Compound capable of increasing blood-brain barrier permeability, preparation method thereof and application thereof

Abstract

The present invention relates to the technical field of compound synthesis, and in particular, to a compound capable of enhancing the permeability of the blood-brain barrier, a preparation method thereof, and an application thereof. The compound capable of enhancing the permeability of the blood-brain barrier has the following structural formula: wherein X represents a halogen, R1 represents a C1-C5 alkyl, and R represents a substituted or unsubstituted alkyl and a substituted or unsubstituted phenyl. The compound has a high blood-brain barrier permeability, and can then enter the brain more quickly, which is beneficial for the compound to exert its pharmacological effect. At the same time, the compound can simultaneously inhibit ALK and EGFR, and can serve as a dual inhibitor of ALK and EGFR, and has a good therapeutic effect on cancers such as non-small cell lung cancer, anaplastic lymphoma, lung adenocarcinoma, and glioma.

Description

Technical Field

[0001] The present invention relates to the technical field of compound synthesis, and in particular to a compound capable of enhancing the permeability of the blood-brain barrier, a preparation method thereof, and an application thereof. Background Art

[0002] The blood-brain barrier refers to the barrier between plasma and brain cells, formed by the walls of brain capillaries and glial cells, and between plasma and cerebrospinal fluid, formed by the choroid plexus. These barriers prevent certain substances from entering the brain from the blood. Specifically, various solutes in the blood enter the brain from the capillaries, with varying degrees of difficulty; some pass quickly, some more slowly, and some are completely blocked. This selective permeability has led to the hypothesis that certain structures may exist that restrict solute permeation. Such structures could reduce or even inactivate brain tissue from harmful substances in the circulating blood, thereby maintaining a basic stability of the brain's internal environment. This has important biological significance for maintaining the normal physiological state of the central nervous system.

[0003] In summary, the blood-brain barrier can prevent many substances that are harmful to the central nervous system from passing through, but this barrier also prevents many therapeutic drugs from entering the central nervous system, which in turn has adverse effects on the treatment of many brain diseases, such as tumors or degenerative diseases, and is not conducive to the drugs exerting their efficacy.

[0004] In view of this, the present invention is proposed. Summary of the Invention

[0005] The object of the present invention is to provide a compound that can enhance the permeability of the blood-brain barrier, a preparation method thereof, and its application. The compound provided in the embodiment of the present invention has a high permeability of the blood-brain barrier, which can then enter the brain more quickly, which is conducive to the compound exerting its pharmacological effect. At the same time, the compound can simultaneously inhibit ALK and EGFR, and can be used as a dual inhibitor of ALK and EGFR, with good therapeutic effects on cancers such as non-small cell lung cancer, anaplastic lymphoma, lung adenocarcinoma, and glioma.

[0006] The present invention is achieved in that:

[0007] In a first aspect, the present invention provides a compound capable of increasing the permeability of the blood-brain barrier, the structural formula of which is shown below:

[0008]

[0009] wherein X represents a halogen, R1 represents a C1-C5 alkyl group, and R represents a substituted or unsubstituted alkyl group and a substituted or unsubstituted phenyl group.

[0010] In a second aspect, the present invention provides a method for preparing the compound capable of enhancing blood-brain barrier permeability as described in the aforementioned embodiment, which is synthesized according to the following synthetic route:

[0011]

[0012] wherein X, X1 and X2 are independently halogen, R1 is a C1-C5 alkyl group, and R is a substituted or unsubstituted alkyl group and a substituted or unsubstituted phenyl group.

[0013] In a third aspect, the present invention provides a use of the compound capable of enhancing the permeability of the blood-brain barrier as described in the aforementioned embodiment in the preparation of an inhibitor for inhibiting ALK and / or EGFR.

[0014] In a fourth aspect, the present invention provides a use of the compound capable of enhancing the permeability of the blood-brain barrier as described in the aforementioned embodiment in the preparation of a drug for treating cancer, wherein the cancer includes non-small cell lung cancer, anaplastic lymphoma, lung adenocarcinoma and glioma.

[0015] The present invention has the following beneficial effects: The compounds provided by the embodiments of the present invention have high blood-brain barrier permeability, allowing them to enter the brain more quickly, thus facilitating the compound's efficacy. Furthermore, the compounds can simultaneously inhibit ALK and EGFR, acting as dual ALK and EGFR inhibitors, demonstrating promising therapeutic effects against cancers such as non-small cell lung cancer, anaplastic lymphoma, lung adenocarcinoma, and glioma. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the following briefly introduces the drawings required for use in the embodiments. It should be understood that the following drawings only illustrate certain embodiments of the present invention and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other relevant drawings can be obtained based on these drawings without paying any creative work.

[0017] Figure 1 This is the NMR spectrum of the compound capable of enhancing blood-brain barrier permeability provided in Example 1 of the present invention;

[0018] Figure 2 This is the mass spectrum of the compound capable of enhancing blood-brain barrier permeability provided in Example 1 of the present invention;

[0019] Figure 3 This is a carbon spectrum of the compound capable of enhancing blood-brain barrier permeability provided in Example 1 of the present invention;

[0020] Figure 4 This is the NMR spectrum of the compound capable of enhancing blood-brain barrier permeability provided in Example 15 of the present invention;

[0021] Figure 5 This is the mass spectrum of the compound capable of enhancing blood-brain barrier permeability provided in Example 15 of the present invention;

[0022] Figure 6 This is a carbon spectrum of the compound capable of enhancing blood-brain barrier permeability provided in Example 15 of the present invention;

[0023] Figure 7 This is the NMR spectrum of the compound capable of enhancing blood-brain barrier permeability provided in Example 28 of the present invention;

[0024] Figure 8 This is the mass spectrum of the compound capable of enhancing blood-brain barrier permeability provided in Example 28 of the present invention;

[0025] Figure 9 This is the carbon spectrum of the compound capable of enhancing blood-brain barrier permeability provided in Example 28 of the present invention. DETAILED DESCRIPTION

[0026] To make the purpose, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention are described clearly and completely below. Where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer are used. Where the manufacturer of the reagents or instruments is not specified, all are conventional products that can be purchased commercially.

[0027] The present invention provides a compound capable of enhancing the permeability of the blood-brain barrier, and its structural formula is shown below:

[0028]

[0029] wherein X represents a halogen, R1 represents a C1-C5 alkyl group, and R represents a substituted or unsubstituted alkyl group and a substituted or unsubstituted phenyl group.

