CN117924147A - Isoindoline dichlorobenzene derivative and preparation method and application thereof - Google Patents

Isoindoline dichlorobenzene derivative and preparation method and application thereof Download PDF

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CN117924147A
CN117924147A CN202410069645.4A CN202410069645A CN117924147A CN 117924147 A CN117924147 A CN 117924147A CN 202410069645 A CN202410069645 A CN 202410069645A CN 117924147 A CN117924147 A CN 117924147A
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isoindoline
dichlorobenzene
amino acid
formula
preparation
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王玉记
禄甜
张霁颐
张敏
陈麒羽
吴浩
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Capital Medical University
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Capital Medical University
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Abstract

The invention provides an isoindoline dichlorobenzene derivative, a preparation method and application thereof, and belongs to the technical field of biological medicines. The isoindoline dichlorobenzene derivative provided by the invention can obviously inhibit the mutual combination of PD-1/PD-L1, and can be used for preparing a PD-1/PD-L1 inhibitor, wherein the inhibitor comprises a stereoisomer, pharmaceutically acceptable salt, prodrug, hydrate or solvate thereof and pharmaceutically acceptable auxiliary materials. The inhibitor has remarkable antitumor effect. The inhibition effect of the isoindoline dichlorobenzene derivative provided by the invention on PD-1/PD-L1 is measured by adopting an HTRF (homogeneous phase time resolved fluorescence) technology, and the result shows that the inhibition effect of the isoindoline dichlorobenzene derivative on PD-1/PD-L1 is obviously superior to that of the prior art.

Description

Isoindoline dichlorobenzene derivative and preparation method and application thereof
Technical Field
The invention relates to the technical field of biological medicine, in particular to an isoindoline dichlorobenzene derivative, a preparation method and application thereof.
Background
Tumor immune medicine is used for eliminating tumor cells by enhancing natural immune defense of an organism to tumors, and comprises immune checkpoint inhibitors, therapeutic antibodies, cancer vaccines and the like, wherein clinical researches of the immune checkpoint inhibitors targeting PD-1/PD-L1 are the most mature, and the application is the most extensive. By the 8 th year 2021, 10 PD-1/PD-L1 antibody medicaments are approved to be marketed in China, and the indications comprise melanoma, urothelial cancer, hodgkin's lymphoma, hepatocellular carcinoma, non-small cell lung cancer, nasopharyngeal carcinoma and the like. Wherein, the pamizumab and the attlizumab are used for treating various types of tumors. However, antibody drugs have their inherent problems: ① The tumor tissue has poor permeability, has poor curative effect on partial tumor microenvironment PD-L1 high expression patients, and has long metabolism time, strong immunogenicity and great side effect. ② The administration route is single, and the compliance of patients is poor. ③ High production cost, heavy burden to the medical insurance payment department and the patient, etc. And the micromolecular tumor immunity medicine is hopeful to overcome the defects.
At present, some high-efficiency small molecule compounds are reported sequentially, and some small molecule compounds enter clinical researches. However, no small molecule tumor immunity drug has been marketed until now. Thus, small molecule-based tumor immunotherapy remains one of the most interesting scientific fields of tumor immunotherapy.
Patent application publication No. WO 2018/119263 A1 discloses 5- (2-methylbiphenyl-3-yl) isoindoline having the structural formula:
The above PCT patent application discloses that it is capable of inhibiting the binding of apoptosis receptor 1/apoptosis ligand 1 (PD-1/PD-L1) to each other and is useful in the treatment of tumors. However, the compound has an unsatisfactory inhibition effect on PD-1/PD-L1, the IC 50 detected by an HTRF method is higher than 60nM, and the toxicity is high, wherein the toxicity of human liver normal cells L-02 is less than 10 mu mol, and the water solubility is poor. Therefore, the synthesis of the compound has ideal PD-1/PD-L1 inhibition effect, small toxicity and good water solubility.
