Targeted FAK degradation compound and application

Abstract

The invention discloses a compound targeting FAK degradation and application, the compound has a structure as shown in formula (I) and can be used for preparing drugs for treating, preventing and relieving diseases caused by excessive expression of FAK protein. The PROTAC compound capable of degrading the FAK protein in the targeted manner has the capability of remarkably degrading FAK in MIA-PACA-2 cells under different concentrations, and has certain proliferation inhibition capability on the MIA-PACA-2 cells when being combined with a KRAS inhibitor. The compound can be developed as a potential anti-solid tumor drug, and has a wide application prospect.

Description

Technical Field

[0001] This invention relates to the field of chemical synthesis, and more specifically to a compound that targets FAK degradation and its applications. Background Technology

[0002] Focal adhesion kinase (FAK), also known as PTK2 (protein tyrosine kinase 2), is a class of non-receptor protein tyrosine kinases that are widely expressed in various tissues of the human body. It is mainly regulated by integrin signaling and controls basic cellular processes such as cell adhesion, migration, proliferation, and survival through kinase-dependent and kinase-independent functions.

[0003] FAK is composed of a FERM domain, a kinase domain, a proline-rich region, and a FAT domain. These domains make FAK a core scaffold of the cell signaling complex, enabling it to bind to multiple proteins simultaneously, forming a highly dynamic signaling complex that precisely integrates extracellular matrix (ECM) signals into intracellular signaling pathways. In this process, after integrins bind to the ECM, they recruit FAK to focal adhesion and trigger trans-autophosphorylation at the Y397 site. Once phosphorylated, FAK acts as a molecular scaffold and recruits Src family kinases (SFKs) to bind to FAK, activating their catalytic function. This leads to further phosphorylation of multiple tyrosine residues in FAK, enhancing its function as a scaffold protein and activating multiple downstream signaling pathways, such as the PI3K-AKT pathway, the Ras-MAPK pathway, and the STAT1 pathway. The activation of these signaling pathways plays a crucial role in the growth, proliferation, metastasis, and invasion of tumor cells. Furthermore, FAK overexpression is also associated with chemotherapy resistance and radiotherapy resistance in cancer, and inhibiting the FAK signaling pathway can reverse tumor drug resistance and enhance the efficacy of immunotherapy. Therefore, inhibiting or degrading FAK is considered a novel strategy in cancer treatment.

[0004] Although several FAK inhibitors have shown promising results in preclinical or clinical trials, particularly demonstrating potential therapeutic value in refractory tumors such as platinum-resistant ovarian and pancreatic cancers, it is important to note that no small-molecule inhibitors that directly target FAK have yet received clinical approval globally. Furthermore, FAK small-molecule inhibitors primarily target the kinase active domain of FAK without disrupting its scaffold function, only partially blocking its signaling function. This limitation of selective inhibition often leads to drug resistance in tumor cells through activation of non-kinase domain signaling pathways. This significantly restricts the clinical application of FAK-targeting strategies relying solely on kinase inhibition.

[0005] Proteolysis-targeting chimera (PROTAC) technology has attracted widespread attention as an innovative drug development strategy. PROTAC technology utilizes PROTAC molecules to link a target protein with an E3 ubiquitin ligase, forming a ternary complex of target protein-PROTAC-E3 ubiquitin ligase, which ubiquitinates the target protein. The ubiquitinated protein can then be specifically recognized and degraded by the proteasome within the cell. Compared to traditional inhibitors that only inhibit single signaling pathways of certain multifunctional proteins, PROTAC molecules can induce target protein degradation, affecting all functions of the protein and thus reducing feedback mechanisms, which can, to some extent, overcome drug resistance and reduce the likelihood of its development. Furthermore, PROTACs offer potential advantages such as low dosage, low toxicity, the ability to overcome drug resistance, improved selectivity, and targeting of undrugifiable proteins. These breakthrough advantages make PROTAC technology one of the most clinically promising protein degradation therapy strategies. Therefore, it is necessary to develop a PROTAC-like compound that targets and degrades the FAK protein. Summary of the Invention

[0006] The purpose of this invention is to provide a compound targeting FAK degradation and its applications. The described PROTAC-like compound, which targets and degrades FAK protein, exhibits significant FAK degradation ability at different concentrations in MIA-PACA-2 cells, and when combined with a KRAS inhibitor, it shows a certain inhibitory effect on the proliferation of MIA-PACA-2 cells. It can be developed as a potential anti-solid tumor drug and has broad application prospects.

[0007] To achieve the above objectives, the present invention provides a PROTAC compound of Formula I that degrades FAK protein:

[0008]

[0009] The Linker is a connecting chain that links to the 4-position of the right-hand isoindole-1,3-dione structure, and its structure is as follows:

[0010] ;

[0011] R1 is a group of -CF3 or -Cl.

[0012] Furthermore, the PROTAC compound that degrades FAK protein, or its stereoisomers, tautomers, deuterated derivatives, solvates, prodrugs, or metabolites, are compounds with the following structures:

[0013] Compound 1: ;

[0014] Compound 2: ;

[0015] Compound 3: ;

[0016] Compound 4: ;

[0017] Compound 5: ;

[0018] Compound 6: ;

[0019] Compound 7: ;

[0020] Compound 8: ;

[0021] Compound 9: ;

[0022] Compound 10: ;

[0023] Compound 11: .

[0024] The present invention also provides a method for preparing the PROTAC compound that degrades FAK protein, comprising the following steps:

[0025] The PROTAC compound that targets and degrades FAK protein is a compound of general formula (I), where the linker is a piperidine derivative of different lengths and sizes, and the E3 ligand is a 4-substituted amine derivative.

[0026] The present invention also provides a method for preparing the PROTAC compound that targets and degrades FAK protein, the route of which is as follows:

[0027] Step 1: Synthesize POI target molecule intermediate 8 targeting FAK via route 1 or route 2:

[0028]

[0029] Step 2: Synthesis of important intermediate 11a-j

[0030]

[0031] Step 3: The POI target molecule intermediate 8 targeting FAK obtained in Step 1 and the key intermediate 11-a~j obtained in Step 2 are reductively amination in DMF at room temperature in the presence of NaBH3CN and HOAc to obtain the target product.

[0032] R1 is a group of -CF3 or -Cl.

[0033] In a second aspect, the present invention also provides a pharmaceutical composition comprising the compound as described above, or a stereoisomer, tautomer, solvate, pharmaceutically acceptable salt, metabolite, isotope derivative, N-oxide, or prodrug, and a pharmaceutically acceptable carrier, diluent, or excipient.

[0034] Thirdly, the present invention also provides the use of the compounds described above, or their stereoisomers, tautomers, solvates, pharmaceutically acceptable salts, metabolites, isotope derivatives, N-oxides or prodrugs, or pharmaceutical compositions, in medicaments for the prevention and / or treatment of diseases associated with FAK activity. These FAK activity-related diseases include cancer or immune diseases; the cancers preferably include cholangiocarcinoma, diffuse large B-cell lymphoma, clear cell renal cell carcinoma, low-grade glioma, sarcoma, thymoma, mesothelioma, meningioma, gastric adenocarcinoma, melanoma, lung cancer, small cell lung cancer, non-small cell lung cancer, lung adenocarcinoma, hepatocellular carcinoma, gastric cancer, intestinal cancer, pancreatic cancer, glioblastoma, ovarian cancer, liver cancer, bladder cancer, hepatocellular carcinoma, breast cancer, metastatic breast cancer, colon cancer, rectal cancer, colorectal cancer, kidney cancer, head and neck cancer, pheochromocytoma, and paraganglioma.

[0035] In another aspect, in the above-described uses or methods, the compounds described above, or their stereoisomers, tautomers, solvates, pharmaceutically acceptable salts, metabolites, isotope derivatives, N-oxides or prodrugs, or pharmaceutical compositions, may be used alone or in combination with other types of pharmaceutical preparations and / or treatment methods. Particularly in the preparation of drugs for treating pancreatic cancer, they may be used in combination with KRAS G12C inhibitors.

[0036] It is particularly noteworthy that, in this article, when referring to a "compound" having a specific structural formula, it generally also includes its stereoisomers, diastereomers, enantiomers, racemic mixtures, and isotopic derivatives.

[0037] The "pharmaceutically acceptable salts" described in this invention refer to pharmaceutically acceptable addition salts of acids and bases, or their solvates. Such pharmaceutically acceptable salts include salts of the following acids: hydrochloric acid, phosphoric acid, hydrobromic acid, sulfuric acid, sulfurous acid, formic acid, toluenesulfonic acid, methanesulfonic acid, nitric acid, benzoic acid, citric acid, tartaric acid, maleic acid, hydroiodic acid, alkanonic acids (such as acetic acid, HOOC-(CH2)n-COOH (where n is 0-4)), etc., or mixed salts thereof. Salts of bases include: sodium salts, potassium salts, calcium salts, ammonium salts, etc. Many non-toxic pharmaceutically acceptable addition salts are known to those skilled in the art.

