An sg2c class bromobenzyl ether derivative, preparation method and application

Abstract

The application discloses an SG2C type bromobenzyl ether derivative, a preparation method and application thereof, and SG2C-CBM is used as a ligand 99m Complex formed by Tc labeling 99m Tc-SG2C-CBM can be applied to detection of PD-L1 expression as an imaging agent, realizes real-time, comprehensive and convenient detection of the PD-L1 expression level of a tumor, and overcomes the shortcomings of an immunohistochemical method. 99m The labeling rate of Tc-SG2C-CBM is above 93%, and after being placed at room temperature for 6 hours, the radiochemical purity is still above 90%, the in-vitro stability at room temperature is good, the lipid-water partition coefficient is 1.31, the drug in a biological body can be reflected between water and lipid, and cell uptake shows that 99m The PD-L1 positive cell uptake of Tc-SG2C-CBM is 6.32 times that of the negative cell uptake.

Description

Technical Field

[0001] This invention belongs to the fields of radiopharmaceutical chemistry and clinical nuclear medicine, specifically relating to an SG2C-type brominated benzyl ether derivative, its preparation method, and its application. Background Technology

[0002] In recent years, immunotherapy has become a research hotspot in cancer treatment, with tumor cell immune escape playing a crucial role in tumor development and progression. Tumor cells bind to programmed death ligand 1 (PD-1) on T cells via PD-L1 produced on their surface, inhibiting T cell proliferation and releasing cytotoxins. This leads to the depletion of tumor-specific T cell activity and apoptosis, allowing tumor cells to escape the immune system and survive. PD-1 / PD-L1-based immunotherapy is a promising next-generation immunotherapy that aims to utilize the body's own immune system to fight tumors. By blocking the PD-1 / PD-L1 signaling pathway and inducing apoptosis, it has the potential to treat various types of tumors, and the expression level of PD-L1 is significantly correlated with the final efficacy of immunotherapy.

[0003] Currently, the expression level of PD-L1 on the tumor surface is mainly measured by immunohistochemistry, which requires the extraction of tumor tissue and is an invasive procedure. Furthermore, due to the heterogeneity of tumors, the PD-L1 level measured by immunohistochemistry cannot reflect the overall immune status of the tumor. Moreover, PD-L1 expression level is a dynamic indicator that changes with the course of tumor treatment. Therefore, real-time, comprehensive, and convenient detection of tumor PD-L1 levels is a pressing clinical need.

[0004] Molecular imaging, through molecular probes, enables qualitative or quantitative visualization and research of pathogenic molecules, disease characteristics, molecular mechanisms of disease development and progression, and disease outcome, treatment, and prognosis at the molecular level in vivo. Therefore, by utilizing molecular probes specifically targeting PD-L1, molecular imaging can continuously and comprehensively observe PD-L1 levels in tumors, overcoming the shortcomings of existing immunohistochemical methods.

[0005] Nuclear medicine imaging using radionuclide-labeled molecular probes is currently the most mature technology in molecular imaging, and it has long been used clinically for tumor diagnosis and efficacy evaluation. Radionuclide-labeled PD-L1-specific molecular probes can reflect the PD-L1 level of tumors in real time, comprehensively, and quantitatively, providing a basis for judging immunotherapy. International publication number (WO2017 / 202273A1) discloses a benzylphenyl ether derivative, its preparation method, pharmaceutical composition, and uses, which has good binding ability to PD-L1 protein. Existing technology uses radionuclide-labeled PD-L1 antibodies or partial antibodies and other macromolecules. Although the imaging results can reflect the distribution of PD-L1 in vivo, the preparation of such imaging probes is relatively expensive, and the elimination half-life in vivo is relatively long, making them not ideal imaging probes. The prior art (CN111943876B) discloses an N2S2-type brominated benzyl ether derivative. However, the labeling conditions in CN111943876B are not mild, require boiling, and have poor targeting of PD-L1. This problem needs to be solved. Summary of the Invention

[0006] To address the technical problems of existing brominated benzyl ether derivative labeling conditions being harsh, requiring boiling, and exhibiting poor targeting of PD-L1, the present invention aims to provide a suitable... 99m Tc-labeled SG2C-type brominated benzyl ether derivatives, their preparation methods and applications.