Disclosure of Invention
In view of the above, the invention aims to provide an isoindoline dichlorobenzene derivative, a preparation method and application thereof. The compound provided by the invention can inhibit the mutual combination of programmed cell death receptor 1/programmed cell death ligand 1 (PD-1/PD-L1), can be used for preparing PD-1/PD-L1 inhibitors, and has remarkable anti-tumor effect.
In order to achieve the above object, the present invention provides the following technical solutions:
the invention provides an isoindoline dichlorobenzene derivative, which has a structure shown in a formula 1:
In the formula I, R 1 is
L 1 is H or an amino acid residue which is the residual structure of the carboxyl group of the amino acid after removal of one-OH.
Preferably, the amino acid is a natural amino acid or a non-natural amino acid;
the natural amino acid is glycine, alanine, valine, leucine, isoleucine, phenylalanine, proline, tryptophan, serine, tyrosine, cysteine, methionine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine or histidine;
the unnatural amino acids comprise a variety of derivatives of 20 natural amino acids in structure.
Preferably, the compound has a structure shown in any one of formulas IV-1 to IV-3 or V-1 to V-7:
The invention provides a preparation method of the isoindoline dichlorobenzene derivative, when L 1 is H, the preparation method comprises the following steps:
(1) Under the action of a catalyst, carrying out a first substitution reaction on a compound with a structure shown in a formula I and pinacol biborate to obtain a compound with a structure shown in a formula II;
(2) Under the action of a catalyst, the compound with the structure shown in the formula II and the compound with the structure shown in the formula III undergo a second substitution reaction, and the isoindoline dichlorobenzene derivative with the structure shown in the formula IV is obtained after deprotection;
When L 1 is an amino acid residue, the isoindoline dichlorobenzene derivative with the structure shown in the formula IV and amino acid undergo a third substitution reaction to obtain the isoindoline dichlorobenzene derivative with L 1 as the amino acid residue.
Preferably, in the step (1), the catalyst is a Pd (ii) catalyst;
the first substitution reaction is carried out in the presence of potassium acetate, the temperature of the first substitution reaction is 80-110 ℃, and the time is 8-10 h;
In the step (2), the catalyst is Pd (II) catalyst;
the second substitution reaction is carried out in the presence of potassium acetate, the temperature of the second substitution reaction is 80-110 ℃, and the time is 8-10 h.
Preferably, the amino acid is an amino acid after activation of the carboxyl group;
the temperature of the third substitution reaction is room temperature and the time is 8-14 h.
The invention provides application of the isoindoline dichlorobenzene derivative in preparation of a PD-1/PD-L1 inhibitor.
The invention provides a PD-1/PD-L1 inhibitor, which comprises an active ingredient and pharmaceutically acceptable auxiliary materials;
the active ingredients comprise the isoindoline dichlorobenzene derivatives, and stereoisomers, pharmaceutically acceptable salts, prodrugs and hydrates or solvates thereof.
The invention provides application of the isoindoline dichlorobenzene derivative in preparation of antitumor drugs.
The invention provides an anti-tumor drug, which comprises an active ingredient and pharmaceutically acceptable auxiliary materials;
the active ingredients comprise the isoindoline dichlorobenzene derivatives, and stereoisomers, pharmaceutically acceptable salts, prodrugs and hydrates or solvates thereof.
The invention provides an isoindoline dichlorobenzene derivative, which has a structure shown in a formula 1. The isoindoline dichlorobenzene derivative provided by the invention can obviously inhibit the mutual combination of PD-1/PD-L1, and can be used for preparing a PD-1/PD-L1 inhibitor, wherein the inhibitor comprises a stereoisomer, pharmaceutically acceptable salt, prodrug, hydrate or solvate thereof and pharmaceutically acceptable auxiliary materials. The inhibitor has remarkable antitumor effect. The inhibition effect of the isoindoline dichlorobenzene derivative provided by the invention on PD-1/PD-L1 is measured by adopting an HTRF (homogeneous phase time resolved fluorescence) technology, and the result shows that the inhibition effect of the isoindoline dichlorobenzene derivative on PD-1/PD-L1 is obviously superior to that of the prior art.