[0030] The present invention uses 2,4-diamino-5-halopyrimidine and 4-(4-alkylpiperazin-1-yl)aniline to form the parent core, and selects a specific R group, which gives the compound good blood-brain barrier permeability, allowing it to efficiently cross the blood-brain barrier and better exert its pharmacological efficacy. Furthermore, the compound is a dual-target inhibitor of ALK and EGFR, capable of effectively binding to both ALK and EGFR proteins, respectively, retaining key interactions while also increasing the interaction between additional amino acid residues.

[0031] Specifically, X represents any one of fluorine, chlorine and bromine; preferably chlorine.

[0032] R1 represents a C1-C5 straight chain alkyl group, preferably a C1-C3 straight chain alkyl group, preferably a methyl group or an ethyl group, and most preferably an ethyl group. It is understood that R1 can also be a C1-C5 branched chain alkyl group such as isopropyl, isobutyl, or tert-butyl.

[0033] For example, the structural formula of this compound is shown below:

[0034]

[0035] The definitions of R in Formula 1 and Formula 2 are the same. Specifically, R represents a substituted or unsubstituted alkyl group and a substituted or unsubstituted phenyl group; for example, selected from a C1-C10 substituted or unsubstituted alkyl group; preferably a C1-C7 substituted or unsubstituted alkyl group; more preferably any one of a substituted or unsubstituted methyl group, a substituted or unsubstituted ethyl group, a substituted or unsubstituted propyl group, a substituted or unsubstituted butyl group, a substituted or unsubstituted pentyl group, and a substituted or unsubstituted hexyl group.

[0036] The substituted or unsubstituted alkyl group may be a straight-chain alkyl group or a branched-chain alkyl group. For example, the substituted or unsubstituted propyl group may be a substituted or unsubstituted n-propyl group or a substituted or unsubstituted isopropyl group. Similarly, the substituted or unsubstituted butyl group may be n-butyl, isobutyl or tert-butyl group. The substituted or unsubstituted pentyl group may be n-pentyl, isopentyl, sec-pentyl or tert-pentyl group. The substituted or unsubstituted hexyl group may be n-hexyl, isohexyl, sec-hexyl or tert-hexyl group.

[0037] The substituents in the above-mentioned substituted alkyl group are selected from C3-C5 cycloalkyl, C3-C5 heterocycloalkyl, and the heteroatoms are selected from any one of N, O and S or any two or more combinations thereof, hydroxyl, phenyl, halogen, alkenyl, amino, and any one of para-substituted phenyl; wherein the substituents in the para-substituted phenyl group are selected from any one of sulfonylamino, hydroxyl and halogen; for example, including but not limited to para-substituted phenyl, hydroxyl, halogen, phenyl, alkenyl, amino,

[0038] wherein the substituent of the para-monosubstituted phenyl group is selected from any one of sulfonylamino, hydroxyl and halogen.

[0039] It should be noted that the above Indicates the site of attachment to the parent nucleus.

[0040] The above-mentioned substituents may replace one hydrogen atom on one carbon atom, may replace multiple hydrogen atom on one carbon atom, or may replace multiple hydrogen atom on multiple carbon atom.

[0041] Furthermore, the substituted phenyl group in R is selected from any one of the groups represented by the following structural formulas:

[0042] Any of, wherein R2, R4 and R6 can be located at any position of the benzene ring, but R6 is not located at Key alignment.

[0043] Specifically, R2 is selected from any one of nitro, halogen, C1-C5 alkyl (e.g., methyl, ethyl, n-propyl, isopropyl, tert-butyl, n-butyl, n-pentyl, isopentyl, and tert-pentyl), thioether, and C3-C6 cycloalkyl (e.g., cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl), and R3, R4, R5, and R6 are independently selected from halogen (e.g., fluorine, bromine, and chlorine) or C1-C3 alkoxy (e.g., methoxy, ethoxy, and propoxy). The groups selected from R3, R4, R5, and R6 may be the same or different.

[0044] More specifically, the compound capable of increasing the permeability of the blood-brain barrier is selected from any one of the compounds represented by the following structural formulas:

[0045]

[0046]

[0047]

[0048] The numbers in the above structural formula correspond to the numbers of the compounds provided in the examples and their compounds.

[0049] In a second aspect, the present invention provides a method for preparing the compound capable of enhancing blood-brain barrier permeability, which is synthesized according to the following synthetic route:

[0050]

[0051] wherein X, X1 and X2 are independently halogen, R1 is a C1-C5 alkyl group, and R is a substituted or unsubstituted alkyl group and a substituted or unsubstituted phenyl group.

[0052] In step a, the molar ratio of Compound I to Compound II is (1.1-1.2):1, for example, 1.1:1, 1.11:1, 1.12:1, 1.13:1, 1.14:1, 1.15:1, 1.16:1, 1.17:1, 1.18:1, 1.19:1, 1.2:1, and any other value between (1.1-1.2):1. The reaction temperature is 70-85°C, for example, 70°C, 75°C, 80°C, and 85°C. The solvent used can be a commercially available alcohol solvent, and an acid-binding agent, such as, but not limited to, an amine such as N,N-diisopropylethylamine, is also used in step a. The reaction progress is monitored by thin-layer chromatography.

[0053] Specifically, 1 mmol of different R-substituted amine compounds was weighed and dissolved in about 5 mL of anhydrous isopropyl alcohol (isopropyl alcohol was added to 3A molecular sieves and allowed to stand for 24 hours to remove water). 3 mmol of acid binder N, N-diisopropylethylamine was added to the dissolved amine compound. Heat and stir at 80°C for 4 hours. After ensuring that the mixture was uniform, 1.15 mmol of 2,4,5-trichloropyrimidine was slowly added dropwise. At this time, the reaction was maximized to ensure that the raw materials could react completely. After the addition was complete, heating and stirring were continued. The reaction lasted for about 4 hours or more, and the reaction progress was monitored using thin-layer chromatography (TLC) until the reaction was completed. After the reaction was completed, the reaction mixture was poured into water to quench the reaction and allowed to stand to room temperature. Subsequently, an appropriate amount of ethyl acetate was added to it to extract the generated organic compounds. After collecting the organic phase, it was also necessary to wash it with saturated brine to remove impurities, extract it again, collect the organic layer, add an appropriate amount of anhydrous Na2SO4 to it, and let it stand for 30 minutes to remove moisture. Next, the organic phase containing the target compound is obtained by vacuum filtration, and then the organic phase is concentrated under vacuum to obtain a white or yellow solid. This organic layer is Compound III and is refrigerated for future use. The yield is approximately 80-90%.

[0054] In step b, the molar ratio of compound III to compound IV is (1.2-1.8):1, for example, 1.2:1, 1.3:1, 1.4:1, 1.5:1, 1.6:1, 1.7:1, 1.8:1, and any other value between (1.2-1.8):1. The reaction temperature is 70-85°C, for example, 70°C, 75°C, 80°C, and 85°C. The reaction in step b is catalyzed using an acid such as concentrated hydrochloric acid.