[0038] The pharmaceutically acceptable salts of the present invention can be prepared by conventional methods, for example by dissolving the compounds of the present invention in a water-miscible organic solvent (e.g., acetone, methanol, ethanol, and acetonitrile), adding an excess of an aqueous solution of an organic or inorganic acid to precipitate the salt from the resulting mixture, removing the solvent and the remaining free acid, and then separating the precipitated salt.

[0039] Compared with the prior art, the present invention has the following advantages:

[0040] Based on the rapidly developing PROTAC (Proteolysis-targeting chimaeras) technology, this invention addresses the problems of drug resistance and insufficient activity of small molecule inhibitors through protein degradation. The PROTAC compound provided by this invention does not require long-term occupation of the active site like inhibitors; instead, it only needs a brief binding to the target protein to induce protein degradation (with best-in-class degradation effect). Since the degraded protein needs to be resynthesized to restore its function, this significantly delays the development of drug resistance. Attached Figure Description

[0041] Figure 1 To detect the effect of compounds 1–6 on FAK degradation induced by immunoblotting at five concentrations of 0.001, 0.01, 0.1, 1, and 10 µM;

[0042] Figure 2 To detect the effect of compounds 7-11 on FAK degradation induced by immunoblotting at five concentrations of 0.001, 0.01, 0.1, 1 and 10 µM;

[0043] Figure 3 To detect the effect of compound 11 on inducing FAK degradation at 0.013, 0.04, 0.12, 0.37, 1.11, 3.33 and 10 µM by immunoblotting;

[0044] Figure 4 To detect the effect of compound 11 on inducing FAK degradation at 0, 2, 4, 12, 24 and 48 °C using immunoblotting;

[0045] Figure 5 The results are experimental findings regarding the ubiquitination mechanism of compound 11.

[0046] Figure 6 The results of the in vitro antitumor activity assay of the degradation agent were obtained using the CCK8 assay.

[0047] Figure 7 The results of the in vitro antitumor activity assay of compound 11 in combination with AMG510 were obtained by CCK8 assay.

[0048] Figure 8Concentration-time curves of compounds 3, 4, and 5 administered via gavage;

[0049] Figure 9 The concentration-time curve of compound 3 after gavage administration. Detailed Implementation

[0050] The present invention will be further illustrated by way of embodiments below, but the present invention is not limited to the scope of the embodiments described herein.

[0051] Example 1: Preparation of Compound 1

[0052] Synthesis of intermediate 3:

[0053]

[0054] 2,4-Dichloro-5-trifluoromethylpyrimidine (1, 10 g, 46.087 mmol) and zinc chloride (12.56 g, 92.17 mmol) were dissolved in a 1,2-dichloroethane / tert-butanol (1:1, v / v) mixture and stirred at room temperature for 30 minutes. Then, methyl 4-amino-3-methoxybenzoate (2, 8.35 g, 46.087 mmol) and triethylamine (7.27 mL, 50.7 mmol) were added sequentially. Under nitrogen protection, the reaction mixture was heated to 110 °C and stirred continuously until the reaction was complete (monitored by TLC). After the reaction was complete, the reaction was quenched with water, and the mixture was extracted with ethyl acetate (500 mL × 3). The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give a white solid. 8.3 g, yield approximately 50%. 1 H NMR (400 MHz, CHLOROFORM-D) δ 8.63 (s, 1H), 8.54 (d, J = 8.5 Hz, 1H), 8.29 (s, 1H), 7.73 (dd, J = 8.5, 1.8 Hz, 1H), 7.57 (d, J= 1.8 Hz, 1H), 3.98 (s, 3H), 3.91 (s, 3H). 13 C NMR (101 MHz, CHLOROFORM-D) δ166.78, 160.03, 159.42, 157.36 (q, J = 5.1 Hz), 147.62, 131.69, 124.92,123.89, 123.36, 118.09, 110.84, 56.15, 52.23. ESI [M + H] + (m / z): 362.07.

[0055] Synthesis of intermediate 5:

[0056]

[0057] Intermediate 3 (6 g, 16.62 mmol) was dissolved in 1,4-dioxane, and 7-hydroxy-1-indanone (4 g, 3.7 g, 24.93 mmol) and cesium carbonate (27.07 g, 83.1 mmol) were added. The reaction mixture was heated to 80 °C and stirred continuously for 18 hours until completion. The reaction was monitored by TLC. After completion, the reaction was quenched with water. The resulting mixture was extracted with ethyl acetate (500 mL × 3), the organic phases were combined, washed with 100 mL of saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give a yellowish-white solid. 3.4 g, yield approximately 50%. 1 HNMR (400 MHz, CHLOROFORM-D) δ 8.58 (s, 1H), 8.05 (s, 1H), 7.73 (t, J = 7.8Hz, 1H), 7.53 (d, J = 7.6 Hz, 1H), 7.44 (d, J = 1.8 Hz, 1H), 7.21 (d, J =10.2 Hz, 1H), 7.16 (d, J = 7.9 Hz, 1H), 3.89 (d, J = 3.1 Hz, 6H), 3.20 – 3.13(m, 2H), 2.58 – 2.49 (m, 2H). 13 C NMR (101 MHz, CHLOROFORM-D) δ 203.02, 166.82,160.13, 157.54, 157.49, 157.19, 148.66, 147.22, 136.25, 132.50, 129.17,124.81, 124.53, 123.71, 122.61, 121.84, 121.02, 116.89, 110.57, 56.02, 52.15,36.63, 29.79, 25.82. ESI [M + H] + (m / z): 474.11.

[0058] Synthesis of intermediate 6:

[0059]

[0060] Intermediate 5 (2.8 g, 5.92 mmol) and lithium hydroxide (1.242 g, 29.6 mmol) were dissolved in a tetrahydrofuran / water (1:1, v / v) mixture. The reaction mixture was stirred continuously at room temperature for 3 hours until the reaction was complete. The reaction was monitored by TLC. After the reaction was completed, the pH was adjusted to 3-5 with 6N hydrochloric acid aqueous solution. The resulting mixture was then extracted with ethyl acetate (500 mL × 3), the organic phases were combined, washed with 100 mL of saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give a yellowish-white solid. 2.2 g, yield approximately 81%. ESI [M + H] + (m / z): 460.12.

[0061] Synthesis of intermediate 8:

[0062]

[0063] Intermediate 6 (2 g, 4.354 mmol), HATU (1.98 g, 5.224 mmol), and DIPEA (1.52 ml, 8.708 mmol) were dissolved in DMF and stirred at room temperature for 30 min. Then, 1-Boc-4-aminopiperidine (7, 1.3 mg, 6.531 mmol) was added to the reaction mixture, and the reaction was stirred continuously at room temperature for 3 h. The reaction was monitored by TLC. After the reaction was completed, water was added to quench the reaction. The resulting mixture was extracted with ethyl acetate (500 mL × 3), the organic phases were combined, washed with 100 ml of saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give a yellowish-white solid. 1.2 g, yield approximately 43%. 1H NMR (400 MHz, DMSO) δ 8.83 (s, 1H), 8.70 (s,1H), 8.20 (d, J = 7.9 Hz, 1H), 7.77 (t, J = 7.8 Hz, 1H), 7.55 (d, J = 7.6 Hz,1H), 7.39 (d, J = 1.9 Hz, 1H), 7.28 (s, 1H), 7.21 (d, J = 7.9 Hz, 1H), 7.12(d, J = 8.3 Hz, 1H), 4.02 – 3.91 (m, 3H), 3.78 (s, 3H), 3.10 (t, J = 5.8 Hz,2H), 2.83 (s, 2H), 2.47 (d, J = 5.7 Hz, 3H), 1.78 (dd, J = 13.1, 3.8 Hz, 2H), 1.45 (dd, J = 12.2, 4.3 Hz, 2H), 1.41 (s, 9H). 13 C NMR (101 MHz, DMSO) δ203.45, 166.38, 165.27, 161.26, 158.31, 157.85, 154.37, 150.14, 147.98,136.94, 130.77, 130.01, 128.95, 125.48, 120.98, 119.72, 110.32, 79.15, 56.41,47.09, 36.79, 31.95, 28.61, 25.83. ESI [M + H] + (m / z): 642.27.