Drawings
FIG. 1 is a synthetic route for the isoindoline dichlorobenzene derivatives;
FIG. 2 shows Tumor Growth Inhibition (TGI) results for different groups on different days after treatment.
Detailed Description
The invention provides an isoindoline dichlorobenzene derivative, which has a structure shown in a formula 1:
In the formula I, R 1 is
L 1 is H or an amino acid residue which is the residual structure of the carboxyl group of the amino acid after removal of one-OH.
In the present invention, the amino acid is preferably a natural amino acid or other amino acid;
The natural amino acid is preferably glycine, alanine, valine, leucine, isoleucine, phenylalanine, proline, tryptophan, serine, tyrosine, cysteine, methionine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine or histidine;
The structural formula of the other amino acid comprises NH 2 -C-COOH.
In the invention, the isoindoline dichlorobenzene derivative has a structure shown in any one of formulas IV-1 to IV-3 or V-1 to V-7:
The invention provides a preparation method of the isoindoline dichlorobenzene derivative, when L 1 is H, the preparation method comprises the following steps:
(1) Under the action of a catalyst, carrying out a first substitution reaction on a compound with a structure shown in a formula I and pinacol biborate to obtain a compound with a structure shown in a formula II;
(2) Under the action of a catalyst, the compound with the structure shown in the formula II and the compound with the structure shown in the formula III undergo a second substitution reaction, and the isoindoline dichlorobenzene derivative with the structure shown in the formula IV is obtained after deprotection;
In the invention, under the action of a catalyst, a compound with a structure shown in a formula I and pinacol biborate undergo a first substitution reaction to obtain a compound with a structure shown in a formula II. In the present invention, the catalyst is preferably a Pd (II) catalyst, and more preferably a [1,1' -bis (diphenylphosphino) ferrocene ] palladium dichloride dichloromethane complex. In the present invention, the molar ratio of the compound having the structure represented by formula I to the catalyst is preferably 1:0.02 to 0.2, more preferably 1:0.05 to 0.1.
In the invention, the molar ratio of the compound with the structure shown in the formula I to the pinacol biborate is preferably 1:1-2.
In the present invention, the first substitution reaction is preferably performed in the presence of potassium acetate. In the present invention, the molar ratio of the compound having the structure represented by formula I to potassium acetate is preferably 1:2 to 5, more preferably 1:3 to 4. In the present invention, the solvent used in the first substitution reaction is preferably dioxane.
In the present invention, the first substitution reaction is preferably performed under nitrogen gas. In the present invention, the temperature of the first substitution reaction is preferably 80 to 110 ℃, more preferably 100 ℃, and the time is preferably 8 to 10 hours, more preferably 8 hours.
After the first substitution reaction, the present invention preferably performs a post-treatment of the resulting first substitution reaction product, the post-treatment preferably comprising the steps of:
Removing the solvent of the first reaction product, extracting the obtained residue, concentrating the organic phase, and purifying by column chromatography to obtain a pure compound with a structure shown in a formula II.
In the present invention, the means for removing the solvent of the first reaction product is preferably rotary evaporation; the extractant used for the extraction is preferably water and dichloromethane in sequence.
After the compound with the structure shown in the formula II is obtained, the compound with the structure shown in the formula II and the compound with the structure shown in the formula III undergo a second substitution reaction under the action of a catalyst, and the isoindoline dichlorobenzene derivative with the structure shown in the formula IV is obtained after deprotection. In the present invention, the catalyst is preferably a Pd (II) catalyst, and more preferably a [1,1' -bis (diphenylphosphino) ferrocene ] palladium dichloride dichloromethane complex. In the present invention, the molar ratio of the compound having the structure represented by formula II to the catalyst is preferably 1:0.02 to 0.2, more preferably 1:0.05 to 0.1.
In the present invention, the molar ratio of the compound having the structure represented by formula II to the compound having the structure represented by formula III is preferably 1:1 to 2.