[0007] To achieve the above objectives and technical effects, the technical solution adopted by this invention is as follows:

[0008] An SG2C-type bromobenzyl ether derivative, wherein the SG2C-type bromobenzyl ether derivative is N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycylcysteine, has the following structural formula (Ⅰ):

[0009]

[0010] Furthermore, the preparation method of the SG2C-type bromobenzyl ether derivative includes the following steps:

[0011] N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]serine, Fmoc-glycine, and FMOC-thiotert-butyl-cysteine ​​were synthesized via solid-phase polypeptide synthesis. Tris(2-carboxyethyl)phosphine (TCEP) was added to remove the tert-butyl thiocysteine, yielding N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-seroylglycylglycyl(S-tert-butylthio)cysteine.

[0012] Furthermore, the solid-phase polypeptide synthesis method uses resin as a carrier to synthesize N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycyl(S-tert-butylthio)cysteine, and then uses a cleavage solution to cleave N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycyl(S-tert-butylthio)cysteine ​​from the resin. The cleavage solution is preferably a solution containing trifluoroacetic acid.

[0013] This invention also discloses the application of an SG2C-type bromobenzyl ether derivative as a ligand to form a complex with a radionuclide marker in PD-L1 imaging, wherein the radionuclide marker is... 99m Tc.

[0014] The present invention also discloses a 99m Tc-labeled SG2C-type bromobenzyl ether derivatives were prepared using SG2C-type bromobenzyl ether derivatives.

[0015] Furthermore, the aforementioned 99m The preparation method of Tc-labeled SG2C-type bromobenzyl ether derivatives includes the following steps:

[0016] Add glucohepatic acid salt, ethylenediaminetetraacetic acid salt, stannous chloride and pertechnetic acid solution to the above SG2C type bromobenzyl ether derivative solution, mix well and incubate to obtain the desired product.

[0017] Preferably, the SG2C-type brominated benzyl ether derivative is dissolved in an alcohol solvent to obtain a solution of the SG2C-type brominated benzyl ether derivative, and the incubation temperature is preferably 30-45℃, and the incubation time is preferably 60 min or more.

[0018] The present invention also discloses a 99m Application of Tc-labeled SG2C-type brominated benzyl ether derivatives as imaging agents in the detection of PD-L1 expression.

[0019] This invention also discloses a developer, for 99m Tc-labeled SG2C-type brominated benzyl ether derivatives.

[0020] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0021] This invention discloses an SG2C-type brominated benzyl ether derivative, its preparation method, and its application. The SG2C-type brominated benzyl ether derivative is: N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycylcysteine, abbreviated as SG2C-CBM, which is used as a ligand. 99m Tc-labeled complexes 99m Tc-SG2C-CBM can be used as an imaging agent to detect PD-L1 expression, enabling real-time, comprehensive, and convenient detection of PD-L1 levels in tumors, overcoming the shortcomings of immunohistochemistry. 99m The Tc-SG2C-CBM lipid-water partition coefficient is 1.31, which can reflect the distribution of drugs between the aqueous and lipid phases in vivo. Cellular uptake showed that its uptake in PD-L1 positive cells (A375-hPD-L1) (25.84%) was 6.32 times that in negative cells (A375) (4.09%). Compared with the prior art (CN111943876B A type of N2S2 brominated benzyl ether derivative, preparation method and application), the present invention provides… 99m The Tc-SG2C-CBM labeling conditions are mild and do not require boiling. The labeling rate and stability are comparable, but this invention... 99m The difference in Tc-SG2C-CBM uptake between PD-L1 positive cells (A375-hPD-L1) and negative cells (A375) was compared with that in CN111943876B. 99m Tc-N2S2-CBMBC and 99m The difference in Tc-neutral uptake in breast cancer cells (MDA-MB-231) further illustrates the significance of this invention. 99m Tc-SG2C-CBM exhibits good targeting of PD-L1. Attached Figure Description

[0022] Figure 1 This is the mass spectrum of the labeled precursor SG2C-CBM of the present invention;

[0023] Figure 2 The 1H NMR spectrum of the labeled precursor SG2C-CBM of this invention;

[0024] Figure 3 For the present invention 99m HPLC chromatogram of Tc-SG2C-CBM;

[0025] Figure 4 For the present invention 99m HPLC chromatogram of Tc-blank. Detailed Implementation

[0026] The embodiments of the present invention will be described in detail below so that the advantages and features of the present invention can be more easily understood by those skilled in the art, thereby making a clearer and more explicit definition of the scope of protection of the present invention.