[0064] Synthesis of intermediate 10a:

[0065]

[0066] The starting materials 3-hydroxyazacyclobutane (420 mg, 5.74 mmol) and 2-(2,6-dioxopiperidin-3-yl)-5-fluoroisoindole-1,3-dione (2.257 g, 8.6 mmol) were dissolved in DMF. The mixture was stirred at room temperature, and DIPEA (2.0 mL, 11.48 mmol) was added to the solution. The reaction mixture was stirred at 80 °C for 12 h. The reaction was monitored by TLC, and after completion, the reaction was quenched with water. The resulting mixture was extracted with dichloromethane (500 mL × 3), the organic phases were combined, washed with saturated brine (100 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give intermediate 2-6a as a yellow solid powder, with a yield of approximately 89%. 1 H NMR (400 MHz, DMSO-D6) δ 11.07 (s,1H), 7.62 (d, J = 8.3 Hz, 1H), 6.78 (d, J = 2.2 Hz, 1H), 6.64 (dd, J = 8.3,2.2 Hz, 1H), 5.82 (d, J = 6.4 Hz, 1H), 5.05 (dd, J = 12.9, 5.4 Hz, 1H), 4.62 (h, J = 6.5 Hz, 1H), 4.25 (t, J = 8.2 Hz, 2H), 3.74 (dd, J = 9.8, 4.6 Hz, 2H), 2.97 – 2.74 (m, 1H), 2.64 – 2.53 (m, 2H), 2.13 – 1.86 (m, 1H). 13 C NMR(101 MHz, DMSO-D6) δ 173.34, 170.63, 167.99, 167.67, 155.54, 134.32, 125.32,117.24, 114.90, 105.18, 61.65, 61.07, 49.17, 31.45, 22.67. ESI [M + H] + (m / z): 352.14.

[0067] Synthesis of intermediate 11a:

[0068]

[0069] Compound 10a (200 mg, 0.607 mmol) was dissolved in dichloromethane, and Desmartin oxidant (DMP, 386.280 mg, 0.910 mmol) was slowly added under ice bath conditions at 0°C. After the addition was complete, the reaction system was slowly raised to room temperature and stirred continuously for 2 hours. After the reaction was completed, the reaction suspension was filtered three times through a diatomaceous earth filter, and the filtrate was collected. The organic phase was concentrated under reduced pressure to obtain the crude product, which could be used for the next reaction without further purification.

[0070] Synthesis of Compound 1:

[0071]

[0072] Intermediate 8 (200.0 mg, 0.312 mmol) was dissolved in dichloromethane, and EA / HCl (2.5 mL) was added. The mixture was stirred at room temperature for 30 minutes. The solvent was then removed by evaporation under reduced pressure to obtain the crude product. This crude product was dissolved in DMF, and the pH was adjusted to neutral by adding DIPEA. Then, intermediate 11a prepared in the previous step was added, along with sodium cyanoborohydride (68 mg, 1.0 mmol). The reaction mixture was adjusted to weakly acidic with acetic acid and stirred at room temperature for 4 hours. After the reaction was complete, water was added to quench the reaction, and the reaction mixture was extracted with dichloromethane (500 mL × 3). The organic phases were combined. The organic phase was washed with saturated brine (100 mL × 5), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to obtain the final product as a yellow solid powder, with a yield of approximately 59%. 1H NMR (400 MHz, CDCl3) δ 9.16 (s, 1H), 8.56 (s, 1H), 7.92 (s, 1H), 7.67 (t, J = 7.8 Hz, 1H), 7.59 (d, J = 8.2 Hz, 1H), 7.45 (d, J = 7.6 Hz, 1H), 7.30 (s, 1H), 7.12 (d, J = 7.9 Hz, 1H), 7.00 – 6.91(m, 1H), 6.73 (d, J = 2.1 Hz, 1H), 6.49 (dd, J = 8.3, 2.1 Hz, 1H), 6.28 (s,1H), 4.88 (dd, J = 12.0, 5.4 Hz, 1H), 4.06 (q, J = 7.5 Hz, 3H), 3.89 (d, J =6.7 Hz, 2H), 3.85 (s, 3H), 3.35 (t, J = 6.2 Hz, 1H), 3.13 (t, J = 5.9 Hz,2H), 2.98 – 2.58 (m, 5H), 2.58 – 2.47 (m, 2H), 2.08 (dt, J = 21.3, 10.1 Hz, 6H), 1.58 (s, 2H). 13 C NMR (101 MHz, CDCl3) δ 201.98, 170.51, 167.86, 166.96,166.46, 165.65, 165.36, 159.10, 156.42, 156.09, 153.87, 147.49, 146.55,135.09, 133.19, 130.03, 127.97, 127.30, 124.27, 123.59, 120.76, 119.91,117.90, 116.96, 116.25, 113.18, 107.93, 104.03, 76.24, 54.93, 54.43, 53.79,48.11, 48.06, 45.77, 35.54, 30.41, 28.67, 24.71, 21.77. HRMS (ESI) (m / z):calcd for C 43 H 40 O8F3N8 [M + H] + , 853.2922; found, 853.2896.

[0073] Example 2: Preparation of Compound 2

[0074] Synthesis of intermediate 10b:

[0075]

[0076] The preparation method is the same as 10a, except that compound 3-hydroxypyrrolidine is used instead of compound 3-azacyclobutanol to obtain the product, which is a yellow solid powder with a yield of about 90%. 1 H NMR (400 MHz, DMSO) δ 11.07 (s, 1H), 7.64 (d, J = 8.6Hz, 1H), 6.84 (d, J = 38.8 Hz, 2H), 5.08 (s, 2H), 4.44 (s, 1H), 3.58 – 3.42(m, 3H), 3.25 (d, J = 10.7 Hz, 1H), 2.86 (d, J = 15.2 Hz, 1H), 2.58 (d, J =18.9 Hz, 2H), 1.99 (d, J = 27.1 Hz, 3H). 13 C NMR (101 MHz, DMSO) δ 173.34,170.68, 168.22, 167.75, 152.50, 134.54, 125.45, 115.97, 115.68, 105.89,69.54, 56.80, 49.18, 46.50, 33.97, 31.50, 22.76. ESI [M + H] + (m / z): 344.08.

[0077] Synthesis of intermediate 11b:

[0078]

[0079] The preparation method is the same as 11a in Example 1, and it can be used for the next reaction without further purification.

[0080] Synthesis of compound 2:

[0081]

[0082] The synthesis steps were the same as for compound 1, except that intermediate 11b was used instead of compound 11a to obtain the product, which was a yellow solid powder with a yield of about 59%. 1H NMR (400 MHz, CDCl3) δ 9.22 (s, 1H), 8.60 – 8.52 (m, 1H), 7.91 (s,1H), 7.68 (q, J = 7.1 Hz, 1H), 7.58 (d, J = 6.0 Hz, 1H), 7.46 (d, J = 7.5 Hz,1H), 7.28 (s, 1H), 7.12 (t, J = 7.0 Hz, 1H), 6.88 (s, 2H), 6.64 (d, J = 8.8Hz, 1H), 6.13 (t, J = 7.6 Hz, 1H), 4.90 (dd, J = 12.1, 5.5 Hz, 1H), 3.99 (s,1H), 3.66 – 3.45 (m, 3H), 3.43 – 3.22 (m, 2H), 3.17 – 2.62 (m, 8H), 2.51 (d,J = 8.0 Hz, 3H), 2.30 (dt, J = 34.0, 9.3 Hz, 4H), 2.13 – 1.89 (m, 4H), 1.59 (s, 2H). 13 C NMR (101 MHz, CDCl3) δ 201.86, 171.85, 170.56, 170.15, 167.88,167.17, 166.58, 165.65, 165.35, 159.10, 156.41, 156.05, 150.80, 147.51,146.55, 135.07, 133.41, 130.01, 127.99, 127.38, 124.40, 123.57, 119.89,117.81, 116.26, 115.85, 114.01, 107.90, 104.98, 76.24, 63.00, 54.94, 51.30,50.78, 50.70, 49.49, 48.03, 46.24, 45.85, 35.53, 30.99, 30.45, 29.89, 28.67,28.38, 24.72, 21.83. HRMS (ESI) (m / z): calcd for C 44 H 42 O8F3N8 [M + H] + ,867.3078; found, 867.3041.

[0083] Example 3: Preparation of Compound 3

[0084] Synthesis of intermediate 10c:

[0085]

[0086] The preparation method is the same as 10a, except that compound 4-hydroxypiperidine is used instead of compound 3-azacyclobutanol to obtain product 2-6c, which is a yellow solid powder with a yield of about 91%. 1 H NMR (400 MHz, DMSO-D6) δ 11.08 (s, 1H), 7.64 (d, J= 8.6 Hz, 1H), 7.30 (d, J = 2.4 Hz, 1H), 7.22 (dd, J = 8.6, 2.5 Hz, 1H), 5.05(dd, J = 12.9, 5.4 Hz, 1H), 4.77 (d, J = 4.2 Hz, 1H), 3.81 (dt, J = 13.3, 4.3Hz, 2H), 3.74 (h, J = 4.1 Hz, 1H), 3.18 (ddd, J = 13.1, 9.6, 3.2 Hz, 2H),2.87 (ddd, J = 17.7, 14.2, 5.7 Hz, 1H), 2.62 – 2.52 (m, 2H), 2.06 – 1.94 (m,1H), 1.86 – 1.73 (m, 2H), 1.41 (q, J = 9.1 Hz, 2H). 13 C NMR (101 MHz, DMSO-D6)δ 172.71, 170.00, 167.48, 166.82, 154.59, 133.92, 124.89, 117.39, 117.22,107.54, 65.42, 48.56, 44.78, 33.11, 30.82, 22.03. ESI [M + H] + (m / z): 358.12.

