBN protein ligand's B structure is linked to any one of the X1, X2, or X3 sites by a CN, NC, or CO single bond;

[0033] When the occupying group When selected from the above structures, the occupying group With the Linker L 1 Substructures are linked by CN, NC, or CO single bonds;

[0034] In Formula II, the substructure L of the linker 1 L 3 and L 5 Each of the following structures can be selected:

[0035]

[0036] Among them, R a Selected from any one of H, alkyl, SO2-alkyl, or SO2-aryl;

[0037] In Formula II, the substructure L of the linker 2 and L 4 Each of the following structures can be selected:

[0038]

[0039] In Formula II, the target protein ligand is a ligand of a target protein that causes the related disease, and the target protein ligand is selected from any one of the following structures:

[0040]

[0041]

[0042]

[0043]

[0044]

[0045]

[0046] Furthermore, the diseases mentioned include, but are not limited to, prostate cancer, breast cancer, non-small cell lung cancer, chronic myeloid leukemia, acute myeloid leukemia, T-cell acute lymphoblastic leukemia, Alzheimer's disease, gout, autoimmune diseases, inflammatory bowel disease, B-cell lymphoma, androgenetic alopecia, acne, synovial sarcoma, solid tumors, lung cancer, multiple myeloma, lymphoma, tumor metastasis, cervical cancer, neuroblastoma, hepatocellular carcinoma, colorectal cancer, pancreatic cancer, malignant rhabdomyosarcoma, and oral squamous cell carcinoma.

[0047] Furthermore, the PROTAC degrading agent targets AR, ER, BTK, and IRAK4.

[0048] In some embodiments of the present invention, the specific structure of the PROTAC degrading agent is selected from any of the following:

[0049]

[0050]

[0051] Thirdly, the present invention provides a pharmaceutical composition comprising the compound shown in the first aspect formula (I).

[0052] Furthermore, the pharmaceutical composition may also comprise an enantiomer, diastereomer, stereoisomer, or pharmaceutically acceptable salt of the compound represented by formula (I) and a pharmaceutically acceptable diluent or carrier.

[0053] Furthermore, the pharmaceutically acceptable salt refers to the parent compound prepared by addition of a non-toxic acid or its base.

[0054] Furthermore, the pharmaceutical composition can be formulated into various dosage forms, including but not limited to one or more of tablets, capsules, aerosols, pills, powders, solutions, suspensions, emulsions, granules, liposomes, transdermal preparations, and / or suppositories.

[0055] Fourthly, the present invention provides a pharmaceutical composition for targeting and degrading proteins, said pharmaceutical composition comprising a PROTAC degrading agent represented by formula (II).

[0056] Furthermore, the pharmaceutical composition targets AR, ER, BTK, and IRAK4.

[0057] Furthermore, the pharmaceutical composition may also comprise an enantiomer, diastereomer, stereoisomer, or pharmaceutically acceptable salt of the PROTAC degrader shown in formula (II) and a pharmaceutically acceptable diluent or carrier.

[0058] Furthermore, the pharmaceutically acceptable salt refers to the parent compound prepared by addition of a non-toxic acid or its base.

[0059] Furthermore, the pharmaceutical composition can be formulated into various dosage forms, including but not limited to one or more of tablets, capsules, aerosols, pills, powders, solutions, suspensions, emulsions, granules, liposomes, transdermal preparations, and / or suppositories.

[0060] Fifthly, the present invention provides the use of the compound described in the first aspect or the PROTAC degrading agent described in the second aspect, or its stereoisomers, pharmaceutically acceptable salts, deuterated compounds, tautomers, polymorphs, solvates, N-oxides, isotope-labeled compounds, metabolites, or prodrugs in the preparation of medicaments for treating diseases related to proteins such as AR, ER, BTK, and IRAK4.

[0061] Furthermore, the drug achieves its therapeutic effect or alleviates symptoms by limiting or inhibiting the expression of AR, ER, BTK, and IRAK4.

[0062] Furthermore, the diseases related to the AR, ER, BTK, IRAK4, and other proteins include inflammatory diseases and tumors.

[0063] Furthermore, the diseases related to the proteins AR, ER, BTK, and IRAK4 include, but are not limited to, prostate cancer, androgenetic alopecia, polycystic ovary syndrome, breast cancer, endometrial cancer, osteoporosis, B-cell lymphoma, lymphocytic leukemia, autoimmune diseases, monocytic leukemia, acute myeloid leukemia, multiple myeloma, non-small cell lung cancer, chronic myeloid leukemia, gout, inflammatory bowel disease, acne, synovial sarcoma, solid tumors, small cell lung cancer, cervical cancer, neuroblastoma, hepatocellular carcinoma, colorectal cancer, pancreatic cancer, malignant rhabdomyosarcoma, and oral squamous cell carcinoma.

[0064] Furthermore, the drug can be formulated into various dosage forms, including but not limited to one or more of tablets, capsules, aerosols, pills, powders, solutions, suspensions, emulsions, granules, liposomes, transdermal preparations, and / or suppositories.

[0065] Beneficial effects

[0066] This invention provides a series of novel compounds capable of binding to CRBN proteins, and protein degrading agents (PROTACs) based on these compounds, which mainly have the following advantages and beneficial effects:

[0067] 1. It possesses highly efficient CRBN binding activity and can be used for E3 ubiquitin ligase-mediated protein degradation.

[0068] The CRBN small molecule ligand structure of the present invention is novel and has better CRBN binding affinity than lenalidomide, which can mediate the degradation of target proteins more efficiently.

[0069] These compounds can serve as E3 ubiquitin ligands for the design of novel PROTACs and molecular glue degraders, expanding the application scope of targeted degradation strategies.

[0070] 2. Exhibits excellent targeted protein degradation activity, suitable for the treatment of various cancers.

[0071] In in vitro experiments, the compounds of this invention exhibited significant degradation activity against a variety of cancer-related proteins (AR, ER, BTK, IRAK4); among which:

[0072] The PROTAC degrading agent of this invention exhibits significant degradation activity against target proteins AR, BTK, and IRAK4, and can induce target protein degradation even at low concentrations, demonstrating high degradation efficiency.

[0073] Meanwhile, the D of the PROTAC degrading agent of this invention max All of them were at Grade A, indicating that these compounds can reduce the expression level of target proteins to the greatest extent, with a degradation rate of over 75%.

[0074] 3. Applicable to a variety of cancer targets that are difficult to treat, overcoming the limitations of traditional small molecule inhibitors.

[0075] Proteins that traditional small molecule inhibitors cannot directly target (such as transcription factors, RNA-binding proteins, and cell cycle regulatory proteins) can be degraded using the PROTAC technology of this invention.

[0076] 4. Promote personalized treatment and expand the application of precision medicine.

[0077] Since the small molecule CRBN ligand of the present invention can be used in the design of different PROTACs, researchers can use precise drugs according to the gene mutations and target protein expression of different cancer patients.

[0078] Combining biomarker detection can further optimize drug screening strategies and enable personalized protein degradation therapies. Detailed Implementation

[0079] The specific embodiments of the present invention will be further described below. It should be noted that these descriptions are for the purpose of aiding understanding the present invention, but do not constitute a limitation thereof. Furthermore, the technical features involved in the embodiments described below can be combined with each other as long as they do not conflict with each other.

[0080] The term “stereoisomer” as used in this article refers to compounds that have the same chemical composition but different spatial arrangements of atoms or groups, including “diastereomers” and “enantiomers”.

[0081] The term "diastereomer" as used herein refers to a stereoisomer having two or more chiral centers and whose molecules are not mirror images of each other. Diastereomers possess different physical properties, such as melting point, boiling point, spectral characteristics, and reactivity. In the presence of resolving agents or chromatography, mixtures of diastereomers can be separated using high-resolution analytical steps such as electrophoresis and crystallization, in the presence of chiral HPLC columns.

[0082] The term "enantiomer" as used herein refers to two stereoisomers of a compound that do not overlap as mirror images of each other. A 50:50 mixture of enantiomers is called a racemic mixture or racemate, which can occur during chemical reactions or processes where stereoselectivity or stereoorientation has ceased.

[0083] The terms “pharmaceutically acceptable salt” and “salt of a compound” used herein are interchangeable and both refer to derivatives of the disclosed compound, wherein the parent compound is prepared by the addition of a non-toxic acid or its base to form a salt. Examples of pharmaceutically acceptable salts include, but are not limited to, non-toxic acid addition salts: salts formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, and perchloric acid, or with organic acids such as acetic acid, maleic acid, tartaric acid, citric acid, succinic acid, or malonic acid. Other pharmaceutically acceptable salts include, but are not limited to, adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, hydrogen sulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentylpropionate, diglucuronate, dodecyl sulfate, ethanesulfonate, formate, fumarate, glucohepanoate, glyceryl phosphate, glucuronate, hemisulfate, heptaate, hexanoate, hydroiodate, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, dihydroxynaphthalate, pectate, persulfate, 3-phenylpropionate, phosphate, picrate, neopentanoate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate, etc. Representative alkali metal or alkaline earth metal salts include sodium, lithium, potassium, calcium, and magnesium. Where appropriate, other pharmaceutically acceptable salts include non-toxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, alkyl groups having 1 to 6 carbon atoms, sulfonate, and arylsulfonate.

[0084] Table 1. Exemplary compound structures binding to CRBN protein

[0085]

[0086]

[0087]

[0088]

[0089]

[0090]

[0091]

[0092]

[0093]

[0094] Table 2. Exemplary compound structures of PROTAC protein degraders

[0095]

[0096]

[0097]

[0098]

[0099]

[0100]

[0101]

[0102]

[0103]

[0104]

[0105]

[0106]

[0107]

[0108]

[0109] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, and the experimental materials used in the following examples are all commercially available. In the following examples, A3, B3, C1, C2, C4, C7, E1, E2, E4, E5, E6, E9, F1, F2, F4, G1, G2, G12, H1, H2, H8, H14, H22, I1, K1, N1, and N2 are commercially available compounds.

[0110] Example 1: Synthesis of Compound A5

[0111] Route A:

[0112]

[0113] In a flask dried in an oven and purged with argon, bromoindole starting material A1 (1.0 equiv) was added, followed by anhydrous THF (0.1 M). After argon purging and an ice-water bath, NaH (1.2 equiv) was added, and the mixture was stirred for 30 min. Iodomethane (1.1 equiv) was then added, and the reaction was allowed to proceed for 6 h. TLC monitoring showed no remaining bromoindole starting material. The reaction was quenched with water, extracted with EtOAc, and the combined organic phases were washed with saturated brine and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate A2 was obtained.

[0114] In an oven-dried and argon-purged flask, bromoindole starting material A2 (1.0 equiv.), 2,6-bis(benzyloxy)-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine (A1, 1.2 equiv.), and Pd(dppf)Cl2 (10 mol%) were added. After argon purging, a mixture of dried 1,4-dioxane-pre-deoxygenated saturated sodium bicarbonate aqueous solution (3:1, v:v, 0.1 M) was added, followed by argon purging. The reaction mixture was heated overnight in an oil bath at 85°C. TLC monitoring showed no remaining bromoindole starting material. After the reaction system cooled to room temperature, water and EtOAc were added for dilution. The resulting aqueous phase was extracted with EtOAc, and the combined organic phases were washed with saturated brine and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate A4 was obtained.

[0115] In a flask that has been dried in an oven and purged with argon, intermediate A4 was added, followed by MeOH-THF (1:1, v:v, 0.2M) and 10% Pd / C (20wt%). Hydrogen was introduced during vigorous stirring, and the reaction mixture was stirred vigorously under hydrogen balloon pressure for 8 hours. TLC monitoring showed no remaining starting material, and LC-MS monitoring showed no unreacted intermediates. The reaction mixture was filtered through silica gel cake and evaporated under reduced pressure to obtain crude product A5. A4, the target final product, requires preparative-grade HPLC purification (MeCN / H2O) to obtain a pure product. Intermediate A5, however, requires no purification and can be used directly in the next reaction step.