In the present invention, the second substitution reaction is preferably performed in the presence of potassium acetate. In the present invention, the molar ratio of the compound having the structure represented by formula II to potassium acetate is preferably 1:2 to 5, more preferably 1:3 to 4. In the present invention, the solvent used in the second substitution reaction is preferably dioxane.
In the present invention, the second substitution reaction is preferably performed under nitrogen gas. In the present invention, the temperature of the second substitution reaction is preferably 80 to 110 ℃, more preferably 90 to 100 ℃; the time is preferably 8 to 12 hours, more preferably 9 to 10 hours.
After the second substitution reaction, the present invention preferably performs a post-treatment of the resulting second substitution reaction product, the post-treatment preferably comprising the steps of:
and removing the solvent of the second reaction product, and extracting, concentrating an organic phase and purifying by column chromatography on the obtained residue.
In the present invention, the means for removing the solvent of the second reaction product is preferably rotary evaporation; the extractant used for the extraction is preferably water and dichloromethane in sequence.
In the present invention, the deprotection method preferably comprises the steps of:
Mixing the second substitution reaction product with an organic solvent, adding a deprotection reagent, carrying out deprotection reaction, and regulating the pH value to 8.
In the present invention, the organic solvent is preferably methylene chloride. In the present invention, the deprotection reagent is preferably trifluoroacetic acid.
In the present invention, the temperature of the deprotection reaction is preferably room temperature and the time is preferably 8 to 12 hours.
After adjusting the pH value to 8, the invention extracts the obtained deprotection reaction, combines organic phases, dries, filters and concentrates, and adds HCl to separate out solid.
In the present invention, the extractant used for the extraction is preferably methylene chloride; the drying is preferably anhydrous sodium sulfate drying.
In the invention, when L 1 is an amino acid residue, the isoindoline dichlorobenzene derivative with the structure shown in the formula IV and amino acid undergo a third substitution reaction to obtain the isoindoline dichlorobenzene derivative with L 1 as the amino acid residue.
In the invention, the molar ratio of the isoindoline dichlorobenzene derivative with the structure shown in the formula IV to the amino acid is preferably 1:1-1.5.
Before the third substitution reaction, the isoindoline dichlorobenzene derivative with the structure shown in the formula IV is preferably activated, the activating agent used in the activation is preferably N-methylmorpholine, and the activating time is preferably 20-30 min, more preferably 25min.
In the present invention, the amino acid is preferably a Boc-protected amino acid, i.e., an N-Boc-amino acid.
In the present invention, the amino acid is preferably a carboxyl-activated amino acid. In the present invention, the carboxyl group-activating reagent is preferably DCC or HOBt. In the present invention, the method for activating the carboxyl group is preferably: mixing amino acid with organic solvent, adding HOBt, activating for 10-15 min, and adding DCC.
In the present invention, the temperature of the third substitution reaction is preferably room temperature, and the time is preferably 8 to 14 hours, more preferably 12 hours.
In the present invention, when the amino acid is a Boc-amino acid, the third substitution reaction is followed by a deprotection reaction. In the present invention, the specific process of the deprotection reaction is referred to above, and will not be described herein.
In the present invention, after the third substitution reaction, the present invention preferably performs a post-treatment of the resulting third substitution reaction product, the post-treatment preferably comprising the steps of:
Extracting the third substituted reaction product, merging organic phases, concentrating, redissolving by using ethyl acetate, carrying out ice bath, adding HCl to precipitate solid, and carrying out solid-liquid separation to obtain the pure isoindoline dichlorobenzene derivative with L 1 as amino acid residue.
In the present invention, the extractant used for the extraction is preferably water and methylene chloride in this order.
In the present invention, the synthetic route of the isoindoline dichlorobenzene derivative is preferably shown in fig. 1. The reactants and conditions in fig. 1 are: a: potassium acetate, 100 ℃, pd (ii), dioxane, pinacol biborate; b: potassium acetate, 100 ℃, pd (ii), dioxane; c: trifluoroacetic acid, dichloromethane: d: DCC, HOBt, anhydrous tetrahydrofuran, amino acids.