[0027] A SG2C-type brominated benzyl ether derivative, named N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycylcysteine, abbreviated as SG2C-CBM, has the following structural formula (I):

[0028]

[0029] The preparation method of the SG2C-type brominated benzyl ether derivative includes the following steps:

[0030]

[0031] S1. Preparation of 2-bromo-3-phenyltoluene

[0032] Under nitrogen purging, 100-5000 mg of 2-bromo-3-iodotoluene was added to a 100 mL three-necked flask, followed by 30-70 mL of an aqueous solution containing dioxane (60%-90%) to dissolve it. After stirring until homogeneous, 50-2500 mg of phenylboronic acid, 100-8000 mg of cesium carbonate, and 10-300 mg of triphenylphosphine palladium were added sequentially. The mixture was stirred at 70-90 °C for 12-24 h, and the reaction was stopped. The mixture was cooled to room temperature, and the solvent was removed by rotary evaporation. The mixture was then extracted three times with water and ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, and the organic phase was evaporated to dryness. The solution was then passed through a silica gel column to obtain a colorless oily substance of 2-bromo-3-phenyltoluene.

[0033] S2, Preparation of 2-bromo-3-phenylbenzyl bromide

[0034] Under nitrogen purging, 100-2000 mg of 2-bromo-3-phenyltoluene obtained in step S1 was dissolved in anhydrous carbon tetrachloride in a 100 mL three-necked flask. 80-1600 mg of N-bromosuccinimide (NBS) was added with stirring. The temperature was raised to 80 °C, and 10-20 mg of benzoyl peroxide (BPO) was added in two batches (2 h apart) to react. The mixture was then cooled to room temperature, extracted with water and dichloromethane, and the organic phases were combined and dried with anhydrous sodium sulfate. The organic phase was evaporated to dryness to obtain a yellow oily substance of 2-bromo-3-phenylbenzyl bromide, which was directly used in the next reaction.

[0035] S3. Preparation of 2-hydroxy-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzaldehyde

[0036] Under nitrogen purging, 100-1000 mg of 2,4-dihydroxy-5-chlorobenzaldehyde was added to a 100 mL three-necked flask, dissolved in acetonitrile, and 150-1000 mg of sodium bicarbonate was added. After stirring at room temperature, an acetonitrile solution of 150-1500 mg of 2-bromo-3-phenylbenzyl bromide obtained in step S2 was added dropwise. The mixture was heated to 75 °C and stirred overnight, then cooled to room temperature, filtered, and the filtrate was washed with water. The aqueous solution was then extracted twice with ethyl acetate, the organic phases were combined, dried over anhydrous Na2SO4, the solvent was evaporated, and the mixture was recrystallized from ethyl acetate and petroleum ether to give 2-hydroxy-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzaldehyde as a pale yellow solid.

[0037] S4. Preparation of 2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzaldehyde

[0038] Under nitrogen purging, 100-1000 mg of 2-hydroxy-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzaldehyde was dissolved in 3-8 mL of DMF and mixed with 100-600 mg of cesium carbonate at room temperature. Then, 3-5 mL of 80-800 mg of 3-cyanobenzyl bromide in DMF solution was added dropwise. After the addition was complete, the temperature was raised to 85 °C and stirred for 12-24 h. The mixture was then cooled to room temperature and poured into 100 mL of water. The mixture was extracted three times with ethyl acetate. The organic phases were combined, dried, evaporated to dryness, and recrystallized from ethyl acetate and petroleum ether to give 2-hydroxy-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzaldehyde as a yellow solid.

[0039] S5. Preparation of N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]serine

[0040] Dissolve 50-500 mg of 2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzaldehyde and 10-300 mg of serine in ethanol, add 20-200 μL of acetic acid and 50-500 mg of anhydrous sodium sulfate, heat under reflux for 2-6 h, cool to room temperature, add 5-20 mL of a methanol solution of 20-500 mg of sodium cyanoborohydride, stir at room temperature for 24 h, evaporate the solvent, extract the aqueous solution three times with ethyl acetate, combine the organic phases, dry with anhydrous Na2SO4, evaporate the solvent to obtain the desired product.