[0116] Example 2: Synthesis of compounds B4 and B5

[0117] Route B:

[0118]

[0119] In a flask that has been dried in an oven and purged with argon, the following starting materials were added sequentially: B1 (1.0 equiv.), NaI (1.0 equiv.), CuI (5.0 equiv.), dioxane (0.1 M), and N.1 N 1 The reaction system was subjected to dimethylethane-1,2-diamine (0.1 equiv) and stirred in an oil bath at 100 °C for 24 h. LC-MS analysis showed no residual starting material. The mixture was extracted with DCM / water, and the combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by preparative HPLC (MeCN / H2O) to obtain B2.

[0120] In an oven-dried flask purged with argon, anhydrous tetrahydrofuran (THF) and anhydrous dimethylformamide (DMA) were thoroughly mixed to prepare a 30% DMA / THF solution. Next, a stock solution of nickel and ligands was prepared: in a small vial, NiBr2·diglyme (0.1 equiv) and (R)-4-heptylBiOX ((R)-L1, 0.11 equiv) or ((S)-4-heptylBiOX ((S)-L1, 0.11 equiv)) solids were added, along with 1.5 mL of the 30% DMA / THF solution via syringe. The flask was heated to 60°C and stirred until a homogeneous purple solution formed. The flask was then removed from the heat source and cooled to room temperature with stirring, resulting in an olive-green solution. In another reaction flask, manganese powder (1.5 equiv), 3-chloropiperidine-2,6-dione (1.0 equiv), and B2 (1.5 equiv) solids were added. Ni / L1 stock solution was added via syringe. Then, trimethylsilane chloride (TMSCl, 0.8 equiv) was added to the reaction flask. The entire process was carried out under argon protection and stirred at 1200 rpm at room temperature for 2 hours. LC-MS analysis showed no reactant residue. The mixture was extracted with DCM / water, and the combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration and solvent removal under reduced pressure, preparative-grade HPLC (MeCN / H2O) was used to purify B2. B4 or B5 was obtained by DCM / water extraction, washing with saturated sodium chloride solution, and drying over anhydrous sodium sulfate. After filtration and solvent removal under reduced pressure, silica gel rapid column chromatography was used to purify B2.

[0121] Example 3: Synthesis of compounds 5-13, 18-26, 31-39

[0122]

[0123] Following the RouteA synthetic route, compound5 was synthesized from 4-Methyl-5-broMo-indole as the starting material.

[0124]

[0125] Following the Route A synthetic route, compound 6 was synthesized from 5-bromo-4-methoxy-1H-indole as the starting material.

[0126]

[0127] Following the synthetic route of Route A, compound 7 was synthesized from 5-Bromo-4-fluoro-1H-indole as the starting material.

[0128]

[0129] Following the Route A synthetic route, compound 8 was synthesized from 6-Methyl-5-broMo-indole as the starting material.

[0130]

[0131] Following the Route A synthetic route, compound 9 was synthesized from 5-bromo-6-methoxy-1H-indole as the starting material.

[0132]

[0133] Following the synthetic route of Route A, compound 10 was synthesized from 5-Bromo-6-fluoro-1H-indole as the starting material.

[0134]

[0135] Following the Route A synthetic route, compound 11 was synthesized from 7-Methyl-5-broMo-indole as the starting material.

[0136]

[0137] Following the synthetic route of Route A, compound 12 was synthesized from 5-bromo-7-methoxy-1H-indole as the starting material.

[0138]

[0139] Following the synthetic route of Route A, compound 13 was synthesized from 5-Bromo-7-fluoro-1H-indole as the starting material.

[0140]

[0141] Following the Route A synthetic route, compound18 was synthesized from 5-Methyl-4-broMo-indole as the starting material.

[0142]

[0143] Following the synthetic route of Route A, compound 19 was synthesized from 4-bromo-5-methoxy-1H-indole as the starting material.

[0144]

[0145] Following the synthetic route of Route A, compound 20 was synthesized from 4-Bromo-5-fluoro-1H-indole as the starting material.

[0146]

[0147] Following the RouteA synthetic route, compound21 was synthesized from 6-Methyl-4-broMo-indole as the starting material.

[0148]

[0149] Following the synthetic route of Route A, compound 22 was synthesized from 4-bromo-6-methoxy-1H-indole as the starting material.

[0150]

[0151] Following the synthetic route of Route A, compound 23 was synthesized from 4-Bromo-6-fluoro-1H-indole as the starting material.

[0152]

[0153] Following the RouteA synthetic route, compound24 was synthesized from 7-Methyl-4-broMo-indole as the starting material.

[0154]

[0155] Following the RouteA synthetic route, compound 25 was synthesized from 4-Bromo-7-methoxy-1H-indole as the starting material.

[0156]

[0157] Following the synthetic route of Route A, compound 26 was synthesized from 4-Bromo-7-fluoro-1H-indole as the starting material.

[0158]

[0159] Following the Route A synthetic route, compound 31 was synthesized from 4-Methyl-6-broMo-indole as the starting material.

[0160]

[0161] Following the RouteA synthetic route, compound 32 was synthesized from 6-Bromo-4-methoxy-1H-indole as the starting material.

[0162]

[0163] Following the synthetic route of Route A, compound 33 was synthesized from 6-Bromo-4-fluoro-1H-indole as the starting material.

[0164]

[0165] Following the RouteA synthetic route, compound34 was synthesized from 5-Methyl-6-broMo-indole as the starting material.

[0166]

[0167] Following the RouteA synthetic route, compound 35 was synthesized from 6-Bromo-5-methoxy-1H-indole as the starting material.

[0168]

[0169] Following the synthetic route of Route A, compound 36 was synthesized from 6-Bromo-5-fluoro-1H-indole as the starting material.

[0170]

[0171] Following the RouteA synthetic route, compound37 was synthesized from 7-Methyl-6-broMo-indole as the starting material.

[0172]

[0173]

[0174] Following the RouteA synthetic route, compound 38 was synthesized from 6-Bromo-7-methoxy-1H-indole as the starting material.

[0175]

[0176] Following the synthetic route of Route A, compound 39 was synthesized from 6-Bromo-7-fluoro-1H-indole as the starting material.

[0177] Example 4: Synthesis of compounds 1-4, 14-17, 27-30, and 40-43

[0178]

[0179] Following the Route B synthetic route, compound 1 was synthesized from 5-bromo-1H-indole as the starting material.

[0180]

[0181] Following the synthetic route of Route B, compound2 was synthesized from 5-bromo-1H-indole as the starting material.

[0182]

[0183] Following the synthetic route of Route B, compound 3 was synthesized from 5-Bromo-1-methyl-1H-indole as the starting material.

[0184]

[0185] Following the synthetic route of Route B, compound 4 was synthesized from 5-Bromo-1-methyl-1H-indole as the starting material.

[0186]

[0187] Following the Route B synthetic route, compound 14 was synthesized from 5-bromo-1H-indole as the starting material.

[0188]

[0189] Following the Route B synthetic route, compound15 was synthesized from 5-bromo-1H-indole as the starting material.

[0190]

[0191] Following the synthetic route of Route B, compound 16 was synthesized from 4-Bromo-1-methyl-1H-indole as the starting material.

[0192]

[0193] Following the synthetic route of Route B, compound 17 was synthesized from 4-Bromo-1-methyl-1H-indole as the starting material.

[0194]

[0195] Following the Route B synthetic route, compound27 was synthesized using 6-bromo-1H-indole as the starting material.

[0196]

[0197]

[0198] Following the Route B synthetic route, compound 28 was synthesized from 6-bromo-1H-indole as the starting material.

[0199]

[0200] Following the synthetic route of Route B, compound 16 was synthesized from 6-Bromo-1-methyl-1H-indole as the starting material.

[0201]

[0202] Following the synthetic route of Route B, compound 30 was synthesized from 6-Bromo-1-methyl-1H-indole as the starting material.

[0203]

[0204] Following the synthetic route of Route B, compound 40 was synthesized from 4-bromo-6-methoxy-1H-indole as the starting material.

[0205]

[0206] Following the synthetic route of Route B, compound 41 was synthesized from 4-bromo-6-methoxy-1H-indole as the starting material.

[0207]

[0208] Following the synthetic route of Route B, compound42 was synthesized from 4-bromo-1-methyl-6-methoxy-1H-indole as the starting material.

[0209]

[0210] Following the synthetic route of Route B, compound 43 was synthesized from 4-bromo-1-methyl-6-methoxy-1H-indole as the starting material.

[0211] Example 5: Synthesis of Compound 57

[0212]

[0213] The starting material Q1 (2.0 g, 9.09 mmol), EtOH (8.0 mL, 9.09 mmol), and ethylamine (22.73 mL, 45.45 mmol, 2 M THF solution) were added to a 100 mL single-necked flask, and the mixture was heated to 50 °C and stirred for 4 h. The reaction was confirmed by TLC to be complete. The solvent was evaporated, and the residue was diluted with 15 mL of H2O and stirred for 15 min. The solid was collected by filtration to give a red solid Q2 (2.00 g, yield 89.77%).

[0214] Intermediate Q2 (2.0 g, 8.16 mmol) was dissolved in acetone (16.0 mL), and H2O (2.0 mL) was added. NH4Cl (4.37 g, 81.61 mmol) and Zn (2.67 g, 40.80 mmol) were added at room temperature, and the mixture was stirred overnight at room temperature. The reaction proceeded to completion by TLC. The residue was filtered, the filtrate was concentrated, water was added, and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and then subjected to rotary evaporation followed by column chromatography (V). 石油醚 / V 乙酸乙酯 =50 / 1) yielded a pale red solid Q3 (1g, yield 56.97%).

[0215] Intermediate Q3 (1.0 g, 4.65 mmol) was dissolved in DMF (23.0 mL), and DIEA (1.80 g, 13.95 mmol) was added. The mixture was stirred at room temperature for 10 min. N-BOC-4-piperidinic acid (1.767 g, 4.65 mmol) and HATU (1.277 g, 5.58 mmol) were added at 0 °C, and the mixture was stirred overnight at room temperature. The reaction proceeded to complete reaction by TLC. Water was added, and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and then subjected to column chromatography (V... 石油醚 / V 乙酸乙酯 =5 / 1) to obtain pale red solid Q4 (1.83 g, yield 92.32%).

[0216] Intermediate Q4 (1.83 g, 4.29 mmol) was dissolved in glacial acetic acid (5.0 mL), heated to 100 °C, and stirred for 8 h. TLC analysis confirmed complete reaction of the starting material. The solvent was evaporated to dryness, and the solution was used directly in the next reaction without further purification. Intermediate Q5 was dissolved in DCM (15 mL), and TEA (433 mg, 4.29 mmol) and Boc₂O (935 mg, 4.29 mmol) were added at room temperature. The mixture was stirred at room temperature for 4 h. TLC analysis confirmed complete reaction of the starting material. Water was added, and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and then subjected to column chromatography (V... 石油醚 / V 乙酸乙酯 =1 / 1) to obtain pale yellow solid Q5 (1.2 g, two-step yield 69.50%).

[0217] Intermediate Q6 (408 mg, 1.00 mmol), 2,6-bis(benzyloxy)-3-(4,4,5,5-tetramethyl-1,3,2-dioxoboron-2-yl)pyridine (417 mg, 1.00 mmol), and [1,1'-bis(di-tert-butylphosphine)ferrocene]palladium(II) dichloride (73 mg, 0.10 mmol) were dissolved in 1,4-dioxane (12 mL) and 2M Na₂CO₃ (4 mL) aqueous solution. The reaction was carried out at 85 °C for 8 h under Ar atmosphere. The reaction proceeded to completion as determined by TLC. Water was added, and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and then subjected to rotary evaporation followed by column chromatography (V). 石油醚 / V 乙酸乙酯 =1 / 1) to obtain a pale yellow oily solid Q7 (600 mg, yield 94.27%).

[0218] Intermediate Q7 (300 mg, 0.48 mmol) was dissolved in tetrahydrofuran (9 mL) and ethanol (3 mL). Pd / C (100 mg, 0.096 mmol) and Pd(OH)₂ (35 mg, 0.048 mmol) were added, and the mixture was purged with H₂ three times. The reaction was carried out at room temperature for 6 h. TLC was used to confirm the complete reaction of the starting material. Pd / C and Pd(OH)₂ were filtered off, and the organic phase was evaporated to dryness before column chromatography (V) 二氯甲烷 / V 甲醇 =20 / 1) yields colorless solid Q8 (210 mg, yield 98.32%).