[0055] Specifically, an appropriate amount of anhydrous isopropanol is used to dissolve the compound III obtained by the reaction, 1.5 mmol of 4-(4-R1piperazine-1-yl)aniline is added, heated to 80 ° C and stirred to dissolve, and then about 1 mL of concentrated hydrochloric acid is added dropwise to catalyze the reaction. It lasts for about 48 hours or more. In order to monitor the progress of the reaction until it ends, thin layer chromatography (TLC) is used. After the reaction is complete, the reaction solution is cooled to room temperature and vacuum filtered to obtain the precipitate produced after the reaction. After adding a small amount of water to dissolve the precipitate, an appropriate amount of NaOH is added to adjust the pH to 12. It is found that a precipitate is precipitated. After vacuum filtering the precipitate, it is washed with a small amount of anhydrous methanol to obtain a white or yellow precipitate. The precipitate is collected by suction and separated by silica gel column chromatography. The yield is about 60-70%.

[0056] In a third aspect, the present invention provides a use of the compound capable of enhancing the permeability of the blood-brain barrier as described in the aforementioned embodiment in the preparation of an inhibitor for inhibiting ALK and / or EGFR.

[0057] In a fourth aspect, the present invention provides a use of the compound capable of enhancing the permeability of the blood-brain barrier as described in the aforementioned embodiment in the preparation of a drug for treating cancer, wherein the cancer includes non-small cell lung cancer, anaplastic lymphoma, lung adenocarcinoma and glioma.

[0058] The features and performance of the present invention are further described in detail below with reference to the embodiments.

[0059] Example 1

[0060] The present invention provides a compound (numbered AEB-1) that can enhance the permeability of the blood-brain barrier, and its structural formula is shown below:

[0061] This embodiment also provides a method for preparing the compound capable of enhancing blood-brain barrier permeability, comprising:

[0062] Weigh 1 mmol of p-nitroaniline and dissolve it in approximately 5 mL of anhydrous isopropyl alcohol (add isopropyl alcohol to 3A molecular sieves and let stand for 24 hours to remove water). Add 3 mmol of the acid-binding agent N,N-diisopropylethylamine to the dissolved p-nitroaminobenzene. Heat and stir at 80°C for 4 hours. After ensuring the mixture is homogeneous, slowly add 1.15 mmol of 2,4,5-trichloropyrimidine dropwise. This is when the reaction is maximized to ensure complete reaction of the raw materials. Continue heating and stirring after the addition is complete. Continue for approximately 4 hours or longer, monitoring the reaction progress using thin-layer chromatography (TLC) until the reaction is complete. After the reaction is complete, quench the reaction by immersing the reaction mixture in water and allow it to cool to room temperature. Subsequently, add an appropriate amount of ethyl acetate to extract the generated organic compounds. After collecting the organic phase, wash it with saturated brine to remove impurities, extract it again, collect the organic layer, add an appropriate amount of anhydrous Na2SO4, and let it stand for 30 minutes to remove moisture. Next, the organic phase containing the target compound is obtained by vacuum filtration, and then the organic phase is concentrated under vacuum to obtain a white or yellow solid. This organic layer is compound III and is refrigerated for later use. The yield is 80-90%.

[0063] The compound III obtained by the reaction was dissolved in an appropriate amount of anhydrous isopropanol, 1.5 mmol of 4-(4-ethylpiperazine-1-yl)aniline was added, and the mixture was heated to 80°C and stirred to dissolve. Then, about 1 mL of concentrated hydrochloric acid was added dropwise to catalyze the reaction. The reaction lasted for about 48 hours or more. In order to monitor the progress of the reaction until its completion, thin layer chromatography (TLC) was used. After the reaction was complete, the reaction solution was cooled to room temperature and vacuum filtered to obtain the precipitate produced after the reaction. A small amount of water was added to dissolve the precipitate, and then an appropriate amount of NaOH was added to adjust the pH to 12. A precipitate was found to be precipitated. The precipitate was vacuum filtered and washed with a small amount of anhydrous methanol to obtain a white or yellow precipitate. The precipitate was collected by suction and separated by silica gel column chromatography. The yield was 60-70%.

[0064] AEB-1 characterization diagram can be found in Figure 1 、 Figure 2 and Figure 3 , the data is as follows: 1 H NMR(400MHz,DMSO-d6)δ8.22(s,1H),8.17–8.04(m,8H),7.58–7.50(m,4H),6.9 1–6.84(m,2H),2.37(q,J=7.2Hz,2H),1.04(t,J=7.2Hz,3H).HRMS(ESI):calcd for C 22 H 24 ClN7O2:453.1680; found:453.1681.

[0065] Example 2-28

[0066] Examples 2-28 were prepared by referring to the preparation method of Example 1, with the only difference being the change of the corresponding raw materials. Specifically,

[0067] Example 2: AEB-2, characterization data are as follows: 1 H NMR(400MHz,DMSO-d6)δ8.95(s,1H),7.89(s,1H),7.84–7.72(m,2H),7.57–7.49(m, 2H),7.44–7.37(m,2H),7.31(s,2H),7.24(t,J=5.8Hz,1H),6.85–6.76(m,2H),3.67 –3.61(m,1H),3.64–3.55(m,1H),3.05(dd,J=10.3,5.5Hz,4H),2.96(dd,J=8.9,6.3 Hz,2H),2.49(s,1H),2.37(q,J=7.2Hz,2H),1.04(t,J=7.2Hz,3H).HRMS(ESI):calcd for C 24H 30 ClN7O2S:515.1870; found:515.1868.

[0068] Example 3: AEB-3: 1 H NMR(400MHz, DMSO-d6)δ8.94(d,J=27.3Hz,1H),8.02–6.72(m,4H),2.44–2.21(m,3H),2.16–1.32(m,5H),1.06–0.98(m,3H).HRMS(ESI):calcd forC 23 H 34 ClN7:443.2564; found:443.2560.

[0069] Example 4: AEB-4: 1 H NMR(400MHz,DMSO-d6)δ9.19(s,1H),8.81(s,1H),8.12(s,1H),7.67–7.21(m,4H),6.96–6.71(m ,3H),5.76(s,2H),3.72(s,3H),2.36(q,J=7.2Hz,2H),1.02(t,J=7.2Hz,3H).HRMS(ESI):calcd for C 23 H 26 BrClN6O:516.1040; found:516.1033.