The invention provides application of the isoindoline dichlorobenzene derivative in preparation of a PD-1/PD-L1 inhibitor.
The invention provides a PD-1/PD-L1 inhibitor, which comprises an active ingredient and pharmaceutically acceptable auxiliary materials;
the active ingredients comprise the isoindoline dichlorobenzene derivatives, stereoisomers, pharmaceutically acceptable salts, prodrugs and hydrates or solvates thereof. The invention has no special requirement on the pharmaceutically acceptable auxiliary materials, and the auxiliary materials well known in the art can be used.
The invention provides application of the isoindoline dichlorobenzene derivative in preparation of antitumor drugs. In the present invention, the tumor is preferably one or more of colorectal cancer tumor, breast cancer tumor and non-small cell lung cancer tumor.
The invention provides an anti-tumor drug, which comprises an active ingredient and pharmaceutically acceptable auxiliary materials;
The active ingredients comprise the isoindoline dichlorobenzene derivatives, and stereoisomers, pharmaceutically acceptable salts, prodrugs and hydrates or solvates thereof. The invention has no special requirement on the pharmaceutically acceptable auxiliary materials, and the auxiliary materials well known in the art can be used.
The isoindoline dichlorobenzene derivatives, the preparation methods and applications thereof provided by the invention are described in detail below with reference to examples, but they should not be construed as limiting the scope of the invention.
Example 1
The compound prepared in this example is a compound shown as V-1, and the preparation method comprises the following steps:
step one: preparation of Compound II
1000Mg of Compound I (3.356 mmol,1 eq), 852mg of pinacol ester of diboronic acid (3.356 mmol,1 eq), 1315mg of AcOK (13.424 mmol,4 eq) are weighed out and dissolved in 20mL of dioxane, nitrogen protection is carried out, stirring is carried out, the reaction system is placed under an oil bath (100 ℃) stirring, then 274.2mg of catalyst [1,1' -bis (diphenylphosphine) ferrocene ] palladium dichloride dichloromethane complex (0.336 mmol,0.1 eq) are added. The progress of the reaction was monitored by TLC and the reaction was complete for 8 hours. The solvent was removed by rotary evaporation, extracted with water and DCM, and the collected organic phase was concentrated. Purification by column chromatography (PE: ea=5:1) gave 771.8mg of white solid with a developing solvent of PE: ea=10: 1, rf value of 0.5, compound II, yield of 66.6％.1HNMR(300MHz,chloroform-d)δ＝7.768-7.648(m,2H),7.312-7.193(m,1H),4.688(d,J＝8.3Hz,3H),4.634(s,1H),1.528(s,9H),1.357(s,12H).
Step two: preparation of Compound IV-1
600Mg of compound III-1 (2.239 mmol,1 eq), 771mg of compound II (2.4629 mmol,1.1 eq) and 877mg of potassium acetate (8.956 mmol,4 eq) were dissolved in 12mL of dioxane and nitrogen protection was performed. The reaction system was placed under an oil bath (100 ℃ C.) with stirring, 187.7mg of catalyst [1,1' -bis (diphenylphosphino) ferrocene ] palladium dichloride dichloromethane complex (0.23 mmol,0.1 eq) was added and the reaction was complete for 10 hours. The solvent was removed by rotary evaporator, extracted with water and dichloromethane, and the collected organic phase was concentrated using rotary evaporator. Purification by column chromatography gave 644mg of white solid. The resulting mixture was dissolved in 4mL of DCM, 1mL of trifluoroacetic acid was added to conduct the removal of Boc protection, the reaction was carried out for 12 hours, the pH was adjusted to 8, DCM was added to extract, the organic phases were combined, dried over anhydrous sodium sulfate, concentrated by filtration, 0.5mL of 1M HCl was added to precipitate a solid, which was filtered to give 418.5mg of a white solid as IV-1 in 86.6% yield. 1 HNMR (300 mhz, dmso) δ9.62 (s, 2H), 7.57-7.33 (m, 11H), 4.58 (s, 4H).