[0041] S6. Preparation of N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycyl(S-tert-butylthio)cysteine

[0042] Using a solid-phase polypeptide synthesis method, 50-500 mg of N-fluorenylmethoxycarbonyl-S-tert-butylthiocysteine ​​was first linked to 100-800 mg of resin, and the N-protecting fluorenylmethoxycarbonyl group (Fmoc) was removed. Then, the process was repeated twice by linking N-fluorenylmethoxycarbonylglycine and removing the N-protecting fluorenylmethoxycarbonyl group (Fmoc) sequentially, linking two glycine groups, followed by 50-600 mg of N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]serine. 20 mL of 1% trifluoroacetic acid (1 mL of trifluoroacetic acid in 99 mL of CH2Cl2) was added as cutting solution, and the mixture was bubbled with N2 for 1 min. The reaction solution was collected in a round-bottom flask. This cutting operation was repeated 5 times, and all reaction solutions were collected and evaporated to dryness by rotation to obtain an oily liquid. Adding an appropriate amount of icy diethyl ether resulted in the precipitation of a pale yellow solid.

[0043] S7. Preparation of N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycylcysteine

[0044] Dissolve 5-50 mg of N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycyl(S-tert-butylthio)cysteine ​​in 0.5-5 mL of ethanol, add 0.5-5 mL of 12.5 mg / L tris(2-carboxyethyl)phosphine (TCEP), mix well, and let stand at room temperature for 30-60 min. Then, purify the solution by semi-preparative high performance liquid chromatography to obtain the desired N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycylcysteine.

[0045] A sort of 99m Tc-labeled SG2C-type brominated benzyl ether derivatives ( 99mTc-SG2C-CBM), prepared using SG2C-type brominated benzyl ether derivatives, includes the following steps:

[0046] Dissolve 50-800 μg of the labeled precursor (SG2C-type brominated benzyl ether derivative, SG2C-CBM) in ethanol, add 1-10 mg of sodium gluconate, 0.2-10 mg of disodium ethylenediaminetetraacetate, 5-200 μg of stannous chloride and pertechnetic acid solution, shake well, and incubate at 37°C for 45-90 min to obtain the desired product. 99m Tc-SG2C-CBM.

[0047] This invention also discloses the application of an SG2C-type brominated benzyl ether derivative as a ligand to form a complex with a radionuclide marker in PD-L1 imaging, used to prepare an imaging agent reflecting the expression level of PD-L1 on the surface of tumor cells; the radionuclide marker is preferably... 99m Tc.

[0048] Example 1

[0049] As shown in the figure, an SG2C-type bromobenzyl ether derivative has the following structural formula (Ⅰ):

[0050]

[0051] A method for preparing an SG2C-type bromobenzyl ether derivative includes the following steps:

[0052] Preparation of S1, 2-bromo-3-phenyltoluene:

[0053] Under nitrogen purging, 1400 mg of 2-bromo-3-iodotoluene was added to a 100 mL three-necked flask, followed by 50 mL of dioxane / water (5 / 1 v / v) to dissolve it. After stirring for 10 min, 700 mg of phenylboronic acid, 3600 mg of cesium carbonate, and 160 mg of triphenylphosphine palladium were added sequentially. The mixture was stirred at 75 °C for 18 h, and the reaction was stopped. After cooling to room temperature, the solvent was removed by rotary evaporation. The mixture was then extracted three times with water and ethyl acetate. The combined organic phases were dried over anhydrous sodium sulfate, and the organic phase was evaporated to dryness. The solution was then passed through a silica gel column to give 1015 mg of a colorless oil, with a yield of 87.2%. 1 HNMR (400MHz, DMSO-d6) δ7.42-7.31(m,5H,Ar-H), 7.25-7.20(d,2H,Ar-H), 7.12-7.08(m,1H,Ar-H), 2.45(s,3H,Ar-CH3).

[0054] Preparation of S2, 2-bromo-3-phenylbenzyl bromide

[0055] Under nitrogen purging, 1000 mg of 2-bromo-3-phenyltoluene was added to a 100 mL three-necked flask, followed by 40 mL of anhydrous carbon tetrachloride to completely dissolve it. 500 mg of N-bromosuccinimide was added with stirring, and the mixture was heated to 80 °C. Then, 10 mg of benzoyl peroxide was added, and the reaction was continued for 2 hours. The reaction was then stopped, cooled to room temperature, and extracted with water and dichloromethane. The combined organic phases were dried over anhydrous sodium sulfate, and the organic phase was evaporated to obtain a yellow oily substance of 2-bromo-3-phenylbenzyl bromide, which was directly used in the next reaction step.