[0070] Example 5: AEB-5: 1 H NMR (400MHz, DMSO-d6) δ8.68(d,J=21.3Hz,1H),8.37(s,0H),7.69(dd,J=50.7,2.0Hz,1H),7.21(dd,J=71.7,8.9Hz,2H),6.96–6.37(m ,3H),2.90(q,J=4.2Hz,2H),2.13(h,J=6.9Hz,2H),0.83–0.72(m,4H),0.18(d,J=7.8Hz,1H),0.02(d,J=4.9Hz,1H).HRMS(ESI):calcd for C 20 H 27 ClN6:386.1986; found:386.1981.

[0071] Example 6: AEB-6: 1H NMR (400MHz, DMSO-d6) δ9.17(s,1H),8.84(s,1H),8.13(s,1H),7.75(d,J=11.8Hz,1H),7.55(d,J=8.2Hz,1H),7.44(d,J=8 .7Hz,2H),7.33(td,J=8.3,6.8Hz,1H),6.97–6.77(m,3H),2.36(q,J=7.2Hz,2H),1.03(t,J=7.1Hz,3H).HRMS(ESI):calcd for C 19 H 27 BrClN6O2:406.1884; found:406.1886.

[0072] Example 7: AEB-7: 1 H NMR (400MHz, DMSO-d6) δ9.02(s,1H),8.77(d,J=3.2Hz,1H),8.07(d,J=3.1Hz,1H),7.56–7.31(m,2H),7.32–7.20(m,2H),7.14(td,J=8.6 ,3.3Hz,1H),6.95(dd,J=9.3,3.1Hz,0H),6.73–6.53(m,2H),2.36(qt,J=7.3,4.0Hz,2H),1.03(tt,J=7.2,3.7Hz,3H).HRMS(ESI):calcd for C 22 H 23 ClF2N6:444.1641; found:444.1633.

[0073] Example 8: AEB-8: 1 H NMR (400MHz, DMSO-d6) δ9.38(d,J=2.2Hz,1H),8.30(d,J=2.3Hz,1H),7.39–7.33(m,2H),6. 98–6.92(m,2H),2.39(qd,J=7.1,2.3Hz,4H),1.05(td,J=7.2,2.3Hz,3H).HRMS(ESI):calcd for C 18 H 22 ClF3N6:414.1547; found:414.1541.

[0074] Example 9: AEB-9: 1H NMR (400MHz, DMSO-d6) δ9.17(s,1H),8.84(s,1H),8.13(s,1H),7.75(d,J=11.8Hz,1H),7.55(d,J=8.2Hz,1H),7.44(d,J=8 .7Hz,2H),7.33(td,J=8.3,6.8Hz,1H),6.97–6.77(m,3H),2.36(q,J=7.2Hz,2H),1.03(t,J=7.1Hz,3H).HRMS(ESI):calcd for C 22 H 24 ClFN6:426.1735; found:426.1733.

[0075] Example 10: AEB-10: 1 H NMR (400MHz, DMSO-d6) δ9.18(s,1H),8.94(s,1H),8.18–8.01(m,2H),7.72(d,J=8.9Hz,1H),7.54(d,J=8.8Hz, 1H),7.38(d,J=8.6Hz,2H),6.89–6.79(m,2H),2.37(q,J=7.2Hz,2H),1.03(t,J=7.2Hz,3H).HRMS(ESI):calcd forC 22 H 23 BrCl3N6:476.1050; found:476.1044.

[0076] Example 11: AEB-11: 1 H NMR(400MHz, DMSO-d6)δ8.91(s,1H),7.90(s,1H),7.74(t,J=6.2Hz,1H),7.40(d,J=8.6Hz,2H),7.35–7.27(m,4H), 7.27–7.17(m,1H),6.81–6.72(m,2H),4.61(d,J=6.1Hz,2H),2.45(s,1H),1.06(t,J=7.2Hz,3H).HRMS(ESI):calcd forC 23 H 27 ClN6:422.1986; found:422.1985.

[0077] Example 12: AEB-12: 1H NMR (400MHz, DMSO-d6) δ9.01(s,1H),8.65(s,1H),8.05(s,1H),7.54(d,J=8.1Hz,2H),7.42(d,J=8.8Hz,2H),7.15 (d,J=8.1Hz,2H),6.83–6.72(m,2H),2.35(t,J=7.2Hz,2H),2.31(s,3H),1.03(t,J=7.1Hz,3H).HRMS(ESI):calcd forC 23 H 27 ClN6:422.1986; found:422.1983.

[0078] Example 13: AEB-13: 1 H NMR (400MHz, DMSO-d6) δ8.10–7.82(m,1H),7.74–7.24(m,1H),7.08–6.78(m,1H),6.67(d,J=8.5Hz,1H),6.49(d,J=8.6 Hz,1H),3.14–2.79(m,4H),2.68(t,J=4.8Hz,1H),2.47–2.21(m,5H),1.02(h,J=10.7,9.1Hz,3H).HRMS(ESI):calcdfor C 22 H 33 ClN8:444.2517; found:444.2515.

[0079] Example 14: AEB-14: 1 H NMR(400MHz,DMSO-d6)δ9.19(s,1H),8.81(s,1H),8.12(s,1H),7.73–7.06(m,4H),6.98–6.63(m ,3H),5.76(s,2H),3.72(s,3H),2.36(q,J=7.2Hz,2H),1.02(t,J=7.2Hz,3H).HRMS(ESI):calcd for C 22 H 32 ClN7O:445.2357; found:445.2363.

[0080] Example 15: AEB-15: 1H NMR(400MHz, DMSO-d6)δ8.90(s,1H),7.85(s,1H),7.62–7.50(m,2H),7.10(t,J=5.8Hz,1H),6.91–6.79(m,2H),3.03(dd,J=6.3,3.6Hz,3H ),2.36(qd,J=7.1,2.0Hz,2H),1.57(t,J=7.3Hz,2H),1.36–1.20(m,6H),1.03(td,J=7.2,2.1Hz,3H),0.90–0.82(m,3H).HRMS(ESI):calcd for C 22 H 33 ClN6:416.2455; found:416.2459. Characterization spectrum see Figure 4 、 Figure 5 and Figure 6 .

[0081] Example 16: AEB-16: 1 H NMR (400MHz, DMSO-d6) δ9.03(s,1H),8.75(s,1H),8.07(s,1H),7.62(d,J=8.4Hz,2H),7.39(d,J=8.6Hz,2H),7 .29–7.18(m,2H),6.87–6.69(m,2H),5.75(s,1H),2.49–2.33(m,8H),1.03(t,J=7.1Hz,3H).HRMS(ESI):calcd forC 23 H 27 BrClN6S:454.1706; found:454.1709.