[0219] Intermediate Q8 (100 mg) was dissolved in dichloromethane (5 mL), and 4 M hydrochloric acid / ethyl acetate solution (2 mL) was added. The mixture was stirred at room temperature for 2 h. The reaction was confirmed by TLC to be complete. The mixture was filtered to obtain 70 mg of a pale red solid, with a yield of 91%.

[0220] Example 6 Synthesis of Compound 58

[0221]

[0222] Compound 58 was synthesized according to Example 57, except that the starting material ethylamine was replaced with isopropylamine. 34 mg of Compound 58 was obtained by filtration as a pale yellow powder.

[0223] Example 7 Synthesis of Compound 49

[0224]

[0225] S1 (1 g, 5.075 mmol), 1-Boc-4-methanesulfonyloxypiperidine (2.13 g, 7.612 mmol), and Cs2CO3 (4.96 g, 15.225 mmol) were dissolved in DMF (12.5 mL), stirred overnight at 80 °C, and the reaction was quenched with water after TLC detection of complete reaction. The mixture was extracted three times with EA, washed three times with saturated brine, dried, and the solvent was removed under reduced pressure to obtain crude product. The crude product was separated by column chromatography (PE:EA = 3:1) to obtain the target compound S2 (1.39 g, yield 97%).

[0226] Compound S2 (300 mg, 0.789 mmol), 2,6-di(benzyloxy)-3-(4,4,5,5-tetramethyl-1,3,2-dioxoboropentane-2-yl)pyridine (362 mg, 0.868 mmol), and 1,4-dioxane (6 mL) were dissolved in water. PdCl2 (dppf) (58 mg, 0.079 mmol) and saturated sodium bicarbonate solution (2 mL) were added, and the mixture was purged with argon gas and stirred overnight at 85 °C. The reaction was confirmed to be complete by TLC. The mixture was diluted with water, extracted three times with EA, washed once with saturated brine, dried, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was then separated by column chromatography (PE:EA = 3:1) to obtain the target compound S3 (274 mg, 59% yield).

[0227] Compound S3 (274 mg, 0.464 mmol) was dissolved in THF (3 mL), and 20% Pd(OH) was added. 2, (65.4 mg, 0.093 mmol), EtOH (1.5 mL), replaced with hydrogen gas, stirred overnight at room temperature. The reaction was confirmed to be complete by TLC. The mixture was filtered through diatomaceous earth, washed with THF and EtOH, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was then separated by column chromatography (PE:EA = 1:1) to obtain the target compound S4 (178 mg, 93% yield).

[0228] Compound S4 (178 mg, 0.432 mmol) was dissolved in DCM (3 mL), and 4 M dioxane hydrochloride solution (0.8 mL) was added. , The mixture was stirred at room temperature for 6 hours. The reaction was confirmed to be complete by TLC. The mixture was then filtered, washed with DCM, and the filter cake was the product 49 (95 mg, yield 63%).

[0229] Example 8 Synthesis of Compound 50

[0230]

[0231] Compound S5 (300 mg, 0.789 mmol), 2,6-di(benzyloxy)-3-(4,4,5,5-tetramethyl-1,3,2-dioxoboropentane-2-yl)pyridine (362 mg, 0.868 mmol), and 1,4-dioxane (6 mL) were dissolved in water. PdCl2 (dppf) (58 mg, 0.079 mmol) and saturated sodium bicarbonate solution (2 mL) were added, and the mixture was purged with argon gas and stirred overnight at 85 °C. The reaction was monitored by TLC until complete. The mixture was diluted with water, extracted three times with EA, washed once with saturated brine, dried, and the solvent was removed under reduced pressure to obtain the crude product. Column chromatography (PE:EA = 3:1) was performed to obtain the target compound S6 (300 mg, 64% yield).

[0232] Compound S6 (300 mg, 0.508 mmol) was dissolved in THF (3 mL), and 20% Pd(OH)2 (71.8 mg, 0.102 mmol) and EtOH (1.5 mL) were added. Hydrogen gas was introduced, and the mixture was stirred overnight at room temperature. The reaction was confirmed to be complete by TLC. The mixture was filtered through diatomaceous earth, washed with THF and EtOH, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was then separated by column chromatography (PE:EA = 1:1) to obtain the target compound S7 (134 mg, 64% yield).

[0233] Compound S7 (134 mg, 0.325 mmol) was dissolved in DCM (2 mL), and 4 M dioxane hydrochloride solution (0.6 mL) was added. , The mixture was stirred at room temperature for 6 hours. The reaction was confirmed to be complete by TLC. The mixture was then filtered, washed with DCM, and the filter cake was 50 mg (40 mg, 35% yield).

[0234] Example 9 Synthesis of Compound 44

[0235]

[0236] T1 (950 mg, 4.501 mmol), 1-Boc-4-methanesulfonyloxypiperidine (1.89 g, 6.752 mmol), and Cs2CO3 (4.4 g, 13.503 mmol) were dissolved in DMF (12 mL), stirred overnight at 80 °C, and the reaction was quenched with water after TLC detection of complete reaction. The mixture was extracted three times with EA, washed three times with saturated brine, dried, and the solvent was removed under reduced pressure to obtain crude product. The crude product was then separated by column chromatography (PE:EA = 3:1) to obtain the target compound T2 (1 g, yield 94%).

[0237] Compound T2 (700 mg, 1.774 mmol), 2,6-bis(benzyloxy)-3-(4,4,5,5-tetramethyl-1,3,2-dioxoboropentane-2-yl)pyridine (814 mg, 1.951 mmol), and 1,4-dioxane (18 mL) were dissolved in water. PdCl2 (dppf) (130 mg, 0.177 mmol) and saturated sodium bicarbonate solution (6 mL) were added, and the mixture was purged with argon gas and stirred overnight at 85 °C. The reaction was confirmed to be complete by TLC. The mixture was diluted with water, extracted three times with EA, washed once with saturated brine, dried, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was then separated by column chromatography (PE:EA = 3:1) to obtain the target compound T3 (440 mg, yield 41%).

[0238] Compound T3 (440 mg, 0.728 mmol) was dissolved in THF (4 mL), and 20% Pd(OH)2 and EtOH (2 mL) were added to replace the hydrogen gas. The mixture was stirred overnight at room temperature. The reaction was confirmed to be complete by TLC. The mixture was filtered through diatomaceous earth, washed with THF and EtOH, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was then separated by column chromatography (DCM:MeOH = 15:1) to obtain the target compound T4 (210 mg, yield 68%).

[0239] Compound T4 (210 mg, 0.492 mmol) was dissolved in DCM (5 mL), and 4 M hydrochloric acid-dioxane solution (1 mL) was added. The mixture was stirred at room temperature for 6 h. The reaction was confirmed to be complete by TLC. The mixture was filtered, washed with DCM, and the filter cake was the product 44 (160 mg, 90% yield).

[0240] Example 10 Synthesis of Compound 45

[0241]

[0242] Compound T5 (266 mg, 0.675 mmol), 2,6-di(benzyloxy)-3-(4,4,5,5-tetramethyl-1,3,2-dioxoboropentane-2-yl)pyridine (310 mg, 0.742 mmol), and 1,4-dioxane (7.5 mL) were dissolved in water. PdCl2 (dppf) (49 mg, 0.068 mmol) and saturated sodium bicarbonate solution (2.5 mL) were added, and the mixture was purged with argon gas and stirred overnight at 85 °C. The reaction was monitored by TLC until complete. The mixture was diluted with water, extracted three times with EA, washed once with saturated brine, dried, and the solvent removed under reduced pressure to obtain the crude product. Column chromatography (PE:EA = 3:1) was performed to obtain the target compound T6 (167 mg, 41% yield).

[0243] Compound T6 (167 mg, 0.276 mmol) was dissolved in THF (2 mL), and 20% Pd(OH) was added. 2, Add 1 mL of EtOH, replace with hydrogen gas, and stir overnight at room temperature. The reaction was confirmed to be complete by TLC. The mixture was filtered through diatomaceous earth, washed with THF and EtOH, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was then separated by column chromatography (DCM:MeOH = 15:1) to obtain the target compound T7 (80 mg, yield 68%).

[0244] Compound T7 (80 mg, 0.188 mmol) was dissolved in DCM (2 mL), and 4 M hydrochloric acid-dioxane solution (0.4 mL) was added. , The mixture was stirred at room temperature for 6 hours. The reaction was monitored by TLC until it was complete. The mixture was then filtered, washed with DCM, and the filter cake was the product 45 (60 mg, yield 90%).

[0245] Example 11 Synthesis of Compound 46

[0246]

[0247] U1 (950 mg, 4.501 mmol), 1-Boc-4-methanesulfonyloxypiperidine (1.89 g, 6.752 mmol), and Cs2CO3 (4.4 g, 13.503 mmol) were dissolved in DMF (12 mL), stirred overnight at 80 °C, and the reaction was quenched with water after TLC detection of complete reaction. The mixture was extracted three times with EA, washed three times with saturated brine, dried, and the solvent was removed under reduced pressure to obtain crude product. The crude product was separated by column chromatography (PE:EA = 3:1) to obtain the target compound U2 (1.1 g, yield 98%).

[0248] Compound U2 (300 mg, 0.761 mmol), 2,6-di(benzyloxy)-3-(4,4,5,5-tetramethyl-1,3,2-dioxoboropentane-2-yl)pyridine (350 mg, 0.837 mmol), and 1,4-dioxane (6 mL) were dissolved in water. PdCl2 (dppf) (56 mg, 0.076 mmol) and saturated sodium bicarbonate solution (2 mL) were added, and the mixture was purged with argon gas and stirred overnight at 85 °C. The reaction was confirmed to be complete by TLC. The mixture was diluted with water, extracted three times with EA, washed once with saturated brine, dried, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was then separated by column chromatography (PE:EA = 3:1) to obtain the target compound U3 (390 mg, yield 85%).

[0249] Compound U3 (390 mg, 0.645 mmol) was dissolved in THF (4 mL), and 20% Pd(OH) was added. 2, (91 mg 0.129 mmol), EtOH (2 mL), replaced with hydrogen gas, stirred overnight at room temperature. The reaction was confirmed to be complete by TLC. The mixture was filtered through diatomaceous earth, washed with THF and EtOH, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was then separated by column chromatography (PE:EA = 1:1) to obtain the target compound U4 (80 mg, yield 30%).

[0250] Compound U4 (80 mg, 0.188 mmol) was dissolved in DCM (2 mL), and 4 M hydrochloric acid-dioxane solution (0.4 mL) was added. The mixture was stirred at room temperature for 6 h. The reaction was confirmed to be complete by TLC. The mixture was filtered, washed with DCM, and the filter cake was the product 46 (60 mg, 90% yield).

[0251] Example 12 Synthesis of Compound 47

[0252]

[0253] The synthesis of compound 47 follows the same route as compound 44, except that T1 is replaced with Z1 to obtain compound 47.

[0254] Example 13 Synthesis of Compound 48

[0255]

[0256] V1 (950 mg, 4.184 mmol), 1-Boc-4-methanesulfonyloxypiperidine (1.75 g, 6.276 mmol), and Cs2CO3 (4.09 g, 12.552 mmol) were dissolved in DMF (10 mL), stirred overnight at 80 °C, and the reaction was quenched with water after TLC detection of complete reaction. The mixture was extracted three times with EA, washed three times with saturated brine, dried, and the solvent was removed under reduced pressure to obtain crude product. The crude product was then separated by column chromatography (PE:EA = 3:1) to obtain the target compound V2 (510 mg, yield 84%).

[0257] Compound V2 (300 mg, 0.731 mmol), 2,6-bis(benzyloxy)-3-(4,4,5,5-tetramethyl-1,3,2-dioxoboropentane-2-yl)pyridine (336 mg, 0.804 mmol), and 1,4-dioxane (6 mL) were dissolved in water. PdCl2 (dppf) (53 mg, 0.073 mmol) and saturated sodium bicarbonate solution (2 mL) were added, and the mixture was purged with argon gas and stirred overnight at 85 °C. The reaction was monitored by TLC until complete. The mixture was diluted with water, extracted three times with EA, washed once with saturated brine, dried, and the solvent was removed under reduced pressure to obtain the crude product. Column chromatography (PE:EA = 3:1) was performed to obtain the target compound V3 (400 mg, 88% yield).