[0056] Preparation of S3, 2-hydroxy-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzaldehyde

[0057] Under nitrogen purging, 750 mg of 2,4-dihydroxy-5-chlorobenzaldehyde was added to a 100 mL three-necked flask, dissolved in 20 mL of acetonitrile, followed by the addition of 500 mg of NaHCO3. The mixture was stirred at room temperature for 5 min, and then 5 mL of the acetonitrile solution of 2-bromo-3-phenylbenzyl bromide obtained in the previous step was added dropwise. After the addition was complete, the mixture was heated to 75 °C and stirred overnight. The reaction solution was then cooled to room temperature, filtered, and the filtrate was washed with water. The aqueous solution was then extracted twice with ethyl acetate. The combined organic phases were dried over anhydrous Na2SO4, the solvent was evaporated, and the solution was recrystallized from ethyl acetate and petroleum ether to give 640 mg of a pale yellow solid, with a yield of 37.9%. 1 HNMR (400MHz, DMSO-d6) δ11.20(s,1H,-OH), 10.05(s,1H,-CHO), 7.72(d,1H,Ar-H), 7.61(d,1H,Ar-H), 7.51-7.32(m,7H,Ar-H), 6.78(s,1H,Ar-H), 5.28(s,2H,-CH2-).

[0058] Preparation of S4, 2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzaldehyde

[0059] Under nitrogen purging, 460 mg of 2-hydroxy-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzaldehyde was dissolved in 6 mL of DMF, and 300 mg of cesium carbonate was added. The mixture was stirred at room temperature for 15 min, and then 3 mL of a DMF solution containing 260 mg of 3-cyanobenzyl bromide was added dropwise. After the addition was complete, the mixture was heated to 85 °C and stirred overnight. The reaction solution was then cooled to room temperature, poured into 100 mL of water, and extracted three times with ethyl acetate. The combined organic phases were dried, evaporated to dryness, and recrystallized from ethyl acetate and petroleum ether to give 460 mg of a yellow solid, with a yield of 78.5%. 1HNMR (400MHz, DMSO-d6) δ10.24(s,1H,-CHO), 8.05(s,1H,Ar-H), 7.90-7.78(dd,2H,Ar-H), 7.70-7.65(s, 1H,Ar-H), 7.65-7.55(m,2H,Ar-H), 7.55-7.30(m,7H,Ar-H), 7.20(s,1H,Ar-H), 5.50-5.35(m,4H,-CH2-).

[0060] S5. Preparation of N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]serine

[0061] 340 mg of 2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzaldehyde and 120 mg of serine were dissolved in ethanol. 150 μL of acetic acid and 300 mg of anhydrous sodium sulfate were added, and the mixture was heated under reflux for 4 h. After cooling to room temperature, 10 mL of a methanol solution containing 300 mg of sodium cyanoborohydride was added, and the mixture was stirred at room temperature for 24 h. The solvent was evaporated to dryness, and the aqueous solution was extracted three times with ethyl acetate. The combined organic phases were dried over anhydrous Na₂SO₄, and the solvent was evaporated to dryness to give 245 mg of the desired product, with a yield of 61.8%. MS: 623(M+1). 1 HNMR (400MHz, DMSO-d6): δ8.03-7.99(s,1H,Ar-H), 7.92-7.87(d,1H,Ar-H),7.84- 7.79(d,1H,Ar-H),7.67-7.58(m,2H,Ar-H),7.55-7.35(m,8H,Ar-H),7.08-7.03(s ,1H,Ar-H),5.37-5.24(m,4H,-CH2-),4.03-3.93(s,2H,-CH2-),3.76-3.67(m,1H, -CH-), 3.67-3.58(m,1H,-CH-), 3.54-3.26(m,1H,-CH-), 3.23-3.10(m,1H,-NH-).