[0082] Example 17: AEB-17: 1 H NMR (400MHz, DMSO-d6) δ8.90(s,1H),7.85(s,1H),7.60–7.49(m,2H),7.14(t,J=5.9Hz,1H),6.87–6.78(m,2H),3.20(t, J=6.5Hz,2H),2.36(q,J=7.1Hz,2H),2.02–1.92(m,1H),1.03(t,J=7.2Hz,3H),0.90(d,J=6.7Hz,5H).HRMS(ESI):calcd for C 20 H 29 ClN6:388.2142; found:388.2137.

[0083] Example 18: AEB-18:1 H NMR(400MHz,DMSO-d6)δ9.03(s,1H),7.93(s,1H),7.69–7.45(m,3H),6.93–6.70(m,2H),4.13(dd,J =5.8,2.4Hz,2H),3.09(t,J=2.4Hz,1H),2.37(q,J=7.2Hz,2H),1.08–1.01(m,3H).HRMS(ESI):calcd for C 19 H 23 ClN6:370.1673; found:370.1670.

[0084] Example 19: AEB-19: 1 H NMR (400MHz, DMSO-d6) δ9.10 (s, 1H), 8.62 (s, 1H), 8.09 (d, J = 5.2Hz, 1H), 7.46 (d, J = 8.8Hz, 2H), 7.01–6.68(m,4H),6.27(s,1H),3.69(d,J=5.2Hz,6H),1.04(t,J=6.9Hz,3H).HRMS(ESI):calcd for C 24 H 29 ClN6O2:468.2041; found:468.2041.

[0085] Example 20: AEB-20: 1 H NMR(400MHz, DMSO-d6)δ8.88(d,J=115.1Hz,2H),8.07(s,1H),7.70–7.06(m,5H),6. 67(d,J=8.6Hz,2H),2.36(q,J=7.2Hz,2H),1.03(t,J=7.1Hz,3H).HRMS(ESI):calcd for C 22 H 24 ClFN6:426.1735; found:426.1737.

[0086] Example 21: AEB-21: 1H NMR (400MHz, DMSO-d6) δ8.90(s,1H),7.85(s,1H),7.66–7.44(m,2H),7.08(t,J=5.8Hz,1H),6.91–6.73(m,2H),3.18(d,J=5.1Hz, 2H),2.36(q,J=7.2Hz,2H),1.65–1.48(m,2H),1.44–1.26(m,2H),1.03(t,J=7.2Hz,3H),0.91(t,J=7.3Hz,3H).HRMS(ESI):calcd for C 20 H 29 ClN6:388.2142; found:388.2138.

[0087] Example 22: AEB-22: 1 H NMR(400MHz, DMSO-d6)δ8.99(s,1H),7.96(s,1H),7.49(d,J=8.4Hz,1H),7.42–7.19(m,12H),6.91–6.73( m,2H),6.54(d,J=8.3Hz,1H),5.76(s,1H),2.36(q,J=7.2Hz,2H),1.03(t,J=7.2Hz,3H).HRMS(ESI):calcd for C 29 H 31 ClN6:498.2299; found:498.2298.

[0088] Example 23: AEB-23: 1 H NMR (400MHz, DMSO-d6) δ8.98(d,J=25.9Hz,1H),8.66(d,J=31.9Hz,1H),8.02(d,J=21.1Hz,0H),7.60–7.10(m,2H),6.93(d,J=9.0Hz, 1H),6.72(d,J=8.9Hz,1H),3.02(t,J=4.9Hz,2H),2.37(p,J=7.4Hz,2H),1.34–1.11(m,4H),1.04(q,J=7.2Hz,3H).HRMS(ESI):calcd for C 25 H 31 ClN6:450.2299; found:450.2302.

[0089] Example 24: AEB-24: 1H NMR (400MHz, DMSO-d6) δ9.37(s,1H),8.90(s,1H),7.88(s,1H),7.65–7.34(m,3H),7.29–7.03(m,2H),6.94–6. 57(m,4H),5.75(s,1H),4.49(d,J=6.1Hz,2H),2.38(q,J=7.2Hz,2H),1.03(t,J=7.2Hz,3H).HRMS(ESI):calcd forC 23 H 27 ClN6O:438.1935; found:438.1931.

[0090] Example 25: AEB-25: 1 H NMR (400MHz, DMSO-d6) δ8.92(s,1H),7.87(s,1H),7.58–7.50(m,2H),6.89–6.79(m,3H),3.48(d,J=6.3Hz,2H),3.07–3.00(m ,4H),2.55(s,1H),2.46(dd,J=15.5,5.9Hz,6H),2.36(q,J=7.1Hz,2H),2.19(s,6H),1.03(t,J=7.1Hz,3H).HRMS(ESI):calcd for C 20 H 30 ClN7:403.2251; found:403.2251.

[0091] Example 26: AEB-26: 1 H NMR(400MHz, DMSO-d6)δ9.01(s,1H),8.71(s,1H),8.04(s,1H),7.42(dd,J=35.8,8.3Hz,4H),7.27–7.07(m,2H),6.88–6.61(m,2 H),3.17(d,J=5.2Hz,1H),2.37(q,J=7.2Hz,2H),2.11–1.64(m,5H),1.60–1.12(m,5H),1.03(t,J=7.2Hz,3H).HRMS(ESI):calcd for C 28 H 35 ClN6:490.2612; found:490.2607.

[0092] Example 27: AEB-27: 1H NMR (400MHz, DMSO-d6) δ8.91(s,1H),7.91(s,1H),7.76(t,J=6.1Hz,1H),7.56–7.44(m,2H),7.32(dd,J=30.3,8.5 Hz,4H),6.83–6.69(m,2H),4.55(d,J=6.1Hz,2H),2.36(q,J=7.2Hz,2H),1.03(t,J=7.1Hz,3H).HRMS(ESI):calcd forC 23 H 26 BrClN6:500.1091; found:500.1096.

[0093] Example 28: AEB-28: 1 H NMR (400MHz, DMSO-d6) δ9.03(s,1H),8.66(s,1H),8.06(s,1H),7.50–7.34(m,2H),7.29–7.16(m,2H),7.07(d,J=8.0 Hz,1H),6.86–6.67(m,2H),3.68(s,3H),2.40(q,J=7.2Hz,2H),2.15(s,3H),1.04(t,J=7.2Hz,3H).HRMS(ESI):calcd forC 24 H 29 ClN6O:452.2091; found:452.2087. Characterization spectrum see Figure 7 、 Figure 8 and Figure 9 .