[0258] Compound V3 (400 mg, 0.644 mmol) was dissolved in THF (4 mL), and 20% Pd(OH) was added. 2, (91 mg 0.129 mmol), EtOH (2 mL), replaced with hydrogen gas, stirred overnight at room temperature. The reaction was confirmed to be complete by TLC. The mixture was filtered through diatomaceous earth, washed with THF and EtOH, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was then separated by column chromatography (PE:EA = 1:1) to obtain the target compound V4 (230 mg, yield 81%).

[0259] Compound V4 (230 mg, 0.520 mmol) was dissolved in DCM (3 mL), and 4 M hydrochloric acid-dioxane solution (1 mL) was added. The mixture was stirred at room temperature for 6 h. The reaction was confirmed to be complete by TLC. The mixture was filtered, washed with DCM, and the filter cake was the product 48 (140 mg, yield 71%).

[0260] Example 14 Synthesis of Compound 51

[0261]

[0262] Compound V5 (300 mg, 0.731 mmol), 2,6-di(benzyloxy)-3-(4,4,5,5-tetramethyl-1,3,2-dioxoboropentane-2-yl)pyridine (336 mg, 0.804 mmol), and 1,4-dioxane (6 mL) were dissolved in water. PdCl2 (dppf) (53 mg, 0.073 mmol) and saturated sodium bicarbonate solution (2 mL) were added, and the mixture was purged with argon gas and stirred overnight at 85 °C. The reaction was monitored by TLC until complete. The mixture was diluted with water, extracted three times with EA, washed once with saturated brine, dried, and the solvent was removed under reduced pressure to obtain the crude product. Column chromatography (PE:EA = 3:1) was performed to obtain the target compound V6 (450 mg, 99% yield).

[0263] Compound V6 (400 mg, 0.644 mmol) was dissolved in THF (4 mL), and 20% Pd(OH) was added. 2, (91 mg, 0.129 mmol), EtOH (2 mL), replaced with hydrogen gas, stirred overnight at room temperature. The reaction was confirmed to be complete by TLC. The mixture was filtered through diatomaceous earth, washed with THF and EtOH, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was then separated by column chromatography (PE:EA = 1:1) to obtain the target compound V7 (72 mg, 25% yield).

[0264] Compound V7 (72 mg, 0.163 mmol) was dissolved in DCM (2 mL), and 4 M hydrochloric acid-dioxane solution (0.4 mL) was added. The mixture was stirred at room temperature for 6 h. The reaction was confirmed to be complete by TLC. The mixture was filtered, washed with DCM, and the filter cake was the product 51 (28 mg, yield 45%).

[0265] Example 15 Synthesis of compounds 52 and 53

[0266]

[0267] Starting from raw material W5, and referring to the synthesis method of intermediate J6 of compound D34, W8 100 mg was obtained. W8' 150 mg was obtained.

[0268] The synthesis of compound 52 is based on the method used for the synthesis of 46.

[0269] The synthesis of compound 53 is based on the method used to synthesize compound 46.

[0270] Example 16 Synthesis of Compound 54

[0271]

[0272] Compound W9 (1 g, 5.08 mmol) was dissolved in anhydrous DMF (5 mL). I2 (1.310 g, 10.16 mmol) was added to the system at 0 °C, and the mixture was stirred overnight at room temperature. The reaction was monitored by TLC until complete, and the reaction was quenched with water. The aqueous phase was extracted with ethyl acetate, the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and then subjected to column chromatography (V...). 石油醚 / V 乙酸乙酯 =10 / 1) yielded 1.21 g of a pale red oily solid, with a yield of 73.83%.

[0273] Intermediate W10 (1.21 g, 3.75 mmol), N-Boc-1,2,5,6-tetrahydropyridine-4-boronic acid pinacol ester (1.159 g, 3.75 mmol), and [1,1'-bis(di-tert-butylphosphine)ferrocene]palladium(II) dichloride (274 mg, 0.375 mmol) were dissolved in 1,4-dioxane (8 mL) and 2M Na₂CO₃ (4 mL) aqueous solution. The reaction was carried out overnight at 85 °C under Ar atmosphere. The reaction proceeded to completion as determined by TLC. The reaction mixture was extracted with ethyl acetate and water, and the organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and then subjected to rotary evaporation followed by column chromatography (V). 石油醚 / V 乙酸乙酯 =5 / 1) yielded 1.30 g of brown oily solid, with a yield of 91.72%.

[0274] Intermediate W11 (1.30 g, 3.44 mmol) and PtO2 (78 mg, 0.344 mmol) were dissolved in acetic acid (8 mL), substituted with H2 three times, and reacted overnight at room temperature with stirring. TLC was performed to confirm complete reaction of the starting material. The pH was adjusted to weakly alkaline by adding saturated sodium bicarbonate solution, and the mixture was extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and then subjected to column chromatography (V... 石油醚 / V 乙酸乙酯 =5 / 1) yielded 900 mg of brown oily solid, with a yield of 68.86%.

[0275] The synthesis of intermediate W13 was carried out by referring to the synthesis method of intermediate O2 in the compound synthesis route of compound D45, yielding 600 mg of intermediate W13 with a yield of 75.44%.

[0276] The synthesis of intermediate W14 was based on the synthesis of intermediate J6 of compound D34, yielding intermediate W14, 400 mg, in 57.96% yield.

[0277] The synthesis of intermediate W15 was based on the synthesis of intermediate J6 of compound D34, yielding intermediate W15, 300 mg, in a yield of 47.26%.

[0278] The synthesis of compound 54 was performed with reference to the synthesis of compound 46, yielding 5450 mg in 65.34% yield.

[0279] Example 17 Synthesis of Compound 55

[0280]

[0281] The starting material W16 (900 mg, 4.2 mmol) and 1,4-dibromobutane (1.0 g, 4.7 mmol) were added sequentially to a reaction flask. 17 mL of a 1.0 M LiHMDS tetrahydrofuran solution was added to the flask. After the reaction was monitored by TLC until complete, water was added to quench the reaction, and the mixture was extracted with EA. After drying and concentration, the mixture was separated by column chromatography (PE:EA = 1:1) to obtain the target compound W17 in 81% yield.

[0282] Starting from intermediate W17, compound 55 was obtained.

[0283] Example 18 Synthesis of Compound 56

[0284]

[0285] The synthesis of compound 56 was carried out starting from raw material W19, following the synthesis method of compound 58 (originally numbered).

[0286] Example 19 Synthesis of Compound 59

[0287]

[0288] The synthesis of compound 59 started from raw material W24 and followed the synthesis method of intermediate O2 in route D45.

[0289] Example 20 Synthesis of Compound C9

[0290] Route C:

[0291]

[0292] In a flask dried in an oven and purged with argon, starting material C2 (1.0 equiv.) and dry DMF were added. The reaction system was placed in an ice-water bath at 0°C, followed by the addition of NaH (1.5 equiv.) and stirring at 0°C for 0.5 h. Then, a DMF solution containing C1 (1.1 equiv.) was added dropwise, and the mixture was stirred at 0°C for another 0.5 h. The reaction was then continued overnight at room temperature with stirring. TLC showed no remaining starting material. The reaction was quenched with water, and the aqueous phase was extracted with EtOAc. The combined organic phases were washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The intermediate was then purified by silica gel column chromatography. This intermediate was added to a reaction flask, dissolved in DCM, and a dioxane solution of HCl (5.0 equiv.) was added dropwise. The reaction was carried out at room temperature for 2 h. TLC showed no remaining starting material. No further treatment was required, and the solvent was directly removed under reduced pressure to obtain intermediate C3.

[0293] In a flask that has been dried in an oven and purged with argon, add starting material C4 (1.0 equiv.), purge with argon, add dry THF using a syringe, purge with argon again, add SOCl2 (1.5 equiv.) dropwise at room temperature, then heat to 80°C and stir for 4 hours. TLC analysis shows no remaining raw material, yielding a THF solution of intermediate C5. Draw it out with a syringe for later use, taking care to avoid water.

[0294] In an oven-dried and purged flask, intermediate C3 (1.0 equiv.) was added and purged with argon. Then, dry THF was added, and DIPEA (3.0 equiv.) was added dropwise at room temperature. The mixture was stirred for 20 min, and then the prepared intermediate C5-THF solution was added dropwise. The reaction was allowed to proceed overnight at room temperature. TLC showed no residual starting material. The reaction was quenched with water, and the aqueous phase was extracted with DCM. The combined organic phases were washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The intermediate C6 was then purified by silica gel column chromatography.

[0295] In an oven-dried and purged flask, intermediate C6 (1.0 equiv.), starting material C7 (1.2 equiv.), K2CO3 (2.0 equiv.), and DMF were added. The mixture was then heated to 80°C and stirred overnight. TLC analysis showed no remaining raw material. The DMF was dried, and the residue was purified by silica gel rapid column chromatography to obtain intermediate C8.

[0296] In an oven-dried and purged flask, intermediate C7 (1.0 equiv.) and DCM were added and dissolved. The flask was then placed in an ice-water bath, and a DCM solution containing DMP (2.0 equiv.) was added dropwise. The reaction was carried out at 0°C for 0.5 h, followed by a reaction at room temperature for 1 h. TLC showed no residual raw material. The reaction residue was filtered off, and the filtrate was concentrated and purified by silica gel rapid column chromatography to obtain intermediate C9.

[0297] Example 21 Synthesis of compound C11

[0298] Route D:

[0299]

[0300] Intermediate C6 (1.0 equiv.) obtained according to Route C, tert-butyl2,7-diazaspiro[4.4]nonane-2-carboxylate (1.5 equiv.), and DIPEA (2.5 equiv.) were added to a reaction flask, dissolved in isopropanol, and then heated to 90°C and reacted for 12 h. TLC showed no raw material remaining. The mixture was extracted with DCM, washed twice with saturated ammonium chloride solution, washed with saturated sodium chloride solution, dried with anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The mixture was then purified by silica gel rapid column chromatography to obtain intermediate C10.

[0301] Intermediate C10 was placed in a reaction flask, dissolved in DCM, and then a dioxane solution of HCl (5.0 equiv.) was added dropwise. The reaction was carried out at room temperature for 2 hours. TLC showed no raw material remaining. No further treatment was required. The solvent was directly evaporated under reduced pressure to obtain intermediate C11.

[0302] Example 22 Synthesis of compounds C13 and C15

[0303] Route E:

[0304]

[0305] Following the first step of Route D, tert-butyl 2,7-diazaspiro[3.5]nonane-2-carboxylate was substituted with intermediate C6 to obtain intermediate C12.

[0306] Following the second step of Route D, intermediate C12 undergoes deBoc removal to yield compound C13.

[0307] Route F:

[0308]

[0309] Following the first step of Route D, tert-butyl piperazine-1-carboxylate was substituted with intermediate C6 to obtain intermediate C14.

[0310] Following the second step of Route D, intermediate C14 undergoes deBoc removal to yield compound C15.

[0311] Example 23 Synthesis of compound D10

[0312] Route G:

[0313]

[0314] In an oven-dried flask purged with argon, starting material P1 (1.0 equiv.), tert-butyl 2,2,2-trichloroacetimidate (1.0 equiv.), and PPTS (0.1 equiv.) were added sequentially. DCM was added to dissolve the reactants, and the reaction was carried out at room temperature. Every 4-5 hours, tert-butyl 2,2,2-trichloroacetimidate (0.2 equiv.) and PPTS (0.02 equiv.) were added. After three additions, TLC showed no remaining reactants. The reactants were extracted with DCM, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The intermediate P2 was then purified by silica gel column chromatography.

[0315] Intermediate P2 (1.0 equiv.) was added to a flask that had been dried in an oven and purged with argon. Argon was then used to purge the flask, and dry THF was added using a syringe. After dissolving, the reaction system was cooled to -78°C. A THF solution of KHDMS (1.3 equiv.) was slowly added dropwise, and the reaction was continued at -78°C for 0.5 hours. Then, a solution containing...

[0316] A THF solution of 1,1,1-trifluoro-N-phenyl-N-((trifluoromethyl)sulfonyl)methanesulfonamide (1.2 equiv.) was reacted overnight at room temperature. TLC showed no residual starting material. The mixture was extracted with EtOAc, washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The intermediate P3 was then purified by silica gel column chromatography.