[0062] S6. Preparation of N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycyl(S-tert-butylthio)cysteine

[0063] Using a solid-phase polypeptide synthesis method, 142 mg of N-fluorenylmethoxycarbonyl-S-tert-butylthiocysteine ​​was first linked to 225 mg of resin, and the N-protecting group fluorenylmethoxycarbonyl (Fmoc) was removed. Then, 81 mg of N-fluorenylmethoxycarbonylglycine was linked, and the N-protecting group fluorenylmethoxycarbonyl (Fmoc) was removed again. This process was repeated twice, linking two glycine groups sequentially. Then, 167 mg of N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]serine was linked. 20 mL of 1% trifluoroacetic acid (1 mL of trifluoroacetic acid was added to 99 mL of CH2Cl2) cleavage solution was added, and the mixture was bubbled with N2 for 1 min. The reaction solution was collected in a round-bottom flask. The cleavage operation was repeated 5 times, and all the reaction solution was collected and evaporated to dryness by rotation to obtain an oily liquid. Upon addition of an appropriate amount of icy diethyl ether, a pale yellow solid precipitated, yielding 165 mg of product (66.3% yield). MS: 928 (M+1). 1 HNMR(400MHz, DMSO-d6):8.73-8.65(s,1H,-NH-),8.35-8.30(s,1H,-NH-),8.26-8.21(s,1H,-NH-),8.15-8.10(s,1H,-NH-),8 .00-7.92(s,1H,Ar-H),7.85-7.76(s,2H,Ar-H),7.63-7.56(d,3H,Ar-H),7.50-7.47(s,1H,Ar-H),7.46-7.42(s,2H,Ar-H),7. 42-7.37(d,1H,Ar-H),7.36-7.32(s,3H,Ar-H),7.06-7.04(s,1H,Ar-H),5.33-5.22(s,4H,-CH2-,),4.48-4.44(d,1H,-CH-),3 .81-3.70(s,8H,-CH2-),3.08-3.03(m,1H,-CH2-),2.94-2.90(m,1H,-CH2-),1.99-1.90(d,1H,-NH-),1.27-1.24(s,9H,-CH3).

[0064] S7. Preparation of N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycylcysteine ​​(SG2C-CBM)

[0065] 10 mg of N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycyl(S-tert-butylthio)cysteine ​​was dissolved in 1 mL of ethanol, and 1 mL of 12.5 mg / L tris(2-carboxyethyl)phosphine (TCEP) was added. The mixture was stirred and allowed to stand at room temperature for 50 min. The solution was then purified by semi-preparative high-performance liquid chromatography (HPLC) at a flow rate of 3 mL / min with a mobile phase of water / acetonitrile (v / v) = 3 / 2. This yielded 7 mg of the desired N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycylcysteine, with a yield of 77.34%. MS: 840 (M+1). 1 HNMR(400MHz, DMSO-d6):8.28-8.18(s,1H,-NH-),8.04-7.95(s,1H,Ar-H),7.88 -7.78(m,2H,Ar-H),7.66-7.56(m,2H,Ar-H),7.55-7.34(m,8H,Ar-H),7.11-7.03

[0066] (s,1H,Ar-H),5.35-5.25(s,4H,-CH2-),4.45-4.36(m,1H,-CH-),4.21-4.05(s,1H,-CH-),3.85-3.72(m,6H,-CH2-),3.45 -3.35(s,1H,-CH-), 2.90-2.70(d,2H,-CH2-), 2.45-2.40(s,2H,-CH2-), 1.27-1.18(s,4H,-CH2-), 0.88-0.83(m,1H,-SH).

[0067] Example 2

[0068] A sort of 99m Tc-labeled SG2C-type brominated benzyl ether derivatives ( 99m Tc-SG2C-CBM), prepared using SG2C-type brominated benzyl ether derivatives, includes the following steps:

[0069] Dissolve 500 μg of the labeled precursor (SG2C-type brominated benzyl ether derivative, SG2C-CBM) in 200 μL of ethanol. Add 120 μL of 50 mg / mL sodium gluconate, 50 μL of 20 mg / mL disodium EDTA, 10 μL of 10 mg / mL stannous chloride, and 0.5 mL of pertechnetic acid solution (1 mCi / mL). Shake well and incubate at 37 °C for 60 min to obtain the desired product. 99m Tc-SG2C-CBM.

[0070] 99m Detection of Tc-SG2C-CBM labeling rate

[0071] Prepared by the above method 99m The Tc-SG2C-CBM labeling rate was determined by isotope high-performance liquid chromatography (HPLC) using a C18 reversed-phase column with a mobile phase of water / acetonitrile = 1 / 1 (v / v) at a flow rate of 1 mL / min. The labeling rate was greater than 93%. Figure 3 As shown. The precursor was not marked, but all other conditions were the same as the marked one. 99m Tc - blank, such as Figure 4 As shown, the retention times of the two differ by more than 5 minutes, allowing them to be completely separated.