[0094] Experimental Example 1 - Blood-brain barrier permeability study

[0095] Experimental methods

[0096] 1. Determination of the standard curve

[0097] To prepare a standard solution of the compound to be tested, first weigh an appropriate amount of the compound and dissolve it in an appropriate amount of DMSO to form a 10mM stock solution. In this experiment, seven compounds, AEB-1, AEB-9, AEB-12, AEB-16, AEB-21, and AEB-28, were selected as the drugs to be tested, and positive control drugs brigatinib and osimertinib, as well as sodium fluorescein for detecting the integrity of artificial membranes, were added. Next, each stock solution was diluted to five concentrations of 0, 5μM, 10μM, 20μM, and 40μM using PBS. Subsequently, each compound was scanned using an ultraviolet spectrophotometer to determine the maximum absorption wavelength of each compound, and the scanning range was set between 190nm and 800nm. The absorbance of the five concentrations of standard solutions at the maximum absorption wavelength was measured (each concentration was repeated three times), the data was recorded, and a standard curve was generated using Excel.

[0098] 2. Construction of a blood-brain barrier-specific artificial membrane

[0099] A solution containing 16 mg of polar brain extract (Brain Extract Polar) and 8 mg of cholesterol was dissolved in 600 μL of dodecane and sonicated until completely dissolved to form a brain-specific lipid solution. This solution was then added dropwise to the organic porous membrane of a 96-well filter plate at a volume of 5 μL per well and allowed to stand for half an hour to form a uniform lipid layer on the membrane surface.

[0100] 3. Artificial membrane integrity detection

[0101] Add 200 μL of sodium fluorescein solution to each well of the supply plate and 300 μL of blank PBS buffer to each well of the receiving plate, and repeat three times. Subsequently, place the supply plate on the receiving plate, ensure that the PBS is in full contact with the artificial membrane, and cover the cover plate. Incubate the assembled plate at 37°C for 4 hours. After the incubation is completed, carefully separate the receiving plate and the supply plate, use a microplate reader to test the absorbance of sodium fluorescein at the maximum wavelength of 490 nm, and calculate the concentration of sodium fluorescein based on the standard curve. Calculate the apparent permeability Papp of sodium fluorescein, see the following formula. If the calculated Papp is lower than 0.6×10-6cm·s -1 , it can be inferred that the phospholipid-coated artificial membrane prepared by the above method has integrity.

[0102]

[0103] Where, ΔN / Δt: permeation rate (mMol·s -1 ); S: effective area of ​​filter membrane (0.28cm 2 ); C0: initial concentration of fluorescein sodium (mMol·mL -1 ).

[0104] 4. Parallel Artificial Membrane-Blood-Brain Barrier Permeability Evaluation Experiment

[0105] First, take an appropriate amount of compound stock solution and dilute the stock solution to a concentration of 40 μM with PBS as the supply solution. Add 200 μL of supply solution to each well of the supply plate, and add 300 μL of blank PBS to the receiving plate as the receiving solution. Then, combine the receiving plate, filter plate, supply plate and cover plate in order and incubate them at 37°C for 4 hours to simulate the process of the compound passing through the blood-brain barrier. After the incubation, the system is separated, and the supply solution and receiving solution are collected into 96-well plates respectively. The absorbance is measured at the maximum absorption wavelength of each compound using an enzyme-linked microplate reader, and repeated three times each time to ensure the reliability of the data. Finally, record the data and calculate the concentration of each compound supply solution and receiving solution using the standard curve.

[0106] 5. Calculation of effective permeability coefficient logPe of each compound

[0107] Substitute the measured concentration into the formula to calculate Pe, see the following formula.

[0108]

[0109]

[0110] Among them, V A :The amount of liquid added to each well of the collection plate (0.30cm 3 );V D :Liquid addition amount per well of supply plate (0.2cm 3 ); S is the effective area of ​​the artificial membrane (0.28 cm 2 ) ; t: incubation time, which is 14400s in this paper; C A (t): drug concentration in the collection plate after incubation; C e : Theoretical equilibrium concentration after incubation is completed; C D (t): Drug concentration in the feeding plate after incubation.

[0111] Experimental results

[0112] The results of the blood-brain barrier permeability of the compound PAMPA-BBB are shown in Table 1 below.

[0113] Table 1 PAMPA test results of compounds

[0114]

[0115] According to Table 1 above, the Papp of fluorescein sodium after 4 h is 0.054×10 -6, indicating that the parallel artificial membrane established in this experiment has good integrity. The logPe of the positive drug osimertinib is the highest, while the effective permeability logPe of compound AEB-9 is -4.06, which is close to that of osimertinib. The logPe of compound AEB-12 is -4.38, higher than that of brigatinib. According to the classification scheme reported in the literature, using the calculated experimental permeability (logPe), molecules can be divided into four groups: impermeable compounds (logPe < -6.14), low permeability compounds (-6.14 < logPe < -5.66), medium permeability compounds (-5.66 < logPe < -5.33), and high permeability compounds (logPe > -5.33). Based on these critical values, it can be concluded that all tested AEB compounds show high blood-brain barrier permeability.

[0116] Experimental Example 2 - In Vitro Kinase Activity

[0117] (1) ALK Kinase Activity Test Experiment

[0118] 1. Prepare 1-fold kinase base buffer

[0119] 50 mM HEPES buffer (pH 7.5), 10 mM MgCl2, 2 mM DTT, 0.01% BSA, 0.01% Tween-20.

[0120] 2. Prepare compounds

[0121] Dilute the compounds to 100 times the highest required inhibitor concentration using dimethyl sulfoxide (DMSO) with a purity of 100%. To set up control experiments, select two empty wells in the same 96-well plate and inject 100 μL of 100% DMSO solution into each of them. These two wells serve as the compound-free control and the enzyme-free control, respectively. Then label this plate containing the controls and name it the source plate. Subsequently, accurately pipette 40 μL of the compounds from the source plate and transfer them to a new 384-well Echo plate, which is used as the intermediate plate. Immediately, transfer the 200 μL of sample in each well of the intermediate plate to a 384-well detection plate.

[0122] 3. Kinase Reaction and Detection

[0123] Add 10 μL of kinase solution to each well of the assay plate, and 10 μL of 1x kinase buffer to the control wells without enzyme. Next, prepare a 2x substrate solution (by adding substrate and ATP to 1x kinase base buffer). Finally, add 10 μL of 2x substrate solution to each well to initiate the reaction between the kinase and substrate. Incubate the reaction at room temperature and prepare the detection solution at a final concentration of 2x using antibody dilution buffer. Stop the reaction by adding 20 μL of detection solution to each well of the assay plate and incubate at room temperature for a period of time.