[0317] In a Schlenk reaction tube that has been dried in an oven and purged with argon, intermediate P3 (1.0 equiv.) is added.

[0318] Argon gas was used to replace (4-hydroxyphenyl)boronic acid (1.2 equiv.), K2CO3 (2.0 equiv.), and Pd(dppf)Cl2 (0.1 equiv.). Dioxane and water were added using a syringe, and after dissolution, the reaction system was heated to 100°C and reacted overnight. TLC showed no residual raw material. The mixture was extracted with EtOAc, washed with saturated ammonium chloride solution and saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The intermediate P4 was then purified by silica gel column chromatography.

[0319] In a reaction flask dried in an oven, intermediate P4 (1.0 equiv.) was added and dissolved in acetonitrile. NBS (0.9 equiv.) was slowly added and the reaction was carried out at room temperature for 1 h. TLC showed no residual starting material. The mixture was extracted with EtOAc, washed with saturated ammonium chloride solution and saturated sodium chloride solution, dried with anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The intermediate P5 was then purified by silica gel column chromatography.

[0320] In a Schlenk reaction tube that has been dried in an oven and purged with argon, intermediate P5 (1.0 equiv.), phenylboronic acid (1.2 equiv.), K2CO3 (2.0 equiv.), and Pd(dppf)Cl2 (0.1 equiv.) were added and purged with argon. Dioxane and water were added using a syringe to dissolve the mixture. The reaction system was then heated to 100°C and reacted overnight. TLC analysis showed no residual raw material. The mixture was extracted with EtOAc, washed with saturated ammonium chloride solution and saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The intermediate P6 was then purified by silica gel column chromatography.

[0321] In an oven-dried reaction flask, intermediate P6 (1.0 equiv.) was added and dissolved in a mixed solvent of MeOH:THF = 3:1. 10% Pd / C (10 wt%) was added, and the reaction was carried out overnight at room temperature in a hydrogen atmosphere. TLC showed no residual raw material. The mixture was filtered through diatomaceous earth onto palladium on carbon, and after solvent removal under reduced pressure, it was purified by silica gel rapid column chromatography to obtain intermediate P7.

[0322] In a reaction flask that has been dried in an oven and purged with argon, intermediate P7 (1.0 equiv.) was added and dissolved in a THF:MeCN mixture of 1:1. Then, K2CO3 (2.0 equiv.) and 1,1,2,2,3,3,4,4,4-nonafluorobutane-1-sulfonyl fluoride (1.5 equiv.) were added. The reaction was carried out at room temperature for 18 h. TLC analysis showed no residual starting material. The mixture was extracted with EtOAc, washed with saturated ammonium chloride and saturated sodium chloride solutions, dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The resulting product was purified by silica gel column chromatography to obtain intermediate P8.

[0323] In a Schlenk reaction tube that had been dried in an oven and purged with argon, intermediate P8 (1.0 equiv.), 4-(dimethoxymethyl)piperidine (2.0 equiv.), Cs2CO3 (3.0 equiv.), Pd(OAc)2 (0.2 equiv.), and BINAP (0.4 equiv.) were added and purged with argon. Dioxane was added using a syringe, and the reaction system was heated to 105°C and reacted overnight. TLC showed no residual starting material. The mixture was extracted with EtOAc, washed with saturated ammonium chloride solution and saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The intermediate P9 was then purified by silica gel column chromatography.

[0324] In a reaction flask that has been dried in an oven and purged with argon, intermediate P9 (1.0 equiv.) was added, dissolved in THF, and H2SO4 (10.0 equiv.) was added dropwise. The reaction was carried out at 60°C for 2 hours. TLC showed no residual raw material. The mixture was extracted with EtOAc, washed with saturated sodium carbonate solution and saturated sodium chloride solution, dried with anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The intermediate P10 was then purified by silica gel column chromatography.

[0325] Example 24 Synthesis of compound E11

[0326] Route H:

[0327]

[0328] In a flask dried in an oven and purged with argon, starting materials E1 (1.0 equiv.), E2 (1.2 equiv.), ACN, and DIPEA (1.2 equiv.) were added. After purging with argon, the reaction mixture was heated overnight in an oil bath at 60°C. TLC monitoring showed no remaining E1. After the reaction system cooled to room temperature, water and DCM were added for dilution. The resulting aqueous phase was extracted with DCM, and the combined organic phases were washed with saturated brine and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate E3 was obtained.

[0329] In an oven-dried flask purged with argon, starting material E4 (1.0 equiv.) was added, along with DCM, TsCl (1.2 equiv.), DMAP (0.5 equiv.), and Et3N (1.2 equiv.). The mixture was stirred at room temperature for 2 hours. TLC monitoring showed no remaining E4. The mixture was diluted with water and DCM, and the resulting aqueous phase was extracted with DCM. The combined organic phases were washed with saturated brine and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, E5 was obtained.

[0330] In an oven-dried flask purged with argon, intermediate E5 (1.0 equiv.), commercial reagent E6 (1.2 equiv.), DMF, and K2CO3 (1.2 equiv.) were added. The mixture was heated and stirred in an oil bath at 60°C for 12 h. TLC monitoring showed no remaining reactants. After the reaction system cooled to room temperature, water and DCM were added for dilution. The resulting aqueous phase was extracted with DCM, and the combined organic phases were washed with saturated brine and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, E7 was obtained.

[0331] In a flask that has been dried in an oven and purged with argon, intermediate E7, THF, and 10% Pd / C (20 wt%) were added. Hydrogen was purged during vigorous stirring, and the reaction mixture was stirred vigorously for 8 hours under hydrogen balloon pressure. TLC monitoring showed no remaining raw material. The reaction mixture was filtered through silica gel cake and evaporated under reduced pressure to obtain crude product E8, which could be used directly in the next reaction without further purification.

[0332] In an oven-dried and purged flask, intermediate E8 (1.0 equiv.) dissolved in THF-MeOH (8:1, v:v, 0.2 M) was added. LiBH4 (1.2 equiv.) was slowly added upwards at 0°C, and the reaction mixture was stirred at 60°C for 2 h. TLC analysis showed no remaining starting material. After the reaction system cooled to room temperature, an appropriate amount of water was added to quench the reaction. The aqueous phase was extracted with DCM, and the combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate E9 was obtained.

[0333] In an oven-dried flask purged with argon, starting materials E9 (1.0 equiv.), E3 (1.2 equiv.), anhydrous ACN, NMI (2 equiv.), and TCFH (1.2 equiv.) were added. The mixture was stirred at room temperature for 6 hours. TLC monitoring showed no remaining raw material. Water and DCM were added for dilution, and the resulting aqueous phase was extracted with DCM. The combined organic phases were washed with saturated brine and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, E10 was obtained.

[0334] In a flask dried in an oven and purged with argon, E10 (1.0 equiv.), DCM, and DMP (1.2 equiv.) were added, and the reaction mixture was stirred at room temperature for 3 hours. TLC monitoring showed no remaining starting material. Water and DCM were added to dilute the reaction mixture, and the resulting aqueous phase was extracted with DCM. The combined organic phases were washed with saturated brine and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, compound E11 was obtained.

[0335] Example 25 Synthesis of Compound F7

[0336] Route I:

[0337]

[0338] In an oven-dried flask purged with argon, starting materials F1 (1.0 equiv.), F2 (1.0 equiv.), DMF, and DIPEA (2.0 equiv.) were added. The mixture was stirred at room temperature for 12 h. TLC monitoring showed no remaining raw materials. Water and DCM were added for dilution, and the resulting aqueous phase was extracted with DCM. The combined organic phases were washed with saturated brine and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate F3 was obtained.

[0339] In a flask that has been dried in an oven and purged with argon three times, intermediates F3 (1.0 equiv.), F4 (1.1 equiv.), Cs₂CO₃ (3.0 equiv.), Pd(OAc)₂ (0.1 equiv.), and BINAP (0.2 equiv.) were added. The mixture was purged with argon three times, and then dried Dioxane was injected. The reaction system was heated to 100°C in an oil bath and stirred for 20 h under an argon atmosphere. TLC showed no residual starting material. After cooling the reaction system to room temperature, water and EtOAc were added for dilution. The aqueous phase was extracted with EtOAc, and the combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate F5 was obtained.

[0340] In an oven-dried flask purged with argon, F5 (1.0 equiv.) and DMSO were added. Cs₂CO₃ (2.0 equiv.) was added upwards to the solution with stirring at 0°C, followed by slow addition of 30% H₂O₂ solution (1.2 equiv.). The reaction mixture was stirred at room temperature for 3 hours. TLC analysis showed no remaining starting material. The reaction mixture was diluted with water and EtOAc, and the aqueous phase was extracted with EtOAc. The combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate F6 was obtained.

[0341] Intermediate F6 (1.0 equiv.) was dissolved in DCM, and a dioxane solution of HCl (4 M, 8.0 equiv.) was added upwards to the solution with stirring at 0 °C. The reaction system was stirred at room temperature for 2 h. TLC showed no reactant remaining, and the solvent was removed under reduced pressure to obtain compound F7.

[0342] Example 26 Synthesis of compounds G8, G9, G10, G11, G12, G13, and G14

[0343] Route J:

[0344]

[0345] In a flask that has been dried in an oven and purged with argon three times, intermediates G1 (1.0 equiv.), G2 (1.1 equiv.), CuI (0.1 equiv.), and (Ph3P)2PdCl2 (0.1 equiv.) were added. The mixture was purged with argon three times, and then injected with dry THF and Et3N (2 equiv.). The reaction system was stirred at room temperature for 20 h. TLC showed no residual starting material. Water and EtOAc were added to dilute the reaction system, and the aqueous phase was extracted with EtOAc. The combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate G3 was obtained.

[0346] In a flask that has been dried in an oven and purged with argon three times, intermediate G3 (1.0 equiv.) and PdCl2 (0.1 equiv.) were added. The mixture was purged with argon three times, and then injected with dry MeCN-DMF (1:1, v:v). The reaction system was heated overnight in an oil bath at 80°C. TLC analysis showed no remaining starting material. Water and EtOAc were added to dilute the reaction system, and the aqueous phase was extracted with EtOAc. The combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate G4 was obtained.

[0347] Add NaH (1.5 equiv.) to 5 mL of a dry DMF solution of G4 (1.0 equiv.), stir the reaction mixture at room temperature for 30 min, add iodomethane (1.2 equiv.), and continue stirring the reaction mixture under an argon atmosphere for 4 h. TLC analysis showed no remaining starting material. Dilute the reaction mixture with water and EtOAc, extract the aqueous phase with EtOAc, combine the organic phases, wash with saturated sodium chloride solution, and dry with anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, obtain the target intermediate G5.

[0348] Intermediate G5 and A3 are synthesized together as in the second step of RouteA to obtain intermediate G6.

[0349] In a flask that has been dried in an oven and purged with argon, intermediate G6 was added, followed by MeOH-THF (0.2 M, 1:1, v / v) and 10% Pd / C (20 wt%). Hydrogen was purged during vigorous stirring, and the reaction mixture was stirred vigorously under hydrogen balloon pressure for 8 hours. TLC monitoring showed no remaining starting material, and LC-MS monitoring showed no unreacted intermediates. The reaction mixture was filtered through silica gel cake and evaporated under reduced pressure to obtain crude product G7, which could be used directly in the next reaction without further purification.

[0350] Intermediate G7 is synthesized according to step four of Route H to obtain compound G8, G9, G10, or G11.

[0351] Intermediate G7 is synthesized according to step seven of Route G to obtain compound G12, G13, or G14.

[0352] Example 27 Synthesis of compound G20

[0353] Route K:

[0354]

[0355] In a flask that has been dried in an oven and purged with argon three times, starting compounds G15 (1.0 equiv.), G16 (1.1 equiv.), DMF, HATU (2.0 equiv.), and DIPEA (3.0 equiv.) were added. The reaction system was stirred at room temperature for 5 hours. TLC showed no residual starting material. Water and DCM were added to dilute the reaction system, and the aqueous phase was extracted with DCM. The combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate G17 was obtained.

[0356] Intermediate G17 and A3 were synthesized via a Suzuki reaction according to the second step of Route A to obtain intermediate G18; intermediate G18 was synthesized via catalytic hydrogenation according to the fifth step of Route I to obtain intermediate G19; intermediate G19 was synthesized via the fourth step of Route H to obtain the final product G20.