[0072] 99m Stability determination of Tc-SG2C-CBM

[0073] The prepared 99m The radiochemical purity of Tc-SG2C-CBM was determined after being placed at room temperature (25℃) for different times (0h, 1h, 2h, 3h, 4h, 5h, 6h).

[0074] Experimental results show that 99m After being placed at room temperature for 6 hours, the radiochemical purity of Tc-SG2C-CBM remained above 90%, indicating that... 99m Tc-SG2C-CBM exhibits good in vitro stability at room temperature, making it suitable for clinical applications.

[0075] 99m The lipid-water partition coefficient of Tc-SG2C-CBM

[0076] Take 50 μL (50 μCi) 99m Tc-SG2C-CBM was added to a test tube containing 1 mL of n-octanol and 1 mL of phosphate buffered saline (PBS, pH = 7.0). After vortexing for 3 min, 100 μL of n-octanol and 100 μL of PBS were removed, and radioactivity was measured on a gamma counter. Then, 200 μL of n-octanol was added to a test tube containing 800 μL of n-octanol and 1 mL of PBS. This process was repeated until the ratio of the n-octanol count to the PBS count was constant. The lipid-water partition coefficient logP = log[(n-octanol count) / (PBS count)] = 1.31, which reflects the partitioning of the drug between the aqueous and lipid phases in vivo.

[0077] 99m Cellular uptake assay of Tc-SG2C-CBM

[0078] Tumor cell lines with different PD-L1 expression levels (A375-hPD-L1, A375) were plated and incubated overnight; the next day, [the following text is missing - likely a continuation of the previous sentence] were added. 99mTc-SG2C-CBM (10 μCi) cells were incubated in a cell culture incubator at 37°C for 2 h. The cell culture medium was aspirated, and the cells were washed twice with PBS. NaOH solution was added for lysis for 15 min, and the cell lysate was collected. Radioactivity was measured on a γ-counter. Cell uptake (AD%) was equal to the percentage of measured cell radioactivity to the total radioactivity added to each well. 99m Tc-SG2C-CBM uptake of PD-L1 in A375-h cells (25.84%) was 6.32 times that in A375 cells (4.09%). The two cell lines showed different levels of PD-L1 expression, which impacted PD-L1 uptake. 99m There were significant differences in Tc-SG2C-CBM intake.

[0079] Example 3

[0080] A sort of 99m Tc-labeled SG2C-type brominated benzyl ether derivatives ( 99m The preparation method of Tc-SG2C-CBM was modified by changing the amount of SG2C-CBM to 50-800 μg, while keeping the contents of other components the same as in Example 2, and following the preparation method and labeling rate determination method of Example 2. Experimental results showed that when the amount of SG2C-CBM reached 300 μg, the labeling rate was greater than 93%.

[0081] Example 4

[0082] A sort of 99m Tc-labeled SG2C-type brominated benzyl ether derivatives ( 99m The preparation method of Tc-SG2C-CBM was modified by changing the amount of sodium gluconate to 1-10 mg, while keeping the contents of other components the same as in Example 2, and following the preparation method and labeling rate determination method of Example 2. Experimental results showed that when the amount of sodium gluconate was in the range of 1.5-10 mg, the labeling rate was greater than 93%.

[0083] Example 5

[0084] A sort of 99m Tc-labeled SG2C-type brominated benzyl ether derivatives ( 99m The preparation method of Tc-SG2C-CBM was modified by changing the amount of disodium ethylenediaminetetraacetate (EDTA) to 0.2-10 mg, while keeping the contents of other components the same as in Example 2. The preparation method and labeling rate determination method of Example 2 were followed. Experimental results showed that when the amount of disodium ethylenediaminetetraacetate was in the range of 0.5-10 mg, the labeling rate was greater than 93%.

[0085] Example 6

[0086] A sort of 99m Tc-labeled SG2C-type brominated benzyl ether derivatives ( 99mThe preparation method of Tc-SG2C-CBM was modified by changing the amount of stannous chloride to 5-200 μg, while keeping the contents of other components the same as in Example 2, and following the preparation method and labeling rate determination method of Example 2. Experimental results showed that when the amount of stannous chloride was in the range of 20-150 μg, the labeling rate was greater than 93%.