Example 2
The method is the same as that of IV-1, and the raw material III-1 is changed to III-2, so that IV-2, 1 H NMR (300 MHz, DMSO) delta 10.09 (s, 2H), 7.55-7.29 (m, 6H), 6.99-6.84 (m, 3H), 4.55 (s, 4H) and 4.29 (s, 4H) are obtained.
Example 3
The method is the same as IV-1, and the raw material III-1 is changed into III-3 to obtain IV-3,1H NMR(300MHz,DMSO)δ8.22-8.07(m,2H),7.80-7.67(m,1H),7.64-7.56(m,2H).7.56-7.35(m,4H),4.52(s,4H).
Example 4
100Mg of IV-1 was dissolved in 2mL of N-methylmorpholine (0.32 mmol,1 eq) in an ice bath and activated for 20 minutes as a stock solution. 74mgN-Boc-Gly (0.3278 mmol,1 eq) was weighed and dissolved in 2mL tetrahydrofuran, HOBt was added, and after 10 minutes of activation, DCC was added. Then the mixed solution is added dropwise into the standby solution, the ice bath is removed, the reaction is carried out for 12 hours at normal temperature, eighteen washes are carried out, the organic phase is concentrated, the mixture is purified by a column chromatography method, 133mg of white solid is obtained, 4mL of DCM is added into the mixture for dissolution and stirring, 1mL of trifluoroacetic acid is added for removing Boc protection, after the reaction is carried out for 13 hours, water and dichloromethane are added into the system for extraction, the organic phase is combined for concentration, ethyl acetate is used for redissolution, ice bath is added, 0.5mL of 1M HCl is added, 83.37mg of white solid is obtained by filtration, and the compound V-1 is obtained in yield 79.9％.1HNMR(800MHz,DMSO-d6)δ＝8.21(s,3H),7.50-7.45(m,11H),5.07-4.72(m,4H),3.93(s,2H).
Example 5
The preparation of compound V-2 was the same as in example 4, except that the starting material N-Boc-Gly was changed to N-Boc-L-proline.
1HNMR(300MHz,DMSO)δ10.26(bs,1H),8.65(bs,1H),7.56-7.31(m,11H),5.07(d,J＝14.2Hz,1H),4.98-4.80(m,2H),4.75(d,J＝14.2Hz,1H),4.55(t,J＝7.0Hz,1H),3.28-3.17(m,2H),2.49-2.46(m,1H),1.95(m,3H).
Example 6
The preparation of compound V-3 was the same as in example 4, except that the starting material N-Boc-Gly was changed to N-Boc-L-homoproline.
1H NMR(800MHz,DMSO-d6)δ＝8.21(bs,1H),δ＝8.76(bs,1H),7.50-7.45(m,11H),5.10(d,J＝14.4Hz,1H),4.95((d,J＝14.4Hz,1H),4.88-4.69(m,2H),4.26(d,J＝10.8Hz,1H),3.26(d,J＝12.4Hz,1H),2.98(d,J＝10.8Hz,1H),2.22(d,J＝12.4Hz,1H),1.84-1.48(m,5H).
Example 7
The preparation of compound V-4 was the same as in example 4, except that the starting material N-Boc-Gly was changed to N-Boc-L-alanine.
1H NMR(300MHz,DMSO)δ8.46(bs,3H),7.56-7.31(m,11H),5.10(d,J＝14.4Hz,1H),4.95(d,J＝14.4Hz,1H),4.83(d,J＝16.1Hz,1H),4.72(d,J＝16.1Hz,1H),4.24(q,J＝6.7Hz,1H),1.45(d,J＝6.7Hz,3H).
Example 8
The procedure for the preparation of Compound V-5 was as in example 4, except that IV-1 was changed to IV-2 and N-Boc-Gly was changed to N-Boc-L-alanine.