[0087] Synthesis of intermediate 11c:

[0088]

[0089] The preparation method is the same as 11a, and it can be used for the next reaction without further purification.

[0090] Synthesis of compound 3:

[0091]

[0092] The synthesis steps were the same as for compound 1, except that compound 11a was replaced with compound 11c. The product was a yellow solid powder with a yield of about 59%. 1 H NMR (400 MHz, CHLOROFORM-D) δ 8.57 (s, 1H), 7.94 (s, 1H), 7.70 (t,J = 7.8 Hz, 1H), 7.64 (d, J = 8.5 Hz, 1H), 7.48 (d, J = 7.7 Hz, 1H), 7.28 (d,J = 1.8 Hz, 1H), 7.24 (d, J = 2.3 Hz, 1H), 7.14 (d, J = 7.9 Hz, 1H), 7.02(dd, J = 8.7, 2.3 Hz, 1H), 6.90 (s, 1H), 6.13 (d, J = 7.9 Hz, 1H), 4.93 (dd,J = 12.2, 5.4 Hz, 1H), 3.96 (d, J = 12.9 Hz, 3H), 3.87 (d, J = 0.9 Hz, 3H), 3.15 (t, J = 6.0 Hz, 2H), 2.57 – 2.51 (m, 2H), 2.38 (t, J = 11.3 Hz, 2H),1.68 – 1.54 (m, 5H), 1.42 (s, 1H), 1.36 (s, 2H), 1.28 (s, 3H), 1.25 (d, J =1.7 Hz, 6H). 13C NMR (101 MHz, CHLOROFORM-D) δ 203.06, 171.55, 168.83, 168.14,167.37, 166.80, 166.45, 160.21, 157.56, 157.19, 155.09, 148.64, 147.66,136.21, 134.46, 131.10, 129.11, 128.46, 125.57, 124.71, 124.57, 124.47,123.58, 121.02, 118.97, 118.88, 118.00, 117.33, 109.00, 108.71, 77.35, 61.69,56.06, 49.23, 48.33, 47.56, 47.17, 36.64, 35.06, 32.32, 32.01, 31.59, 30.39,30.21, 29.79, 29.75, 29.45, 27.44, 25.82, 22.88, 22.78, 14.22, 1.11. HRMS(ESI) (m / z): calcd for C 45 H 44 O8F3N8 [M + H] + , 881.3235; found, 881.3202.

[0093] Example 4: Preparation of Compound 4

[0094] Synthesis of intermediate 10d:

[0095]

[0096] The preparation method is the same as 10a, except that compound 3-azacyclobutane methanol is used instead of compound 3-azacyclobutanol to obtain the product, which is a yellow solid powder with a yield of about 78%. 1 H NMR (400 MHz, CDCl3) δ 8.59 (s, 1H), 8.00 (s, 1H), 7.60 (d, J = 8.3 Hz, 1H), 6.75 (s, 1H), 6.47 (dd, J = 8.3, 2.4 Hz, 1H), 4.91(dd, J = 12.2, 5.3 Hz, 1H), 4.06 (t, J = 8.1 Hz, 2H), 3.86 – 3.79 (m, 4H), 2.85 – 2.66 (m, 3H), 2.16 – 2.01 (m, 2H). 13C NMR (101 MHz, CDCl3) δ 171.48,168.74, 168.05, 167.63, 155.17, 134.26, 125.30, 117.61, 113.95, 104.88,77.31, 64.00, 53.56, 49.11, 36.60, 31.58, 31.54, 31.49, 22.80. ESI [M + H] + (m / z): 344.13.

[0097] Synthesis of intermediate 11d:

[0098]

[0099] The preparation method is the same as 11a, and it can be used for the next reaction without further purification.

[0100] Synthesis of compound 4:

[0101]

[0102] The synthesis steps were the same as for compound 1, except that compound 11a was replaced with compound 11d. The product was a yellow solid powder with a yield of about 45%. 1H NMR (400 MHz, DMSO-D6) δ 11.08 (s, 1H), 8.81 (s, 1H), 8.68 (s, 1H),8.15 (d, J = 7.8 Hz, 1H), 7.76 (t, J = 7.8 Hz, 1H), 7.61 (d, J = 8.2 Hz, 1H),7.54 (d, J = 8.1 Hz, 1H), 7.37 (s, 1H), 7.20 (d, J = 7.8 Hz, 1H), 7.12 (s,1H), 6.75 (d, J = 2.2 Hz, 1H), 6.61 (dd, J = 8.4, 2.1 Hz, 1H), 5.04 (dd, J =12.9, 5.4 Hz, 1H), 4.11 (t, J = 8.2 Hz, 2H), 3.77 (s, 3H), 3.66 (dd, J = 8.6,5.4 Hz, 3H), 3.14 – 3.06 (m, 2H), 2.98 (p, J = 6.5 Hz, 1H), 2.92 – 2.81 (m,3H), 2.69 – 2.51 (m, 4H), 2.46 (dd, J = 3.8, 1.8 Hz, 2H), 2.08 – 1.98 (m,2H), 1.77 (d, J = 10.3 Hz, 2H), 1.58 (q, J = 10.1 Hz, 2H), 1.20 (s, 1H). 13 CNMR (101 MHz, DMSO-D6) δ 203.55, 173.41, 170.69, 168.06, 167.74, 166.43,165.43, 161.26, 158.37, 157.92, 155.69, 150.11, 148.00, 137.00, 134.35,130.84, 130.00, 128.98, 125.53, 125.36, 122.71, 121.01, 119.74, 117.15,114.56, 110.29, 104.83, 62.17, 56.41, 56.19, 53.00, 49.23, 47.34, 40.21,36.81, 31.96, 31.51, 29.53, 27.75, 25.85, 22.74. HRMS (ESI) (m / z): calcd forC 44 H 42O8F3N8 [M + H] + , 867.3078; found, 867.3039.

[0103] Example 5: Preparation of Compound 5

[0104] Synthesis of intermediate 10e:

[0105]

[0106] The preparation method is the same as 10a, except that compound 3-hydroxymethylpyridine is used instead of compound 3-azacyclobutanol to obtain the product, which is a yellow solid powder with a yield of about 85%. 1 H NMR (400 MHz, DMSO-D6) δ 11.07 (s, 1H), 7.62 (d, J =8.4 Hz, 1H), 6.87 (d, J = 2.2 Hz, 1H), 6.78 (dd, J = 8.5, 2.3 Hz, 1H), 5.04(dd, J = 12.9, 5.4 Hz, 1H), 4.78 (t, J = 5.2 Hz, 1H), 3.51 – 3.41 (m, 5H), 3.18 (dd, J = 10.4, 6.5 Hz, 1H), 2.87 (ddd, J = 17.2, 14.0, 5.5 Hz, 1H), 2.65– 2.51 (m, 3H), 2.16 – 1.91 (m, 2H), 1.79 (dq, J = 12.2, 7.6 Hz, 1H). 13 C NMR (101 MHz, DMSO-D6) δ 173.41, 170.73, 168.27, 167.80, 152.43, 134.53, 125.49,115.95, 115.76, 105.96, 63.04, 51.11, 49.20, 47.84, 41.16, 31.52, 27.92,22.78. ESI [M + H] + (m / z): 358.18.

[0107] Synthesis of intermediate 11e:

[0108]

[0109] The preparation method is the same as 11a, and it can be used for the next reaction without further purification.

[0110] Synthesis of compound 5:

[0111]

[0112] The synthesis steps were the same as for compound 1, except that compound 11a was replaced with compound 11e, resulting in a product that was a yellow solid powder with a yield of approximately 70%. 1 H NMR (400 MHz, DMSO-D6) δ 11.08 (s, 1H), 8.81 (s, 1H), 8.69 (s, 1H), 8.13 (d, J = 7.8 Hz, 1H), 7.77 (t, J = 7.8 Hz, 1H), 7.63 (d, J = 8.5 Hz, 1H),7.55 (d, J = 7.6 Hz, 1H), 7.39 (d, J = 1.8 Hz, 1H), 7.21 (d, J = 7.8 Hz, 1H),7.11 (d, J = 12.3 Hz, 1H), 6.86 (d, J = 2.2 Hz, 1H), 6.77 (dd, J = 8.7, 2.2Hz, 1H), 5.05 (dd, J = 12.8, 5.5 Hz, 1H), 3.78 (s, 3H), 3.20 – 2.76 (m, 7H), 2.65 – 2.52 (m, 3H), 2.34 (d, J = 7.9 Hz, 3H), 2.04 (ddd, J = 33.4, 10.9, 4.8Hz, 5H), 1.84 – 1.52 (m, 6H), 1.21 (s, 3H). 13 C NMR (101 MHz, DMSO-D6) δ203.53, 173.41, 170.73, 168.30, 167.80, 166.42, 165.34, 161.25, 158.35,157.91, 152.38, 150.05, 148.01, 136.99, 134.54, 130.82, 129.99, 128.98,125.52, 121.00, 119.74, 115.95, 115.71, 110.25, 105.90, 61.28, 56.41, 55.43,53.57, 52.96, 52.64, 49.21, 47.73, 47.44, 40.23, 36.82, 36.35, 32.15, 31.52,29.72, 25.86, 22.79. HRMS (ESI) (m / z): calcd for C 45 H 44O8F3N8 [M + H] + ,881.3235; found, 881.3192.