[0087] Example 7

[0088] A sort of 99m Tc-labeled SG2C-type brominated benzyl ether derivatives ( 99m The preparation method of Tc-SG2C-CBM was modified by changing the incubation time to 45 min, 60 min, 75 min, and 90 min, while keeping the content of other components the same as in Example 2. The preparation method and labeling rate determination method of Example 2 were followed. Experimental results showed that the labeling rate was greater than 93% when the heating time was above 60 min.

[0089] For parts not specifically described in this invention, existing technologies can be used, and they will not be elaborated here.

[0090] The above description is merely an embodiment of the present invention and does not limit the patent scope of the present invention. Any equivalent structural or procedural transformations made based on the content of the present invention's specification and drawings, or direct or indirect applications in other related technical fields, are similarly included within the patent protection scope of the present invention.

Claims

1. An SG2C-type bromobenzyl ether derivative, characterized in that, The SG2C-type bromobenzyl ether derivative is N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycylcysteine, having the following structural formula (Ⅰ): （Ⅰ）。 2. The method for preparing an SG2C-type bromobenzyl ether derivative according to claim 1, characterized in that, Includes the following steps: N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]serine, Fmoc-glycine, and FMOC-thiotert-butyl-cysteine ​​were used to synthesize N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-seroylglycylglycyl(S-tert-butylthio)cysteine ​​via solid-phase polypeptide synthesis. Tris(2-carboxyethyl)phosphine (TCEP) was added to remove the tert-butylthiosulfate, yielding N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-seroylglycylglycylcysteine.

3. The method for preparing an SG2C-type bromobenzyl ether derivative according to claim 2, characterized in that, The solid-phase polypeptide synthesis method uses resin as a carrier to synthesize N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycyl(S-tert-butylthio)cysteine, and then uses a cleavage solution to cleave N-[2-(3-cyanophenyl-1-methyleneoxy)-4-(2-bromo-3-phenylbenzyloxy)-5-chlorobenzyl]-N-serylglycylglycyl(S-tert-butylthio)cysteine ​​from the resin.

4. The method for preparing an SG2C-type bromobenzyl ether derivative according to claim 3, characterized in that, The cutting fluid is a solution containing trifluoroacetic acid.

5. The application of the SG2C-type brominated benzyl ether derivative as a ligand to form a complex with a radionuclide label in the preparation of PD-L1 imaging agents, according to claim 1.

6. The application of the SG2C-type bromobenzyl ether derivative as a ligand to form a complex with a radionuclide label in the preparation of a PD-L1 imaging agent, as described in claim 5, is characterized in that... The nuclide marker is 99m Tc.

7. A kind 99m Tc-labeled SG2C-type bromobenzyl ether derivatives, characterized in that, It was prepared using an SG2C-type bromobenzyl ether derivative as described in claim 1.

8. The method according to claim 7 99m A method for preparing Tc-labeled SG2C-type bromobenzyl ether derivatives, characterized in that, Includes the following steps: Add glucohepatic acid salt, ethylenediaminetetraacetic acid salt, stannous chloride and pertechnetic acid solution to a solution of an SG2C type bromobenzyl ether derivative as described in claim 1, mix well, and incubate to obtain the desired product.

9. The method according to claim 8 99m A method for preparing Tc-labeled SG2C-type bromobenzyl ether derivatives, characterized in that, The SG2C-type brominated benzyl ether derivative is dissolved in an alcohol solvent to obtain a solution of the SG2C-type brominated benzyl ether derivative.

10. The method according to claim 8 99m A method for preparing Tc-labeled SG2C-type bromobenzyl ether derivatives, characterized in that, The incubation temperature is 30-45℃.

11. The method according to claim 10 99m A method for preparing Tc-labeled SG2C-type bromobenzyl ether derivatives, characterized in that, The incubation time should be 60 minutes or more.

12. The method according to claim 7 99m Application of Tc-labeled SG2C-type brominated benzyl ether derivatives in the preparation of imaging agents for detecting PD-L1 expression.

13. A developer, characterized in that, The developing agent is one of those described in claim 7. 99m Tc-labeled SG2C-type brominated benzyl ether derivatives.

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