[0357] Example 28 Synthesis of compounds H11, H12, and H13

[0358] Route L:

[0359]

[0360] In a flask that has been dried in an oven and purged with argon three times, intermediates H1 (1.0 equiv.), H2 (1.1 equiv.), MeCN, and DMCS (0.1 equiv.) were added. The mixture was purged with argon three times, and then dried THF and Et3N (2 equiv.) were injected. The reaction system was stirred at room temperature for 20 h. TLC showed no residual starting material. Water and EtOAc were added to dilute the reaction system, and the aqueous phase was extracted with EtOAc. The combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate G3 was obtained.

[0361] Following the third step of Route I, intermediate H3 undergoes a substitution reaction to yield intermediate H4; intermediate H4 and A3 are then synthesized via a Suzuki reaction, following the second step of Route A, to yield intermediate H5.

[0362] Intermediate H5 (1.0 equiv.) was dissolved in dry DCM. DIBAL-H (1.0 min hexanes, 1.2 equiv.) was added dropwise to the solution at -78°C, and the reaction mixture was stirred at the same temperature for 2 h. TLC showed no residual starting material. The reaction was quenched with methanol, and water and a saturated potassium sodium tartrate solution were added and stirred at room temperature for 30 min. The aqueous phase was extracted with EtOAc, and the combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate H6 was obtained.

[0363] Intermediate H6 is used to obtain intermediate H7 according to the second step of Route G.

[0364] Add NaH (1.5 equiv.) to 5 mL of dried DMF solution of H7 (1.0 equiv.), stir the reaction mixture at room temperature for 30 min, then add H8 (1.2 equiv.), and continue stirring the reaction mixture under an argon atmosphere for 4 h. TLC analysis showed no remaining starting material. Dilute the reaction mixture with water and EtOAc, extract the aqueous phase with EtOAc, combine the organic phases, wash with saturated sodium chloride solution, and dry with anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, obtain the target intermediate H9.

[0365] Following the fifth step of Route I, intermediate H9 is synthesized into intermediate H10 via catalytic hydrogenation; intermediate H10 is then synthesized into final product H11, H12, or H13, following the fourth step of Route H.

[0366] Example 29 Synthesis of compounds H19 and H21

[0367] Route M:

[0368]

[0369] Intermediate H1 and intermediate H14 were synthesized according to the first step of Route K to obtain intermediate H15; according to the third step of Route I, intermediate H15 was substituted to obtain intermediate H16; intermediate H16 and A3 were synthesized according to the second step of Route A via a Suzuki reaction to obtain intermediate H17; intermediate H17 was synthesized according to the fifth step of Route I via a catalytic hydrogenation reaction to obtain intermediate H18; intermediate H18 was synthesized according to the fourth step of Route H to obtain intermediate H19.

[0370] NaBH(OAc)3 (2.5 equiv.) and intermediate I1 (1.2 equiv.) were added to a DCE solution of intermediate H19 (1.0 equiv.) (0.3 M) under stirring at 0 °C. The reaction mixture was stirred at room temperature for 8–18 h. LC-MS showed no residual raw material. The mixture was extracted with a DCM mixed solvent, and the combined organic phases were dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by preparative HPLC, intermediate H20 was obtained.

[0371] Intermediate H20 was synthesized in step 4 of Route H to obtain intermediate H21.

[0372] Example 30 Synthesis of compounds H27 and H28

[0373] Route N:

[0374]

[0375] Following the third step of Route I, intermediate H22 undergoes a substitution reaction to yield intermediate H23.

[0376] In a flask that has been dried in an oven and purged with argon three times, intermediates H23 (1.0 equiv.), G12 (1.1 equiv.), and PTS (0.1 equiv.) were added. The mixture was purged with argon three times, and then a dry toluene solution was injected. The reaction system was heated overnight in an oil bath at 80°C. TLC analysis showed no remaining starting material. Water and EtOAc were added to dilute the reaction system, and the aqueous phase was extracted with EtOAc. The combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate H24 was obtained.

[0377] Intermediate H24 and A3 were synthesized via the Suzuki reaction according to the second step of Route A to obtain intermediate H25; intermediate H25 was synthesized via catalytic hydrogenation according to the fifth step of Route I to obtain intermediate H26; intermediate H26 was synthesized via the fourth step of Route H to obtain the final product H27 or H28.

[0378] Example 31: Synthesis of compounds J6, J7, J8, J9, J10, J11, and J12

[0379] Route O:

[0380]

[0381] In a flask dried in an oven and purged with argon, starting material J1 (1.0 equiv.) and dried DMF were added. The reaction system was placed in an ice-water bath at 0°C, followed by the addition of NaH (1.5 equiv.) and stirring at 0°C for 0.5 h. Then, a DMF solution containing J2 (1.1 equiv.) was added dropwise, and the mixture was stirred at 0°C for 0.5 h. The reaction was then continued overnight at 80°C with stirring. TLC showed no remaining starting material. After the reaction system cooled to room temperature, water was added to quench the reaction. The aqueous phase was extracted with EtOAc, and the combined organic phases were washed with saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The intermediate J3 was then purified by silica gel column chromatography.

[0382] Intermediate J3 and A3 were synthesized via a Suzuki reaction following the second step of Route A to obtain intermediate J4; intermediate J4 was synthesized via a catalytic hydrogenation reaction following the fifth step of Route I to obtain intermediate J5; intermediate J5 was synthesized via the fourth step of Route H to obtain final products J6, J7, J8, J9, J10, J11, and J12.

[0383] Example 32 Synthesis of compounds N6, N7, N8, N9, N10, N11, N12, N13, N14, N15, and N16 Route P:

[0384]

[0385]

[0386] In oven-dried reaction flasks, different starting materials, bromoindole N1 (1.0 equiv.) and different NBoc-R2CO (ketone or aldehyde, 1.3 equiv.), were added and dissolved in DCM. The reaction was carried out at 40°C for 2 h, then cooled to room temperature, and NaBH(OAc)3 was added. The reaction was carried out at room temperature for 3-6 h. TLC showed no residual starting material. The product was extracted with EA, washed with saturated ammonium chloride solution and saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and the solvent was removed under reduced pressure. The intermediate N2 was purified by silica gel column chromatography with a yield of 78-91%.

[0387] Intermediate N2 and A3 were synthesized via the Suzuki reaction, following the second step of Route A, to obtain intermediate N3.

[0388] In an oven-dried and purged flask, intermediate N3 (1.0 equiv.) was added and dissolved in dioxane. The mixture was then cooled to 5°C, and a dioxane solution of DDQ (1.2 equiv.) was slowly added dropwise. The reaction was then carried out at room temperature for 1-4 hours. TLC monitoring showed no remaining starting material, and LC-MS monitoring showed no unreacted intermediate. The mixture was diluted with saturated sodium bicarbonate solution and EtOAc. The organic phase was washed with saturated brine and dried over anhydrous sodium sulfate. After filtration and solvent removal under reduced pressure, intermediate N4 was purified by silica gel column chromatography in 57-85% yield.

[0389] Intermediate N4 was synthesized via catalytic hydrogenation according to step 5 of Route I to obtain intermediate N5, with a yield of 43-75%; intermediate N5 was synthesized to obtain final products N6-N16 with a yield of 90-99% according to step 4 of Route H.

[0390] Example 33 Synthesis of compound K4

[0391] Route Q:

[0392]

[0393] In a reaction flask dried in an oven and purged with argon, starting material K1 (1.0 equiv.), benzylcarbonobromidate (1.2 equiv.), and Cs₂CO₃ (2.0 equiv.) were added and dissolved in acetone. The mixture was ultrasonically reacted at 40°C for 4 hours. TLC monitoring showed no remaining starting material. Saturated ammonium chloride solution and EtOAc were added for dilution. The organic phase was washed with saturated brine and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by silica gel column chromatography, intermediate K2 was obtained in 83% yield.

[0394] Intermediate K2 and A3 were synthesized via the Suzuki reaction according to the second step of Route A to obtain intermediate K3, with a yield of 70-95%; intermediate K3 was synthesized via catalytic hydrogenation according to the fifth step of Route I to obtain final product K4 with a yield of 56%.

[0395] Example 34 Synthesis of compounds Z1, Z2, Z3, and Z4

[0396] Route R:

[0397]

[0398] Following the third step of the reaction in route N, intermediate N2 (1.0 equiv.) is oxidized by DDQ to obtain intermediate S1.

[0399] To a flask dried in an oven and purged with argon, intermediate S1 (1.0 equiv.), NaI (1.0 equiv.), CuI (5.0 equiv.), dioxane (0.1 M), and N1,N1-dimethylethane-1,2-diamine (0.1 equiv.) were added sequentially. The reaction system was placed in an oil bath at 100 °C and stirred for 24 h. LC-MS analysis showed no residual starting material. The mixture was extracted with DCM and water, and the combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by preparative HPLC (MeCN / H2O) yielded intermediate S2.

[0400] In an oven-dried and argon-purged flask, anhydrous THF and anhydrous DMA were thoroughly mixed to prepare a 30% DMA / THF solution. Then, in an oven-dried and argon-purged two-necked flask, NiBr2·diglyme (0.1 equiv) and (R)-4-heptylBiOX ((R)-L1, 0.11 equiv) or ((S)-4-heptylBiOX ((S)-L1, 0.11 equiv)) solid were added, followed by argon purging. Then, 4 mL of the 30% DMA / THF solution was added via syringe, followed by another argon purging. The mixture was then heated to 60°C and stirred for approximately 30 min to form a uniform purple solution, which was then cooled to room temperature. In an oven-dried and argon-purged Schlenk reaction tube, manganese powder (1.5 equiv), 3-chloropiperidine-2,6-dione (1.0 equiv), and S2 (1.5 equiv) solid were added, followed by argon purging. Cooled NiBr2 / L1 mixed solution was added via syringe, followed by stirring for 10 min. Trimethylsilane chloride (TMSCl, 0.8 equiv) was then added via syringe. The entire process was carried out under argon protection and stirred at 1200 rpm at room temperature for 4 hours. LC-MS analysis showed no residual feedstock. The mixture was extracted with DCM / water, and the combined organic phases were washed with saturated sodium chloride solution and dried over anhydrous sodium sulfate. After filtration, solvent removal under reduced pressure, and purification by preparative HPLC (MeCN / H2O) yielded S3 and R3.

[0401] Following the fourth step of Route H, intermediates S3 and R3 (1.0 equiv.) were de-Boc protected with HCl to obtain intermediates Z1 and Z2.

[0402] Following the synthetic routes of intermediates Z1 and Z2 in route Z, intermediate G5 was synthesized into intermediates Z3 and Z4 through halogen exchange, chiral construction, and deprotection reaction.

[0403] Example 35 Synthesis of Protein Degrading Agent D1

[0404]

[0405] In an oven-dried reaction flask, intermediates C9 (1.05 equiv.) and N7 (1.0 equiv.) were added and dissolved in DMF. The reaction was carried out at 45°C for 2 h, then cooled to room temperature, and NaBH(OAc)3 was added. The reaction was carried out at room temperature for 4 h. LC-MS showed no residual raw material. The mixture was filtered through a filter membrane and purified by preparative HPLC (MeCN / H2O) to obtain compound D1 with a yield of 51%.

[0406] Example 36 Synthesis of protein degrading agent D2

[0407]

[0408] Following the synthetic method of final product D1, compound D2 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate G8 (1.2 equiv.) with a yield of 45%.

[0409] Example 37 Synthesis of protein degrading agent D3

[0410]

[0411] Following the synthetic method for the final product D1, compound D3 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate Z3 (1.2 equiv.) with a yield of 35%.

[0412] Example 38 Synthesis of protein degrading agent D4

[0413]

[0414] Following the synthetic method of the final product D1, compound D4 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate Z4 (1.2 equiv.) with a yield of 55%.

[0415] Example 39 Synthesis of protein degrading agent D5

[0416]

[0417] Following the synthetic method of the final product D1, compound D5 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate H27 (1.2 equiv.) with a yield of 42%.