[0113] Example 6: Preparation of Compound 6

[0114] Synthesis of intermediate 10f:

[0115]

[0116] The preparation method is the same as 10a, except that compound 4-piperidinemethanol is used instead of compound 3-azacyclobutanol to obtain the product, which is a yellow solid powder with a yield of about 89%. 1 H NMR (400 MHz, DMSO-D6) δ 11.08 (s, 1H), 7.64 (d, J = 8.6Hz, 1H), 7.29 (s, 1H), 7.22 (d, J = 8.7 Hz, 1H), 5.06 (dd, J = 12.9, 5.4 Hz,1H), 4.52 (t, J = 5.3 Hz, 1H), 4.05 (d, J = 13.4 Hz, 2H), 3.26 (t, J = 5.7Hz, 2H), 2.94 (t, J = 11.6 Hz, 3H), 2.58 (dd, J = 19.9, 6.1 Hz, 3H), 2.14 –1.89 (m, 1H), 1.73 (d, J = 13.1 Hz, 2H), 1.24 – 1.11 (m, 2H). 13 C NMR (101MHz, DMSO-D6) δ 173.40, 170.69, 168.20, 167.52, 155.47, 134.60, 125.56,118.08, 117.78, 108.21, 66.08, 49.25, 47.78, 38.75, 31.51, 28.30, 22.72.

[0117] Synthesis of intermediate 11f:

[0118]

[0119] The preparation method is the same as 11a, and it can be used for the next reaction without further purification.

[0120] Synthesis of compound 6:

[0121]

[0122] The synthesis steps were the same as for compound 1, except that compound 11a was replaced with compound 11f, and the product was a yellow solid powder with a yield of about 64%. 1 H NMR (400 MHz, CHLOROFORM-D) δ 8.69 – 8.41 (m, 1H), 7.93 (s, 1H), 7.70 (t, J = 7.8 Hz, 1H), 7.63 (d, J = 8.5 Hz, 1H), 7.48 (d, J = 7.7 Hz, 1H),7.28 (s, 2H), 7.14 (d, J = 7.9 Hz, 1H), 7.01 (dd, J = 8.7, 2.3 Hz, 1H), 6.91(s, 1H), 6.10 (d, J = 7.9 Hz, 1H), 4.98 – 4.84 (m, 1H), 4.11 (q, J = 7.1 Hz,1H), 3.88 (s, 3H), 3.19 – 3.12 (m, 2H), 2.95 (d, J = 12.2 Hz, 4H), 2.59 –2.51 (m, 2H), 2.27 (d, J = 6.7 Hz, 2H), 2.02 (d, J = 11.9 Hz, 4H), 1.88 (d, J= 13.8 Hz, 2H), 1.29 – 1.22 (m, 7H). 13 C NMR (101 MHz, CHLOROFORM-D) δ 203.02,171.52, 168.87, 168.23, 167.43, 166.44, 160.22, 157.51, 157.18, 155.45,148.64, 147.66, 136.19, 134.47, 131.11, 129.11, 128.50, 125.55, 124.70,121.02, 118.88, 118.52, 117.78, 108.98, 108.64, 77.34, 64.13, 56.06, 53.12,52.98, 49.19, 48.00, 47.01, 36.64, 33.43, 31.99, 31.57, 30.14, 29.78, 25.83,22.89, 14.29. HRMS (ESI) (m / z): calcd for C 46 H 46 O8F3N8 [M + H] +, 895.3391;found, 895.3354.

[0123] Example 7: Preparation of Compound 7

[0124] Synthesis of 10g of intermediate:

[0125]

[0126] The preparation method is the same as 10a, except that compound 2-(azacyclobutane-3-yl)ethanol-1-ol hydrochloride is used instead of compound 3-azacyclobutanol to obtain the product, which is a yellow solid powder with a yield of about 30%. 1 H NMR (400 MHz, DMSO) δ 11.06 (s,1H), 7.62 (d, J = 8.2 Hz, 1H), 6.74 (s, 1H), 6.61 (dd, J = 8.3, 2.1 Hz, 1H), 5.05 (dd, J = 13.0, 5.4 Hz, 1H), 4.13 (t, J = 8.2 Hz, 3H), 3.68 (dd, J = 8.3,5.6 Hz, 2H), 3.45 (d, J = 6.9 Hz, 4H), 2.05 – 1.94 (m, 4H), 1.78 (q, J = 6.6Hz, 3H). 13 C NMR (101 MHz, DMSO) δ 173.32, 170.62, 167.69, 155.74, 134.32,125.31, 117.01, 114.50, 104.77, 59.31, 57.32, 49.19, 37.52, 31.49, 27.45,22.72. ESI [M + H] + (m / z): 358.18.

[0127] Synthesis of 11g of intermediate:

[0128]

[0129] The preparation method is the same as 11a, and it can be used for the next reaction without further purification.

[0130] Synthesis of compound 7:

[0131]

[0132] The synthesis steps were the same as for compound 1, except that compound 11a was replaced with compound 11g, and the product was a yellow solid powder. 1HNMR (400 MHz, DMSO-D6) δ 11.09 (s, 1H), 8.82 (s, 1H), 8.69 (s, 1H), 8.16 (d,J = 7.7 Hz, 1H), 7.78 (t, J = 7.8 Hz, 1H), 7.63 (d, J = 8.3 Hz, 1H), 7.55 (d,J = 7.7 Hz, 1H), 7.39 (s, 1H), 7.21 (d, J = 7.8 Hz, 1H), 7.12 (d, J = 9.0 Hz,1H), 6.76 (d, J = 2.1 Hz, 1H), 6.63 (dd, J = 8.4, 2.1 Hz, 1H), 5.06 (dd, J =12.9, 5.4 Hz, 1H), 4.12 (t, J = 8.1 Hz, 2H), 3.78 (s, 3H), 3.68 (dd, J = 8.5,5.4 Hz, 3H), 3.14 – 3.06 (m, 2H), 2.99 (p, J = 6.8 Hz, 1H), 2.93 – 2.82 (m,4H), 2.66 – 2.53 (m, 5H), 2.47 (dd, J = 3.5, 1.7 Hz, 2H), 2.12 – 1.92 (m,4H), 1.83 – 1.76 (m, 2H), 1.60 (tt, J = 13.1, 6.6 Hz, 2H), 1.22 (s, 1H). 13 CNMR (101 MHz, DMSO-D6) δ 203.55, 173.41, 170.69, 168.06, 167.74, 166.43,165.43, 161.26, 158.37, 157.92, 155.69, 150.11, 148.00, 137.00, 134.35,130.84, 130.00, 128.98, 125.53, 125.36, 122.71, 121.01, 119.74, 117.15,114.56, 110.29, 104.83, 62.17, 56.41, 56.19, 53.00, 49.23, 47.34, 40.21,36.81, 31.96, 31.51, 29.53, 27.75, 25.85, 22.74. HRMS (ESI) (m / z): calcd forC 45 H44 O8F3N8 [M + H] + , 881.3235; found, 881.3193.

[0133] Example 8: Preparation of Compound 8

[0134] Synthesis of intermediate 10h:

[0135]

[0136] The preparation method is the same as 10a, except that compound 2-(pyrrolidine-3-yl)ethanol is used instead of compound 3-azacyclobutanol to obtain the product, which is a yellow solid powder with a yield of about 83%. 1 H NMR (400 MHz, CHLOROFORM-D) δ 8.50 (d, J =11.0 Hz, 1H), 7.61 (d, J = 8.3 Hz, 1H), 6.90 (d, J = 2.3 Hz, 1H), 6.63 (dd, J= 8.5, 2.3 Hz, 1H), 4.93 (dd, J = 12.3, 5.3 Hz, 1H), 3.74 (td, J = 6.5, 2.3Hz, 2H), 3.63 – 3.42 (m, 2H), 3.36 (td, J = 9.7, 6.9 Hz, 1H), 3.02 (t, J =9.3 Hz, 1H), 2.90 – 2.70 (m, 3H), 2.47 (p, J = 7.5 Hz, 1H), 2.30 – 2.04 (m,2H), 1.74 (q, J = 6.5 Hz, 3H), 1.28 – 1.19 (m, 1H). 13 C NMR (101 MHz, CHLOROFORM-D) δ 171.60, 168.84, 168.37, 167.78, 152.12, 134.52, 125.52,116.38, 115.06, 106.14, 77.35, 61.52, 53.81, 49.10, 47.91, 36.08, 35.93,31.65, 31.54, 22.87, 1.11. ESI [M + H] + (m / z): 372.17.