[0124] 4. Curve Fitting

[0125] The Lance signal ratio (665 nm / 615 nm) was copied from the Envision program. The ratio was converted to an inhibition percentage (inhibition percentage = (maximum value - sample Lance signal ratio) / (maximum value - minimum value) * 100). The data were displayed in MS Excel, and the curve was fitted using XLFit Excel add-in version 5.4.0.8.

[0126] The results are shown in Table 2 below.

[0127] Table 2 Inhibition rate of AEB series compounds on ALK at 10 μM

[0128]

[0129]

[0130] As shown in Table 2, the tested compounds have good targeting properties for ALK. The kinase activity of compounds AEB-1, AEB-9, AEB-12, AEB-16, and AEB-28 against ALK is close to or even exceeds that of the positive drug brigatinib.

[0131] (2) EGFR kinase activity test

[0132] 1. Prepare 1x Kinase Buffer

[0133] 1x kinase buffer, 50 mM HEPES buffer (pH 7.5), 10 mM MgCl2, 2 mM DTT, 0.01% Tween-20, 0.01% BSA.

[0134] 2. Preparation of Compounds

[0135] Using 100% pure DMSO as a solvent, the compound was diluted until it reached 100 times the desired maximum inhibitor concentration. For a 10μM test compound, a 1000μM compound DMSO solution was prepared. Subsequently, two specific empty wells were selected in the same 96-well plate, marked as the no-compound control well and the no-enzyme control well, and 100μL of 100% DMSO was added to each of these two wells. This plate was named the source plate. Then, 40μL of the compound solution was removed from the source plate and transferred to a new 384-well Echo plate, marked as the intermediate plate.

[0136] 3. Preparation of Assay Plates

[0137] Repeatedly transfer 200 nL of sample from each well of the 384-well Echo Plate to the 384-well assay plate. For example, transfer the sample from A1 of the 384-well Echo Plate to A1 and A2 of the 384-well assay plate. Next, transfer the sample from A2 of the 384-well Echo Plate to A3 and A4 of the 384-well assay plate, and so on.

[0138] 4. Kinase reaction

[0139] Prepare 2x Kinase Solution: Prepare a kinase solution in 1x Kinase Buffer at a concentration twice the final concentration of each reagent in the assay. Add 10 μL of kinase solution to each well of the assay plate, except for the control wells without enzyme (add 10 μL of 1x Kinase Buffer). Shake the plate. Incubate at room temperature for the desired time.

[0140] Prepare 2x substrate solution: Prepare substrate solutions of fluorescein-polyGT and ATP in 1x kinase reaction buffer at a concentration of 2x the final concentration of each reagent required in the assay. Add 10 μL of substrate solution to each well of the assay plate to start the reaction, shake the plate, cover the assay plate, and incubate at room temperature for a desired period of time.

[0141] 5. Kinase Assay

[0142] Prepare the detection solution with a final concentration of 2x using antibody dilution buffer; add 20 μL of detection solution to each well of the detection plate to stop the reaction; briefly mix by centrifuge and incubate at room temperature for a period of time, and then read the fluorescence value on a plate reader.

[0143] 6. Data reading

[0144] Data were collected on an Envision with the excitation wavelength set to 340 nm and the emission wavelengths set to 520 nm and 495 nm, respectively.

[0145] 7. Curve Fitting

[0146] The RFU values ​​were copied from the Envision program, and the ratio of RFU 520 nm / RFU 495 nm was calculated. The ratio was converted to an inhibition percentage (inhibition percentage = (maximum - sample) / (maximum - minimum) * 100). The data were displayed in MS Excel, and the curves were fitted using XLFit excel add-in version 5.4.0.8.

[0147] The results are shown in Table 3 below.

[0148] Table 3 Inhibition rate of AEB series compounds on EGFR at 10 μM

[0149]

[0150]

[0151] As shown in Table 3, the tested compounds have good targeting properties to EGFR. The kinase activity of AEB-1, AEB-4, AEB-9, AEB-15, AEB-16, AEB-19, AEB-21, AEB-23, and AEB-28 against EGFR exceeds that of the positive drugs brigatinib and osimertinib.

[0152] In summary, the compounds provided in the embodiments of the present invention have good targeting properties to ALK and EGFR, and the inhibition rates of the 12 compounds to ALK and EGFR kinases are over 90% at a concentration of 10 μM.

[0153] Experimental Example 3—IV. In vitro anti-tumor cell proliferation activity test

[0154] A549, NCI-H1975, and U251 cells are all adherent cells, PC-9-LRTM cells are semi-adherent and suspension cells, and Karpas-299 cells are suspension cells. The culture medium required for NCI-H1975, PC-9-LRTM, and Karpas-299 cells is RPIM1640 complete medium, while the culture medium required for A549 and U251 cells is DMEM complete medium. The culture methods for adherent, semi-adherent, and suspension cells differ slightly.

[0155] 1. Cell Recovery

[0156] The six cell recovery methods are consistent. After removing the cell cryovials stored in a -80°C cryopreservation box or liquid nitrogen tank, quickly place the sample in a 37°C water bath and continue to gently shake in the water bath until the cells are completely thawed and melted. The thawed cell suspension is then thoroughly mixed with 5mL of complete culture medium (DMEM / RPIM1640). Then centrifuge at 1200 rpm for 3 minutes. After centrifugation, carefully remove the supernatant and discard it, and then add 1mL of complete culture medium. After gently resuspending the cells by pipetting, transfer them to a culture dish with 9mL of fresh complete culture medium. Move the culture dish horizontally in four directions, up, down, left, and right, in a cross shape, and repeat 5-6 times to shake the culture dish back and forth to evenly distribute the cells. Finally, place the culture dish in an incubator with culture conditions of 37°C and 5% CO2.

[0157] 2. Cell Medium Replacement

[0158] Adherent cells: Pour or aspirate the old culture medium from the dish and gently rinse the cells with 2 mL of PBS. Repeat twice. Add 10 mL of fresh culture medium and shake gently to distribute the new medium evenly around the cells. Return the dish to the incubator and continue culturing.

[0159] For semi-suspended, semi-adherent cells: Collect the supernatant from the culture dish into a 15mL centrifuge tube. Gently rinse the cells with 2mL of PBS twice. Aspirate the PBS and add 1mL of trypsin to digest the cells. Observe under a microscope to see if the cells are slowly detaching. Once digestion is complete, quickly add 2mL of fresh complete medium to stop the digestion. Gently pipette the cells and transfer them to a 15mL centrifuge tube. Centrifuge at 1200 rpm for 3 minutes. After centrifugation, pour off the supernatant and resuspend the cells in 1mL of complete medium. Transfer 1mL of the cell suspension to a culture dish containing 9mL of fresh complete medium. Use the "cross method" to evenly distribute the cells. Finally, place the culture dish in an incubator at 37°C, 5% CO2 and continue incubation.