[0418] Example 40 Synthesis of protein degrading agent D6

[0419]

[0420] Following the synthesis method of the final product D1, the target final product D6 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate N6 (1.2 equiv.) with a yield of 46%.

[0421] Example 41 Synthesis of protein degrading agent D7

[0422]

[0423] Following the synthetic method of final product D1, the target final product D7 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate Z2 (1.2 equiv.) with a yield of 36%.

[0424] Example 42 Synthesis of protein degrading agent D8

[0425]

[0426] Following the synthetic method of final product D1, the target final product D8 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate Z2 (1.2 equiv.) with a yield of 51%.

[0427] Example 43 Synthesis of protein degrading agent D9

[0428]

[0429] Following the synthetic method for final product D1, the target final product D9 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate Z2 (1.2 equiv.) with a yield of 43%.

[0430] Example 44 Synthesis of protein degrading agent D10

[0431]

[0432] Following the synthetic method of final product D1, the target final product D10 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate Z2, with a yield of 38%.

[0433] Example 45 Synthesis of protein degrading agent D11

[0434]

[0435] In an oven-dried reaction flask, intermediate K4 (1.0 equiv.) was added and dissolved in DCM. HATU (1.5 equiv.) and DIPEA (2.0 equiv.) were then added, and the mixture was stirred at room temperature for 2 hours. Tert-butyl piperazine-1-carboxylate was then added, and the reaction continued for 4 hours. LC-MS analysis showed no starting material remaining. Water was then added, and the mixture was diluted with DCM. The organic phase was washed with saturated brine and dried over anhydrous sodium sulfate. After filtration and solvent removal under reduced pressure, intermediate K5 was purified by silica gel column chromatography.

[0436] Following the fourth step of Route H, intermediate K5 is used to obtain intermediate K6.

[0437] Following the synthetic method for the final product D1, compound D11 was obtained by reductive amination of intermediate K6 (1.0 equiv.) and intermediate C9 (1.2 equiv.) with a yield of 41%.

[0438] Example 46 Synthesis of protein degrading agent D12

[0439]

[0440] Following the synthetic method of the final product D1, compound D12 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate H19 (1.2 equiv.) with a yield of 40%.

[0441] Example 47 Synthesis of protein degrading agent D13

[0442]

[0443] Following the synthetic method for the final product D1, the target final product D13 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate N13 (1.2 equiv.) with a yield of 30%.

[0444] Example 48 Synthesis of protein degrading agent D14

[0445]

[0446] Following the synthetic method of the final product D1, compound D14 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate G9 (1.2 equiv.) with a yield of 46%.

[0447] Example 49 Synthesis of Protein Degrading Agent D15

[0448]

[0449] Following the synthesis method of the final product D1, the target final product D15 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate N10 (1.2 equiv.) with a yield of 36%.

[0450] Example 50 Synthesis of protein degrading agent D16

[0451]

[0452] Following the synthetic method of the final product D1, the target final product D16 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate N11 (1.2 equiv.) with a yield of 42%.

[0453] Example 51 Synthesis of protein degrader D17

[0454]

[0455] Following the synthetic method of the final product D1, compound D17 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate H28 (1.2 equiv.) with a yield of 55%.

[0456] Example 52 Synthesis of protein degrader D18

[0457]

[0458] Following the synthetic method of the final product D1, compound D18 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate G17 (1.2 equiv.) with a yield of 41%.

[0459] Example 53 Synthesis of protein degrader D19

[0460]

[0461] In a reaction flask dried in an oven, intermediate K4 (1.0 equiv.) was added and dissolved in DMF. HATU (1.5 equiv.) and DIPEA (2.0 equiv.) were then added. The mixture was stirred at room temperature for 2 hours. Then, intermediate C11 (1.0 equiv.) was added, and the reaction was continued for 4 hours. LC-MS showed no residual starting material. The mixture was filtered through a filter membrane and purified by preparative HPLC (MeCN / H2O) to obtain compound D19, with a yield of 39%.

[0462] Example 54 Synthesis of protein degrading agent D20

[0463]

[0464] Following the synthesis method of the final product D19, the target final product D20 was obtained by condensation of intermediate C13 (1.0 equiv.) and intermediate K4 (1.0 equiv.) with a yield of 51%.

[0465] Example 55 Synthesis of Protein Degrading Agent D21

[0466]

[0467] Following the synthetic method of final product D1, the target final product D21 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate N14 (1.0 equiv.) with a yield of 36%.

[0468] Example 56 Synthesis of protein degrader D22

[0469]

[0470] Following the synthetic method of final product D1, the target final product D22 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate N15 (1.2 equiv.) with a yield of 40%.

[0471] Example 57 Synthesis of protein degrading agent D23

[0472]

[0473] Following the synthetic method for final product D1, the target final product D23 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate N8 (1.2 equiv.) with a yield of 31%.

[0474] Example 58 Synthesis of protein degrading agent D24

[0475]

[0476] Following the synthetic method of final product D1, the target final product D24 was obtained by reductive amination of intermediate P10 (1.0 equiv.) and intermediate G8 (1.2 equiv.) with a yield of 25%.

[0477] Example 59 Synthesis of protein degrading agent D25

[0478]

[0479] Following the synthetic method of the final product D1, compound D25 was obtained by reductive amination of intermediate F7 (1.0 equiv.) and intermediate G10 (1.2 equiv.) with a yield of 34%.

[0480] Example 60 Synthesis of protein degrader D26

[0481]

[0482] Following the synthetic method of reference end product D1, compound D26 was obtained by reductive amination of intermediate F7 (1.0 equiv.) and intermediate G12 (1.2 equiv.) in 42% yield.

[0483] Example 61 Synthesis of protein degrader D27

[0484]

[0485] Following the synthetic method of the final product D1, compound D27 was obtained by reductive amination of intermediate F7 (1.0 equiv.) and intermediate G12 (1.2 equiv.) with a yield of 49%.

[0486] Example 62 Synthesis of protein degrader D28

[0487]

[0488] Following the synthetic route for final product D1, intermediate I2 was obtained by reductive amination of intermediate N7 (1.0 equiv.) and intermediate I1 (1.2 equiv.). Intermediate I2 was then synthesized to intermediate I3 via step four of Route H. Following the synthetic route for final product D1, intermediate I3 (1.0 equiv.) was synthesized to final product D28 via reductive amination of E12 (1.2 equiv.), with a yield of 40%.

[0489] Example 63 Synthesis of protein degrader D29

[0490]

[0491] Following the synthetic method of final product D1, the target final product D29 was obtained by reductive amination of intermediate E12 (1.0 equiv.) and intermediate H21 (1.2 equiv.) with a yield of 39%.

[0492] Example 64 Synthesis of protein degrading agent D30

[0493]

[0494] Following the synthetic method for final product D1, intermediate I4 was obtained by reductive amination of intermediate G8 (1.0 equiv.) and intermediate I1 (1.2 equiv.). Intermediate I4 was then synthesized to intermediate I5 according to step four of Route H. Following the synthetic method for final product D1, intermediate I5 (1.0 equiv.) was synthesized to final product D30 via reductive amination with E12 (1.2 equiv.), with a yield of 50%.

[0495] Example 65 Synthesis of protein degrader D31

[0496]

[0497] Following the synthetic method of final product D1, the target final product D31 was obtained by reductive amination of intermediate E12 (1.0 equiv.) and intermediate H11 (1.2 equiv.) with a yield of 55%.

[0498] Example 66 Synthesis of protein degrading agent D32

[0499]

[0500] Following the synthetic method of final product D1, the target final product D32 was obtained by reductive amination of intermediate E12 (1.0 equiv.) and intermediate H13 (1.2 equiv.) with a yield of 50%.

[0501] Example 67 Synthesis of protein degrading agent D33

[0502]

[0503] Following the synthetic method of final product D1, the target final product D33 was obtained by reductive amination of intermediate E12 (1.0 equiv.) and intermediate H12 (1.2 equiv.) with a yield of 48%.

[0504] Example 68 Synthesis of protein degrading agent D34

[0505]

[0506] Following the synthetic method of final product D1, the target final product D34 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate J6 (1.2 equiv.) with a yield of 52%.

[0507] Example 69 Synthesis of protein degrading agent D35

[0508]

[0509] Following the synthetic method of final product D1, the target final product D35 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate J7 (1.2 equiv.) with a yield of 49%.

[0510] Example 70 Synthesis of protein degrading agent D36

[0511]

[0512] Following the synthetic method of final product D1, the target final product D36 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate J8 (1.2 equiv.) with a yield of 45%.

[0513] Example 71 Synthesis of protein degrading agent D37

[0514]

[0515] Following the synthetic method of final product D1, the target final product D37 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate J9 (1.2 equiv.) with a yield of 38%.

[0516] Example 72 Synthesis of protein degrader D38

[0517]

[0518] Following the synthetic method of final product D1, the target final product D38 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate J10 (1.2 equiv.) with a yield of 40%.

[0519] Example 73 Synthesis of protein degrader D39

[0520]

[0521] Following the synthesis method of the final product D1, the target final product D39 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate J11 (1.2 equiv.) with a yield of 47%.

[0522] Example 74 Synthesis of protein degrading agent D40

[0523]

[0524] Following the synthesis method of the final product D1, the target final product D40 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate J12 (1.2 equiv.) with a yield of 50%.

[0525] Example 75 Synthesis of Protein Degrading Agent D41

[0526]

[0527] Following the synthetic method of the final product D1, the target final product D41 was obtained by reductive amination of intermediate C15 (1.0 equiv.) and intermediate N6 (1.2 equiv.) with a yield of 43%.

[0528] Example 76 Synthesis of protein degrading agent D42

[0529]

[0530] Following the synthetic method of final product D1, the target final product D42 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate N16 (1.2 equiv.) with a yield of 38%.

[0531] Example 77 Synthesis of protein degrading agent D43

[0532]

[0533] Following the synthetic method of final product D1, the target final product D43 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate G11 (1.2 equiv.) with a yield of 35%.

[0534] Example 78 Synthesis of protein degrading agent D44

[0535]

[0536] Following the synthetic method of final product D1, the target final product D44 was obtained by reductive amination of intermediate C9 (1.0 equiv.) and intermediate G10 (1.2 equiv.) with a yield of 47%.

[0537] Example 79 Synthesis of protein degrading agent D45

[0538]

[0539] Intermediate G1 (200 mg, 0.49 mmol) was dissolved in anhydrous DMF (6 mL), and NCS (70 mg, 0.49 mmol) was added at room temperature. The mixture was stirred overnight at room temperature. The reaction of the starting material was monitored by TLC until complete. The aqueous phase was extracted with ethyl acetate, and the organic phases were combined. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The concentrate was then subjected to rapid column chromatography (V petroleum ether / V ethyl acetate = 1 / 1) to obtain O1.

[0540] Intermediate O1 was synthesized to obtain intermediate O2, following the fourth step of Route H.

[0541] Following the synthetic method of the final product D1, the target final product D45 was obtained by reductive amination of intermediate O2 (1.0 equiv.) and intermediate C9 (1.2 equiv.) with a yield of 47%.

[0542] Example 80 Synthesis of protein degrading agent D46

[0543]

[0544] Intermediate G4 and A3 were synthesized via the Suzuki reaction, following the second step of Route A, to obtain intermediate O3; intermediate O3 was synthesized via catalytic hydrogenation, following the fifth step of Route I, to obtain intermediate O4; intermediate O4 was synthesized via the fourth step of Route H to obtain intermediate O5.

[0545] Following the synthetic method of final product D1, the target final product D46 was obtained by reductive amination of intermediate O5 (1.0 equiv.) and intermediate C9 (1.2 equiv.) with a yield of 43%.

[0546] Example 81 Synthesis of protein degrader D47

[0547]

[0548] Following the first step of Route P, compounds O6 and H14 are reductively aminationd to obtain intermediate O7; intermediate O7 and A3 are synthesized via a Suzuki reaction following the second step of Route A to obtain intermediate O8; following the fifth step of Route I, intermediate O8 is synthesized via a catalytic hydrogenation reaction to obtain intermediate O9; intermediate O9 is synthesized via the fourth step of Route H to obtain intermediate O10.

[0549] Following the synthetic method for final product D1, the target final product D47 was obtained by reductive amination of intermediate O10 (1.0 equiv.) and intermediate C9 (1.2 equiv.) with a yield of 39%.