[0137] Synthesis of intermediate 11h:

[0138]

[0139] The preparation method is the same as 11a, and it can be used for the next reaction without further purification.

[0140] Synthesis of compound 8:

[0141]

[0142] The synthesis steps were the same as for compound 1, except that compound 11a was replaced with compound 11h, and the product was a yellow solid powder. 1 HNMR (400 MHz, CHLOROFORM-D) δ 8.56 (s, 1H), 7.94 (s, 1H), 7.68 (t, J = 7.8Hz, 1H), 7.59 – 7.51 (m, 1H), 7.47 (d, J = 7.7 Hz, 1H), 7.28 (d, J = 1.8 Hz,1H), 7.12 (d, J = 7.9 Hz, 1H), 6.92 (s, 1H), 6.85 (t, J = 2.0 Hz, 1H), 6.59(ddd, J = 8.7, 3.6, 2.2 Hz, 1H), 6.22 (d, J = 8.0 Hz, 1H), 4.91 (dd, J =12.1, 5.4 Hz, 1H), 3.86 (s, 3H), 3.57 – 3.27 (m, 3H), 3.14 (t, J = 6.0 Hz, 2H), 3.04 – 2.96 (m, 3H), 2.88 – 2.68 (m, 4H), 2.49 (dt, J = 23.3, 6.8 Hz, 4H), 2.06 – 1.96 (m, 3H), 1.67 (dt, J = 23.8, 11.1 Hz, 6H), 1.28 – 1.19 (m, 3H). 13C NMR (101 MHz, CHLOROFORM-D) δ 203.06, 171.93, 169.22, 168.41, 167.78,166.78, 166.46, 160.20, 157.51, 157.20, 152.07, 148.60, 147.63, 136.21,134.50, 131.04, 129.09, 128.52, 125.48, 124.73, 124.57, 121.87, 120.98,118.96, 117.34, 116.36, 115.05, 108.99, 106.06, 77.36, 57.00, 56.04, 53.70,52.68, 52.50, 49.13, 47.91, 46.97, 37.22, 36.64, 31.91, 31.60, 30.48, 29.78,25.81, 22.95. HRMS (ESI) (m / z): calcd for C 46 H 46 O8F3N8 [M + H] + , 895.3391;found, 895.3347.

[0143] Example 9: Preparation of Compound 9

[0144] Synthesis of intermediate 10i:

[0145]

[0146] The preparation method is the same as 10a, except that compound 2-(piperidin-4-yl)ethane-1-ol is used instead of compound 3-azacyclobutanol to obtain the product, which is a yellow solid powder with a yield of about 80%. 1H NMR (400 MHz, CHLOROFORM-D) δ 8.42 (s, 1H), 7.65 (d, J = 8.5 Hz, 1H), 7.03 (d, J = 8.7 Hz, 1H), 4.93 (dd, J = 12.3, 5.4Hz, 1H), 3.93 (d, J = 12.8 Hz, 2H), 3.72 (t, J = 6.5 Hz, 2H), 3.03 – 2.63 (m,5H), 2.18 – 2.05 (m, 1H), 1.82 (d, J = 13.7 Hz, 2H), 1.75 (s, 2H), 1.54 (q, J= 6.5 Hz, 2H), 1.36 – 1.23 (m, 2H). 13 C NMR (101 MHz, CHLOROFORM-D) δ 171.46,168.65, 168.20, 167.42, 155.42, 134.47, 125.56, 118.50, 117.84, 108.69,77.35, 60.21, 49.17, 48.29, 39.08, 32.41, 31.58, 31.53, 22.83. ESI [M + H] + (m / z): 386.16.

[0147] Synthesis of intermediate 11i:

[0148]

[0149] The preparation method is the same as 11a, and it can be used for the next reaction without further purification.

[0150] Synthesis of compound 9:

[0151]

[0152] The synthesis steps were the same as for compound 1, except that compound 11i was used instead of compound 11a. The product was a yellow solid powder with a yield of about 58%. 1H NMR (400 MHz, CHLOROFORM-D) δ 8.54 (s, 1H), 7.96 (s, 1H), 7.69 (t,J = 7.8 Hz, 1H), 7.58 (d, J = 8.4 Hz, 1H), 7.49 (d, J = 7.7 Hz, 1H), 7.30 (d,J = 2.2 Hz, 1H), 7.17 (d, J = 2.3 Hz, 1H), 7.12 (d, J = 7.9 Hz, 1H), 6.95(dd, J = 8.7, 2.4 Hz, 2H), 6.85 (d, J = 7.9 Hz, 1H), 4.93 (dd, J = 11.8, 5.5Hz, 1H), 3.84 (s, 3H), 3.33 (d, J = 11.0 Hz, 2H), 3.14 (t, J = 5.8 Hz, 2H),2.85 (t, J = 14.5 Hz, 3H), 2.76 (d, J = 7.8 Hz, 3H), 2.52 (d, J = 6.7 Hz,4H), 2.00 (s, 4H), 1.75 (d, J = 12.2 Hz, 2H), 1.29 – 1.18 (m, 10H). 13 C NMR(101 MHz, CHLOROFORM-D) δ 203.20, 176.79, 172.00, 169.25, 168.18, 167.42,166.80, 166.58, 160.17, 157.46, 157.29, 155.31, 148.55, 147.53, 136.25,134.40, 131.14, 129.05, 127.90, 125.47, 124.87, 124.57, 121.88, 120.95,119.41, 118.58, 117.86, 109.02, 108.55, 77.37, 56.08, 55.21, 52.12, 49.23,47.95, 45.66, 36.67, 33.94, 32.00, 31.60, 31.33, 30.00, 29.78, 29.44, 22.86,22.78, 14.22, 1.10. HRMS (ESI) (m / z): calcd for C 47 H 48 O8F3N8 [M + H] +,909.3548; found, 909.3505.

[0153] Example 10: Preparation of Compound 10

[0154] Synthesis of intermediate 10j:

[0155]

[0156] The preparation method is the same as 10a, except that compound 3-(4-piperidinyl)-1-propanol is used instead of compound 3-azacyclobutanol to obtain the product, which is a yellow solid powder with a yield of about 70%. 1 H NMR (400 MHz, CHLOROFORM-D) δ 8.36 (s, 1H), 7.65 (d, J = 8.6 Hz, 1H), 7.04 (dd, J = 8.6, 2.5 Hz, 1H), 4.93 (dd, J = 12.4,5.3 Hz, 1H), 3.93 (d, J = 13.1 Hz, 2H), 3.65 (t, J = 6.6 Hz, 2H), 3.01 – 2.62(m, 5H), 2.21 – 2.06 (m, 1H), 1.81 (d, J = 10.9 Hz, 2H), 1.65 – 1.55 (m, 3H),1.40 – 1.19 (m, 5H). 13 C NMR (101 MHz, CHLOROFORM-D) δ 171.39, 168.58, 168.19,167.41, 155.39, 134.47, 125.55, 118.55, 117.88, 108.71, 63.07, 49.17, 48.37,35.67, 32.49, 31.60, 31.53, 29.81, 22.83. ESI [M + H] + (m / z): 400.21.

[0157] Synthesis of intermediate 11j:

[0158]

[0159] The preparation method is the same as 11a, and it can be used for the next reaction without further purification.

[0160] Synthesis of compound 10:

[0161]

[0162] The synthesis steps were the same as for compound 1, except that compound 11j was used instead of compound 11a. The product was a yellow solid powder with a yield of about 58%. 1 H NMR (400 MHz, CHLOROFORM-D) δ 8.56 (d, J = 0.8 Hz, 1H), 7.94 (s,1H), 7.69 (t, J = 7.8 Hz, 1H), 7.60 (d, J = 8.6 Hz, 1H), 7.48 (d, J = 7.7 Hz,1H), 7.29 (d, J = 1.9 Hz, 1H), 7.20 (d, J = 2.3 Hz, 1H), 7.13 (d, J = 7.8 Hz,1H), 6.99 (dd, J = 8.7, 2.3 Hz, 1H), 6.92 (s, 1H), 6.38 (d, J = 8.0 Hz, 1H),4.93 (dd, J = 12.1, 5.4 Hz, 1H), 3.86 (s, 3H), 3.15 (t, J = 6.1 Hz, 4H), 2.97– 2.62 (m, 6H), 2.52 (q, J = 6.4 Hz, 4H), 2.33 (t, J = 12.1 Hz, 2H), 2.12 –2.00 (m, 3H), 1.79 (t, J = 11.1 Hz, 4H), 1.29 – 1.19 (m, 9H). 13 C NMR (101 MHz, CHLOROFORM-D) δ 203.08, 171.71, 168.98, 168.23, 167.44, 166.81, 166.48,160.20, 157.54, 157.22, 155.39, 148.60, 147.60, 136.22, 134.46, 131.14,129.09, 128.21, 125.51, 124.77, 124.57, 121.88, 120.99, 119.06, 118.40,117.76, 117.27, 108.99, 108.54, 77.36, 58.38, 56.07, 52.52, 49.21, 48.19,46.40, 36.65, 35.62, 33.99, 31.60, 31.53, 31.17, 29.78, 25.82, 23.27, 22.89.HRMS (ESI) (m / z): calcd for C 48 H50 O8F3N8 [M + H] + , 923.3704; found, 923.3657.