[0160] Suspension cells: Collect the supernatant from the culture dish into a 15mL centrifuge tube and centrifuge at 1200 rpm for 3 minutes. After centrifugation, pour off the supernatant and resuspend the cells in 1mL of complete medium. Gently pipette to resuspend the cells and transfer them to a culture dish containing 9mL of fresh complete medium. Use the "cross technique" to evenly distribute the cells. Finally, place the culture dish in an incubator at 37°C and 5% CO2 and continue incubation.

[0161] 3. Cell Passaging

[0162] Adherent cells: Pour out or aspirate the old culture medium in the culture dish, add 2mL PBS washing solution to gently rinse the cells, repeat twice, add 1mL trypsin to digest, observe under a microscope, and after digestion, add 2mL complete culture medium to terminate digestion. Transfer the cells to a 15mL centrifuge tube by pipetting, centrifuge at 1200rpm for 3 minutes, pour out the supernatant, and resuspend the cell clusters with a certain amount of fresh culture medium. Then aspirate an appropriate amount of the cell resuspension into a culture dish containing 9mL fresh complete culture medium, and evenly distribute the cells using the "cross method". Return to the incubator for further culture.

[0163] Semi-suspended and semi-adherent cells: Aspirate the old culture medium into a 15 mL centrifuge tube and centrifuge together. Other operations are the same as for adherent cells.

[0164] Suspending cells: Collect the supernatant from the culture dish into a 15mL centrifuge tube and centrifuge at 1200 rpm for 3 minutes. After centrifugation, pour off the supernatant and resuspend the cells in 1mL of complete medium. Gently pipette to resuspend the cells. Transfer an appropriate amount of the cell suspension to a culture dish containing 9mL of fresh complete medium. Use the "cross technique" to evenly distribute the cells. Finally, place the culture dish in an incubator at 37°C, 5% CO2 and continue incubation.

[0165] 4. Cell Seeding

[0166] Prepare a cell suspension according to the "Cell Passaging" procedure, mix well, and count the cells. Subsequently, dilute the cell suspension to the desired concentration and evenly inoculate into a 96-well plate, adding 100 μL per well. Depending on the cell type, the inoculation density is 1500-3000 cells per well. To ensure the accuracy of the experiment, add 100 μL of PBS to the edge of the 96-well plate to prevent evaporation of the culture medium. After completing the cell inoculation, place the 96-well plate in an incubator. If the cells are adherent, continue to culture them until the cells attach to the bottom and set aside.

[0167] 5. Cellular Administration

[0168] Weigh an appropriate amount of compound and prepare a 40mM stock solution with DMSO. Dilute the appropriate amount of stock solution with DMSO to 20mM, 10mM, 5mM, 2.5mM, 1.25mM, and 0.625mM, respectively. Add 1μL of the compound stock solution to 999μL of complete culture medium and dilute to a concentration gradient of 40μM, 20μM, 10μM, 5μM, 2.5μM, 1.25μM, and 0.625μM. When adding drugs to adherent cells, aspirate the old medium and add 100μL of drug-containing medium. For semi-suspended, semi-adherent, or adherent cells, do not aspirate the old medium and add 100μL of drug-containing medium at twice the concentration to be tested. Mix thoroughly by pipetting. Each compound and each concentration were added to three replicate wells, and each plate was set up with a control group (with cells, culture medium, and no drug) and a blank group (without cells, no drug, and only culture medium). After adding the drug, the 96-well plate was returned to the cell culture incubator for continued culture.

[0169] 6.CCK-8 assay

[0170] After incubating the cells with drug-containing medium for 48 hours, add 10% CCK-8 solution to each well (10 μL for adherent cells, 20 μL for semi-adherent, semi-suspended, and suspended cells). Incubate in an incubator for 0-4 hours until the absorbance at 450 nm of the control group is between 0.8 and 1.2. Data were processed and the average of triplicate wells was used as the experimental result. The cell inhibition rate was calculated as shown in the following formula.

[0171]

[0172] Among them, OD 实验组 : absorbance value of the experimental group; OD 对照组 : absorbance value of the control group; OD 空白组 : Absorbance value of blank group.

[0173] The test results are shown in Table 4 below.

[0174] Table 4 Cell inhibition rate of the compounds determined by CCK-8 method

[0175]

[0176]

[0177] From the above in vitro anti-tumor cell proliferation activity test results in Table 4, the compounds generally have low activity against A549 cells, but have good activity against NCI-H1975 and PC-9-LRTM, indicating that the compounds are targeted against EGFR mutations. 50 =1.206 μM), AEB-9 (IC 50 =1.183 μM), AEB-12 (IC50 =1.662 μM) and AEB-19 (IC 50 =1.152 μM) in H1975 cell line, which exceeded the positive control drug brigatinib (IC 50 =1.906 μM). Similarly, these compounds and AEB-16 (IC 50 =1.947 μM) was also significantly more active against H1975 cells than another positive control drug, osimertinib (IC 50 =2.221 μM). In the test on PC-9-LRTM cell line, compound AEB-1 (IC 50 =28 nM), AEB-9 (IC 50 =245 nM) and AEB-12 (IC 50 =105nM) also showed a higher efficacy than brigatinib (IC 50 =346nM). It is worth noting that most of the tested drugs were more active than osimertinib against PC-9-LRTM cells. In addition, in the activity test of Karpas cell line, compound AEB-1 (IC 50 =1.138 μM), AEB-9 (IC 50 =1.301 μM), AEB-12 (IC 50 =1.421 μM), AEB-16 (IC 50 =1.285 μM), AEB-20 (IC 50 =1.29 μM) and AEB-28 (IC 50 =1.35 μM) showed a significant difference compared to the positive control drug brigatinib (IC 50 Most drugs have good activity against neuroblastoma U251 and have the potential to become central anti-tumor drugs.

[0178] The foregoing description is merely a preferred embodiment of the present invention and is not intended to limit the present invention. Those skilled in the art will readily appreciate that various modifications and variations of the present invention are possible. Any modifications, equivalent substitutions, or improvements made within the spirit and principles of the present invention are intended to be within the scope of protection of the present invention.

Claims

1. A compound capable of increasing the permeability of the blood-brain barrier, characterized in that: It is selected from any one of the compounds represented by the following structural formulas: 。 2. Use of the compound capable of increasing blood-brain barrier permeability according to claim 1 in the preparation of a drug for treating cancer, characterized in that: The cancer is glioma.

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