[0550] Example 82 Synthesis of protein degrader D48

[0551]

[0552] 4-Bromoindigo (1 g, 4.424 mmol) and tert-butyl 4-bromopiperidine-1-carboxylate (3.505 g, 13.272 mmol) were dissolved in DMF (22 ml), and Cs2CO3 (2.883 g, 8.848 mmol) was added. The mixture was stirred at 80 °C for 16 h. TLC showed that half of the reaction starter remained. The mixture was quenched with water, extracted three times with EA, and the organic phases were combined. The mixture was washed once with saturated brine, dried over anhydrous sodium sulfate, and the solvent was removed under reduced pressure to obtain the crude product. The crude product was then separated by column chromatography (PE:EA = 3:1) to obtain intermediate O13.

[0553] O13 (375.6 mg, 0.918 mmol) was placed in a reaction flask, and hydrazine hydrate (8 ml) was added. The mixture was heated to reflux at 120 °C for 1 day. TLC was used to detect the reaction. After the reaction was complete, the solvent was removed under reduced pressure to obtain the crude product, which was then separated by column chromatography (PE:EA = 3:1) to obtain the target compound O14.

[0554] Intermediate O14 and A3 were synthesized via the Suzuki reaction according to the second step of Route A to obtain intermediate O15; intermediate O15 was synthesized via catalytic hydrogenation according to the fifth step of Route I to obtain intermediate O16; intermediate O16 was synthesized via the fourth step of Route H to obtain intermediate O17.

[0555] Following the synthetic method for final product D1, the target final product D48 was obtained by reductive amination of intermediate O17 (1.0 equiv.) and intermediate C9 (1.2 equiv.) with a yield of 31%.

[0556] Example 83 Synthesis of protein degrader D49

[0557]

[0558] Following the synthetic method for final product D1, the target final product D49 was obtained by reductive amination of Compound 44 (1.0 equiv.) and intermediate C9 (1.2 equiv.) with a yield of 36%.

[0559] Example 84: Verification of the binding activity of compounds 1-59 to the E3 ubiquitin ligase CRBN

[0560] Verification of the binding affinity activity of the compound for CRBN:

[0561] The binding affinity of the test compound to CRBN protein was detected using the HTRF Cereblon Binding Kit and the 500Assay kit (64BDCRBNPEG; CISBIO). The specific method is as follows:

[0562] 1. Prepare a 40 mM stock solution of the test compound and lenalidomide using DMSO. Serially dilute the compound with diluent#9 (1X) solution to obtain eight different working solutions (800 μM, 160 μM, 32 μM, 6.4 μM, 1.28 μM, 0.256 μM, 0.051 μM); dilute Thalidomide-Red and GST Eu antibodies 50-fold with PROTAC binding buffer 1 (1X), and dilute human CereblonWT GST-tagged protein 50-fold with PROTAC binding buffer 1 (1X).

[0563] 2. First, add 5 μL of the compound and human CereblonWT GST-tagged protein solution to each well. Then, add 10 μL of the homogenized Thalidomide-Red reagent and GST Eu antibody working solution to each well.

[0564] 3. Seal the plate and incubate at room temperature for 3 hours, then remove the seal and perform detection using a Spark microplate reader (Tecan).

[0565] 4. Calculate the ratio of acceptor to donor emission signals for each individual well (Ratio = 665nm signal value / 620nm signal value * 10⁴), and perform IC analysis using GraphPad Prism 8. 50 Value calculation.

[0566] Table 3. Binding activities of Compound 1–59 to the E3 ubiquitin ligase CRBN

[0567]

[0568]

[0569] Using GraphPad Prism, calculate IC based on compound concentration, ratio, and coefficient of variation. 50 value.

[0570] As shown in Table 1, all 59 compounds tested exhibited superior affinity for CRBN. Compared to the positive control Lenalidomide (1–3 μM), more than half of the compounds showed significantly higher IC50 values. 50 A value lower than positive indicates that these compounds have a higher affinity for CRBN, with Compound49 exhibiting the highest affinity for CRBN, and its IC50 value being [value missing]. 50 With a value of only 0.004, the affinity was 500 times higher than that of the positive control in the same batch of tests.

[0571] Example 85 Evaluation of the reducing effects of protein degrading compounds on the expression levels of AR, ER, BTK, and IRAK4

[0572] Western blotting was used to determine the dendritic ratio (DC) of the compound against the target protein in cells. 50 The specific methods are as follows: AR and ER targets were selected from human prostate cancer VCaP and human breast cancer MCF-7 cells, respectively, both cultured in DMEM high-glucose medium containing 10% fetal bovine serum (10099141C, Gibco); BTK and IRAK4 targets were selected from human mantle cell lymphoma JeKo-1 and human monocytic leukemia THP-1 cells, respectively, both cultured in RPMI-1640 medium containing 10% fetal bovine serum (10099141C, Gibco), and incubated at 37℃, 5% CO2, and 95% humidity. Cells were cultured at 3*10-1... 5Cells were seeded in 12-well plates at 1 mL per well. The next day, cells were treated with different concentrations of the compound for 6 hours and then collected. Protein lysis buffer was added to the cells, and the plates were centrifuged and the supernatant was collected. 5×SDS loading buffer was added, and the plates were boiled at 95°C for 10 min and stored. Protein electrophoresis was performed using SDS-PAGE, followed by transferring the proteins onto a PVDF membrane at a constant current of 400 mA for 30 min. Primary antibody GSPT1 (ab126090, abcam) and α-tubulin antibody (S0B0800, STARTER) were added to the membrane according to the antibody manufacturer's instructions and incubated overnight. The membrane was washed with TBST and imaged using ECL chemiluminescence buffer on a Tianneng imaging system. Band grayscale was analyzed using ImageJ software, with α-tubulin used as an internal control for calibration. DC analysis was performed using a GraphPad Prism 8. 50 Value calculation.

[0573] Table 4 summarizes the degradation activities of the protein degrader CompoundD1-33 on AR, ER, BTK, and IRAK4 proteins. Among them, DC... 50 This represents the concentration of the protein degrading agent required to induce 50% degradation of AR, ER, BTK, and IRAK4 proteins. D represents this concentration. max The maximum extent to which the representative protein degrader can induce the degradation of AR, ER, BTK, and IRAK4 proteins is relative to the total amount of AR, ER, BTK, and IRAK4 proteins.

[0574] Table 4. Specific evaluation of the effect of protein degrader Compound D1-33 on reducing the expression levels of different target proteins.

[0575]

[0576]

[0577] Table 4 shows that PROTAC degraders Compound D2, D6, and D9 have significant degradation activity against the target protein AR, while DC... 50 The concentrations were 1.31, 1.42, and 7.33 nM, respectively; the PROTAC degrader Compound D25 exhibited significant degradation activity against the target protein BTK. max The degradation rate reached 100%; the PROTAC degrader CompoundD29 exhibited significant degradation activity against the target protein IRAK4, indicating that it could induce target protein degradation at low concentrations, demonstrating high degradation efficiency. Meanwhile, their D... max All are at level A (D) max The percentage of these compounds exceeding 75% indicates that they can maximally reduce the expression level of the target protein, with a degradation rate exceeding 75%.

[0578] Table 5. Specific evaluation of the effect of protein degrader Compound D34-49 on reducing AR protein expression.

[0579]

[0580]

[0581] Table 5 shows that the PROTAC degrader Compound D35 exhibits significant degradation activity against the target protein AR. Specifically, the compound achieves 86% degradation at 1 nM, demonstrating high degradation efficiency. Furthermore, the compound degrades 98% of AR protein at 100 nM, indicating that it can maximally reduce AR protein expression. In summary, the compounds of this invention demonstrate excellent activity in target degradation, suggesting that the corresponding CRBN-binding ligand structures have high potential application value.

Claims

1. A compound, or its stereoisomer, or its pharmaceutically acceptable salt, or its deuterated compound, or its tautomer, or its polymorph, or its solvate, or its N-oxide, or its isotopically labeled compound, or its metabolite, or its prodrug, capable of binding CRBN protein; wherein the compound comprises a fused ring skeleton consisting of two ring systems, A and B, and the G group is connected to any one of the 4, 5, or 6 positions of the A ring via a carbon-carbon single bond, and the structure of the compound capable of binding CRBN protein is shown in Formula (I); In Equation I, the G substructure includes the following G 1 G 2 G 3 Three types, namely R configuration, S configuration, and racemic configuration: In Formula I, ring A is a benzene ring, selected from any of the following: in, R1 is selected from any one of H, C1-C3 alkyl, C1-C3 alkoxy or halogen; R2 is selected from any one of H, C1-C3 alkyl, C1-C3 alkoxy or halogen; R3 is selected from any one of H, C1-C3 alkyl, C1-C3 alkoxy or halogen; R4 is selected from any one of H, C1-C3 alkyl, C1-C3 alkoxy or halogen; In Formula I, ring B... Representing a covalent single bond or a covalent double bond, X1, X2 and X3 of the B ring are selected from either C or N; The specific structure of the B ring is preferably selected from any of the following structures: Wherein, P1, P2, and P3 are selected from H, halogens, or any of the following structures:

2. The compound of claim 1, wherein the pharmaceutically acceptable salt is a parent compound prepared by addition of a non-toxic acid or its base.

3. The compound of claim 1, in some embodiments of the present invention, wherein the specific structure of the compound capable of binding to the CRBN protein is selected from any of the following:

4. A PROTAC degrading agent, wherein the structure of the PROTAC degrading agent is as follows: in, The CRBN protein ligand is the compound described in claim 1; The structure of the PROTAC degrading agent is shown in formula (II): In Formula II, the occupying group The structure is none, or it can be selected from any of the following structures: When the occupying group When the structure is empty, the Linker's L 1 The CRBN protein ligand's B structure is linked to any one of the X1, X2, or X3 sites by a CN, NC, or CO single bond; When the occupying group When selected from the above structures, the occupying group With the Linker L 1 Substructures are linked by CN, NC, or CO single bonds; In Formula II, the substructure L of the linker 1 L 3 and L 5 Each of the following structures can be selected: Among them, R a Selected from any one of H, alkyl, SO2-alkyl, or SO2-aryl; In Formula II, the substructure L of the linker 2 and L 4 Each of the following structures can be selected: In Formula II, the target protein ligand is a ligand of a target protein that causes the related disease, and the target protein ligand is selected from any one of the following structures:

5. The PROTAC degrading agent as described in claim 4, in some embodiments of the present invention, the specific structure of the PROTAC degrading agent is selected from any of the following:

6. A pharmaceutical composition comprising a compound of formula (I); and may further comprise an enantiomer, diastereomer, stereoisomer, or pharmaceutically acceptable salt of the compound of formula (I) and a pharmaceutically acceptable diluent or carrier.

7. A pharmaceutical composition for targeting protein degradation, the pharmaceutical composition comprising a PROTAC degrader of formula (II); the pharmaceutical composition targeting targets such as AR, ER, BTK and IRAK4.

8. The use of a compound as described in claim 1 or a PROTAC degrader as described in claim 4, or a stereoisomer thereof, a pharmaceutically acceptable salt, a deuterated compound, a tautomer, a polymorph, a solvate, an N-oxide, an isotope-labeled compound, a metabolite, or a prodrug in the preparation of a medicament for treating diseases related to proteins such as AR, ER, BTK, and IRAK4.

9. The application as described in claim 8, wherein the drug achieves the effect of treating the disease or relieving symptoms by limiting or inhibiting the expression of AR, ER, BTK, and IRAK4.

10. The application as described in claim 8, wherein the diseases related to the AR, ER, BTK, IRAK4, and other proteins include, but are not limited to, prostate cancer, androgenetic alopecia, polycystic ovary syndrome, breast cancer, endometrial cancer, osteoporosis, B-cell lymphoma, lymphocytic leukemia, autoimmune diseases, monocytic leukemia, acute myeloid leukemia, multiple myeloma, non-small cell lung cancer, chronic myeloid leukemia, gout, inflammatory bowel disease, acne, synovial sarcoma, solid tumors, small cell lung cancer, cervical cancer, neuroblastoma, hepatocellular carcinoma, colorectal cancer, pancreatic cancer, malignant rhabdomyosarcoma, and oral squamous cell carcinoma.

Images