[0163] Example 11: Preparation of Compound 11

[0164] Synthesis of intermediate 12:

[0165]

[0166] 4-Amino-3-methoxybenzoic acid (2 g, 4.354 mmol), HATU (1.98 g, 5.224 mmol), and DIPEA (1.52 ml, 8.708 mmol) were dissolved in DMF and stirred at room temperature for 30 min. Then, 1-Boc-4-aminopiperidine (1.3 mg, 6.531 mmol) was added to the reaction solution, and the reaction mixture was stirred continuously at room temperature for 3 h. The reaction was monitored by TLC. After the reaction was completed, water was added to quench the reaction. The resulting mixture was extracted with ethyl acetate (500 mL × 3), the organic phases were combined, washed with 100 ml of saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to obtain the compound as a white solid powder in approximately 75% yield. 1 H NMR (400 MHz, CDCl3) δ7.34 (d, J = 4.4 Hz, 1H), 7.16 – 7.01 (m, 1H), 6.64 (dd, J = 8.5, 4.7 Hz,1H), 5.96 (d, J = 7.3 Hz, 1H), 4.23 – 3.97 (m, 3H), 3.89 (q, J = 2.6 Hz, 3H), 2.89 (t, J = 12.9 Hz, 2H), 1.99 (d, J = 12.4 Hz, 2H), 1.46 (d, J = 3.4 Hz,9H), 1.34 (d, J = 16.2 Hz, 2H). 13 C NMR (101 MHz, CDCl3) δ 166.77, 154.80,146.82, 139.71, 124.22, 119.58, 113.26, 109.84, 79.69, 77.29, 55.68, 47.11,32.31, 29.74, 28.49.

[0167] Synthesis of intermediate 13:

[0168]

[0169] The synthesis steps were the same as those for compound 3, except that compound 4-amino-3-methoxybenzoate was replaced with compound 2,4,5-trichloropyrimidine to obtain an intermediate, a yellowish-white solid powder, with a yield of approximately 56%. 1 H NMR (400 MHz, DMSO) δ8.03 (s, 1H), 6.94 (t, J = 7.8 Hz, 1H), 6.71 (d, J = 7.7 Hz, 1H), 6.44 (d, J= 7.9 Hz, 1H), 2.35 – 2.24 (m, 2H), 1.80 – 1.72 (m, 2H). 13 C NMR (101 MHz, DMSO) δ 203.87, 165.37, 159.60, 158.36, 156.54, 147.71, 137.18, 128.01,126.06, 120.82, 116.88, 36.90, 25.89. ESI [M + H] + (m / z): 294.94.

[0170] Synthesis of intermediate 14:

[0171]

[0172] Intermediate 12 (300 mg, 0.86 mmol), intermediate 13 (254 mg, 0.86 mmol), Xantphos (75 mg, 0.129 mmol), palladium acetate (14 mg, 0.0602 mmol), and cesium carbonate (981 mg, 3.01 mmol) were placed in a round-bottom flask, and tetrahydrofuran was added as a solvent. Under nitrogen protection, the reaction system was stirred at 120 °C for 3 h. The reaction was monitored by TLC. After the reaction was completed, water was added to quench the reaction. The resulting mixture was extracted with ethyl acetate (500 mL × 3), the organic phases were combined, washed with saturated brine (100 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography to give the intermediate as a white solid powder, with a yield of approximately 53%. 1H NMR (400 MHz, CDCl3)δ 8.34 (s, 1H), 7.73 (d, J = 6.5 Hz, 2H), 7.49 (d, J = 7.6 Hz, 1H), 7.18 (d,J = 7.8 Hz, 1H), 6.89 (d, J = 8.5 Hz, 1H), 5.91 (d, J = 7.7 Hz, 1H), 4.23 –3.98 (m, 3H), 3.89 (s, 3H), 3.18 (t, J = 5.8 Hz, 2H), 2.91 (t, J = 12.7 Hz,2H), 2.68 – 2.37 (m, 2H), 2.03 (d, J = 16.8 Hz, 2H), 1.48 (s, 9H), 1.28 (d, J= 9.6 Hz, 2H). 13 C NMR (101 MHz, CDCl3) δ 203.08, 166.45, 164.68, 157.61,157.14, 156.69, 154.77, 149.23, 147.29, 136.10, 131.93, 129.06, 127.36,124.48, 121.07, 118.86, 116.40, 108.71, 107.54, 79.77, 77.28, 55.96, 47.32,36.64, 32.28, 29.75, 28.50, 25.80. ESI [M + H] + (m / z): 608.25.

[0173] Synthesis of compound 11:

[0174]

[0175] The synthesis steps were the same as for compound 1, yielding a product that was a yellow solid powder with a yield of approximately 65%. 1H NMR (400MHz, CDCl3) δ 8.32 (d, J = 1.1 Hz, 1H), 7.73 (s, 1H), 7.68 (t, J = 7.7 Hz,1H), 7.58 (d, J = 8.4 Hz, 1H), 7.45 (d, J = 7.7 Hz, 1H), 7.14 (d, J = 7.8 Hz,1H), 6.93 (d, J = 8.5 Hz, 1H), 6.88 (d, J = 2.8 Hz, 1H), 6.62 (d, J = 8.6 Hz,1H), 6.18 (d, J = 7.0 Hz, 1H), 5.28 (d, J = 1.1 Hz, 1H), 4.92 (dd, J = 11.9,5.4 Hz, 1H), 3.84 (s, 3H), 3.56 – 3.48 (m, 1H), 3.48 – 3.28 (m, 2H), 3.15 (d,J = 6.3 Hz, 2H), 3.04 – 2.69 (m, 5H), 2.64 – 2.56 (m, 1H), 2.55 – 2.47 (m,2H), 2.41 (d, J = 10.5 Hz, 2H), 2.15 (ddd, J = 24.8, 12.8, 6.0 Hz, 4H), 2.01(d, J = 12.2 Hz, 2H), 1.77 (dq, J = 16.3, 8.4 Hz, 1H), 1.61 (s, 2H). 13 C NMR(101 MHz, CDCl3) δ 203.11, 171.59, 168.92, 168.32, 167.65, 166.43, 164.58,157.63, 157.12, 156.68, 152.05, 149.13, 147.20, 136.06, 134.41, 131.74,128.98, 127.51, 125.40, 124.47, 120.98, 118.96, 116.35, 115.04, 108.75,107.37, 106.09, 77.26, 61.39, 55.90, 53.46, 52.58, 52.38, 49.05, 47.51,36.60, 32.07, 31.47, 29.82, 25.73, 22.87. HRMS (ESI) (m / z): calcd forC 44 H44 O8ClN8 [M + H] + , 847.2971; found, 847.2932.

[0176] Example 12: Kinase inhibitory activity of the compound

[0177] (1) Method

[0178] Dilute the compound to 100-fold the maximum inhibitor concentration required for the final reaction using 100% DMSO. Transfer 100 μL of this diluted compound to the wells of a 96-well plate. Add 50 μL of 100% DMSO to two empty wells in the same 384-well plate for a compound-free control and an enzyme-free control. Label the wells as source wells. Transfer 200 nL of the compound from each well to a 384 assay plate using Echo. Add 10 μL of 1× kinase buffer to each well, except for the enzyme-free control well. Then, add an appropriate amount of peptide and ATP to the 1× kinase buffer to prepare 2× substrate working solution. Add 10 μL of 2× substrate working solution to each well of the assay plate to start the reaction and incubate at room temperature for 60 minutes. Simultaneously, prepare a 2-fold final concentration of the assay solution using antibody dilution buffer. After the reaction is complete, add 20 μL of the assay solution to each well of the assay plate and incubate at room temperature for 60 minutes to terminate the reaction. Data were collected on Envision at 340 nm excitation and 520 nm and 495 nm emission.

[0179] (2) Results

[0180]

[0181] Example 13: Protein degradation effect of PROTACs

[0182] Western blot at the cellular level

[0183] 1) Compound processing and administration

[0184] MIA-PACA-2 cells were revived and cultured until they reached the logarithmic growth phase, at which point cell counts were performed. When the cell density reached approximately 500,000 cells per milliliter of culture medium, the cells were seeded into 6-well plates. A 10 mM stock solution of the test compound was prepared using 100% DMSO and stored at -80°C. During the experiment, cells were treated with the compound at concentration gradients of 0.001 μM, 0.01 μM, 0.1 μM, 1 μM, and 10 μM. After 24 hours of treatment, the expression level of the target protein in the cells was detected by Western blotting (WB).

[0185] 2) WB detection

[0186] After administering different drug concentrations to cells, centrifuge at 5000 rpm for 10 min 24 h later, discard the culture medium, and wash the cells twice with pre-chilled PBS to remove residual culture medium. Add an appropriate amount of RIPA lysis buffer (usually 100-200 μL) to each well and lyse on ice for 30 min, gently shaking the culture plate during lysis to ensure complete lysis. Then transfer to pre-chilled 1.5 mL centrifuge tubes and centrifuge at 12000 rpm for 30 min at 4 °C, collecting the supernatant (i.e., total protein extract). The lysed protein samples are quantified using the BCA method to ensure consistent protein concentration across samples. After loading equal volumes of protein samples, electrophoresis is performed using a Tricine-SDS-PAGE gel system, and the proteins are transferred from the gel to a nitrocellulose membrane (Amersham, 10600002) using a wet transfer method. Nitrocellulose membranes were placed in TBS-T (Tris-buffered saline solution containing 0.1% Tween-20) blocking buffer containing 5% skim milk powder and incubated on a shaker at room temperature for 1 hour to block non-specific binding sites on the membrane. Subsequently, the membrane was incubated overnight with specific primary antibody at 4°C. The next day, the membrane was washed three times with TBS-T solution for 15 minutes each time to remove unbound primary antibody. After washing, secondary antibody was added and incubated at room temperature for 1 hour, followed by washing three more times with TBS-T solution for 15 minutes each time to remove unbound secondary antibody. Finally, the membrane was incubated with ECL chemiluminescence reagent in the dark for 1 minute, followed by imaging and analysis using a chemiluminescence imaging system.

[0187] Table 1. Degradative activity of the degradative compounds against FAK in MIA-PACA-2 cells.

[0188]

[0189] As can be seen from the experimental results, all compounds exhibit good degradation activity against FAK protein. Among them, compound 11 shows particularly outstanding degradation activity (DC). 50 < 1 nM, Dmax = 94.60%.

[0190] Example 14: Inhibition of cell proliferation by compound 11

[0191] Cells were seeded at 2000 cells / well in 100 μL of complete culture medium into 96-well microplates. Fresh culture medium containing different concentrations of the test compounds was then added, and the plates were incubated for 72 h. Next, 10 μL of CCK-8 solution was added to each well, and the plates were incubated at 37°C for 4 h. Cell viability was measured using a microplate reader at 450 nm. Cell viability (%) = (OD value of treatment group) - (OD value of blank group) / (OD value of control group) - (OD value of blank group) × 100.

[0192] The results are attached. Figure 6 As can be seen, the experimental results show that compound 11 did not significantly inhibit the proliferation of MIA-PACA-2 cells (the inhibition rate was less than 20%). This phenomenon may be closely related to the biological functional characteristics of FAK protein in cells: FAK mainly acts as a cytoskeleton-related protein, participating in the regulation of cell migration, adhesion and survival through integrin-mediated extracellular matrix signaling, rather than directly regulating cell cycle progression or proliferation-related signaling pathways.

[0193] Example 15: Co-administration of PROTACs with KRAS G12C inhibitors

[0194] We co-administered compound 11 at a concentration of 3 μM with different concentrations of the KRAS G12C inhibitor AMG 510 to further evaluate the synergistic effect of the FAK degrader and AMG 510. Results are attached. Figure 7 .

[0195] Experimental results show that compound 11 enhances the sensitivity of MIA-PACA-2 to AMG 510.

[0196] Example 16: Pharmacokinetic Evaluation

[0197] 1) Sample plasma preparation

[0198] Accurately weigh an appropriate amount of the compound and prepare a solution of 10 mg / kg using 5% DMSO and 95% HS 15 solvent. Administer the compound via gavage. Mice were grouped according to body weight, with 3 mice per group for each compound. At 0.25 h, 0.5 h, 1 h, 2 h, 4 h, 6 h, 8 h, and 12 h post-gavage administration, blood was collected from the orbital fossa in EP tubes containing 20 μl of anticoagulant. Finally, blood samples were collected from the corresponding mice via orbital fossa. All collected blood samples were centrifuged at 4000 rpm for 10 min within 1 h, and the supernatant was separated from the serum. Acetonitrile solution was added to the plasma samples to precipitate proteins. The samples were vortexed for 1 min, centrifuged at 10000 rpm for 10 min, and the supernatant was analyzed by LC-MS.

[0199] 2) Establishment of sample method

[0200] Prepare a 500 ng / mL standard solution to determine the optimal detection method. First, select the full scan mode for mass spectrometry and set the scan range according to the compound's molecular weight. After determining the retention time, use the icon-production method to find the daughter ions. Determine an optimal energy value by setting different energies; generally, the intensity of the parent ion should be 1 / 4 of the daughter ion intensity. After determining the optimal energy and daughter ions, use the MRM method. Input the daughter ion, parent ion, and energy into the selected method. After the scan, check the results. Once the MRM spectrum is confirmed to be normal, establish the analytical method for this sample based on the previously determined conditions.

[0201] 3) Preparation of standard curve and processing of drug-treated plasma samples

[0202] Nine standard solutions of different concentrations were prepared (47.5 μL blank plasma + 2.5 μL working solution + 200 μL 5 ng / mL loratadine internal standard solution): 2.5 ng / mL, 5 ng / mL, 10 ng / mL, 25 ng / mL, 50 ng / mL, 100 ng / mL, 250 ng / mL, 500 ng / mL, and 1000 ng / mL. A double-blank solution (50 μL blank plasma + 200 μL acetonitrile) and a single-blank solution (50 μL blank plasma + 200 μL loratadine internal standard solution) were also prepared. All solutions were centrifuged, and the supernatant was collected.

[0203] Take 50 μL of plasma sample from each time point and add 200 μL of loratadine internal standard solution to prepare analytical samples.

[0204] 4) Instrumental analysis and data processing

[0205] The standard curve sample and analytical sample were analyzed using the previously established sample analysis method. After the sample run was completed, the triple quadrupole quantitative analysis software was used to add the sample, specify the target ion and internal standard compound for quantification, set the concentration level, input the corresponding internal standard concentration and correction concentration, check the qualitative ion settings, and finally save the analysis results.

[0206] From the appendix Figure 8 It can be seen that, compared with compounds 3 and 4, compound 5 has relatively better PK properties, with Cmax of 29.9 ng / mL and AUC0-t of 48.0 ng / mL*h.

[0207] From the appendix Figure 9 It can be seen that compound 3 reached its maximum plasma concentration (Tmax) 0.25 hours after administration, with a Cmax of 63.12 ng / mL. Further analysis of the area under the curve (AUC0-t) revealed that the AUC0-t value of compound 3 was 111.86 ng / mL*h.

Claims

1. A compound that targets FAK degradation, characterized in that, The structure shown in equation (I): ； in, R1 is either -CF3 or -Cl; Linker is a connecting chain, and its structure is as follows: 。 2. The compound targeting FAK degradation according to claim 1, characterized in that, Compounds with the following structure: Compound 1: ; Compound 2: ; Compound 3: ; Compound 4: ; Compound 5: ; Compound 6: ; Compound 7: ; Compound 8: ; Compound 9: ; Compound 10: ; Compound 11: .

3. The use of the compound targeting FAK degradation according to claim 1 or 2 in the preparation of medicaments for treating, preventing and alleviating diseases caused by excessive expression of FAK protein.

4. The use of stereoisomers, tautomers, deuterated compounds, solvates, prodrugs, metabolites, and pharmaceutically acceptable salts thereof of the compounds targeting FAK degradation according to claim 1 or 2 in the preparation of medicaments for treating, preventing, and alleviating diseases caused by excessive expression of FAK protein.

5. The application according to claim 3 or 4, characterized in that, The diseases caused by excessive FAK protein expression include cholangiocarcinoma, diffuse large B-cell lymphoma, clear cell renal cell carcinoma, low-grade glioma, sarcoma, thymoma, mesothelioma, meningioma, gastric adenocarcinoma, melanoma, lung cancer, small cell lung cancer, non-small cell lung cancer, lung adenocarcinoma, hepatocellular carcinoma, gastric cancer, intestinal cancer, pancreatic cancer, glioblastoma, ovarian cancer, liver cancer, bladder cancer, hepatocellular carcinoma, breast cancer, metastatic breast cancer, colon cancer, rectal cancer, colorectal cancer, kidney cancer, head and neck cancer, pheochromocytoma, or paraganglioma.

6. The use of the compound targeting FAK degradation according to claim 1 or 2 in the preparation of a drug for treating pancreatic cancer.

7. The use of stereoisomers, tautomers, deuterated compounds, solvates, prodrugs, metabolites, and pharmaceutically acceptable salts thereof of the compounds targeting FAK degradation according to claim 1 or 2 in the preparation of drugs for treating pancreatic cancer.

8. The use of the compound targeting FAK degradation according to claim 1 or 2 in the preparation of a medicament in combination with a KRAS inhibitor.

9. The use of stereoisomers, tautomers, deuterated compounds, solvates, prodrugs, metabolites, and pharmaceutically acceptable salts thereof of the compounds targeting FAK degradation according to claim 1 or 2 in the preparation of medicaments in combination with KRAS inhibitors.

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