1H NMR(300MHz,DMSO)8.50(s,3H),7.57-7.29(m,6H),7.01-6.85(m,3H),5.10(d,J＝14.4Hz,1H),4.95(d,J＝14.4Hz,1H),4.83(d,J＝16.1Hz,1H),4.71(d,J＝16.1Hz,1H),4.29(s,4H),4.24(m,1H),1.46(d,3H).
Example 9
The preparation of compound V-6 was the same as in example 4, except that IV-1 was changed to IV-2 and N-Boc-Gly was changed to N-Boc-L-proline.
1H NMR(300MHz,DMSO)δ10.06(bs,1H),8.65(bs,1H),7.54-7.29(m,6H),7.00-6.84(m,3H),5.06(d,J＝14.3Hz,1H),4.90(d,J＝14.3Hz,1H),4.83-4.68(m,2H),4.55(m,1H),4.29(s,4H),3.37-3.12(m,2H),2.44-2.42(m,1H),1.95(m,3H).
Example 10
The preparation of compound V-7 was the same as in example 4, except that IV-1 was changed to IV-2 and N-Boc-Gly was changed to N-Boc-L-homoproline.
1H NMR(300MHz,DMSO)δ9.67(bs,1H),8.69(bs,1H),7.53-7.29(m,6H),7.00-6.84(m,3H),5.11(d,J＝14.4Hz,1H),4.96(d,J＝14.4Hz,1H),4.84(d,J＝16.0Hz,1H),4.71(d,J＝16.0Hz,1H),4.29(s,4H),4.24(m,1H),3.24(d,J＝12.4Hz,1H),2.96(m,1H),2.20(d,J＝12.4Hz,1H),1.63-1.47(m,5H).
Test case
(1) Table 1 lists the ICs 50 of ten compounds of the invention, IV-1, IV-2, IV-3, V-1-V-7, measured in a PD-1/PD-L1 Homogeneous Time Resolved Fluorescence (HTRF) technique assay, using the PD-1/PD-L1 binding detection kit (64 ICP01PEG &64ICP 01-PEH), available from Cisbio.
IC 50 range a for compound: 0-10 nM; b is 10-20 nM; c, 20-30 nM; d, 30-60 nM. See in particular table 1:
TABLE 1 Structure of different Compounds and IC 50 values
According to the in vitro experimental results, the isoindoline dichlorobenzene derivative provided by the invention can inhibit the mutual combination of a programmed cell death receptor 1/a programmed cell death ligand 1 (PD-1/PD-L1), and the effect is obviously better than that of a compound disclosed in patent publication No. WO 2018/119263 A1 and named as 5- (2-methylbiphenyl-3-connecting) isoindoline.
(2) Animal experiment data
To evaluate the antitumor activity of compound V-5, its in vivo antitumor efficacy was evaluated using the C57BL/6j mouse MC38 model. After the tumor volume reached about 50 cubic millimeters, mice received either the drug BMS202 control or the compound V-5 (15 mg/kg) intraperitoneal injection therapy, once a day, for 2 consecutive weeks. Tumor Growth Inhibition (TGI) results for different groups on different days after treatment are shown in figure 2.
In FIG. 2, the in vivo efficacy study of V-5 on MC38 colorectal cancer tumors in mice is shown. Mice were sacrificed 14 days after intraperitoneal injection of vehicle and compound V-5 (15 mg/kg per day) and excised tumors were weighed. A is the change in tumor volume during treatment; b is the weight of each group of resected tumors; c is an image of each group of resected tumors. Data are expressed as mean ± SD, P <0.05 compared to control, n=3.
It can be seen that compound V-5 significantly inhibited tumor growth without causing weight loss or death during treatment, and that the normal tissues of the group receiving V-5 treatment were not significantly damaged compared to the untreated group. These results indicate that an effective dose of V-5 is well tolerated. After the end of the experiment, mice were sacrificed by cervical dislocation, tumors were dissected and weighed.
(3) Security data
Several conventional cells were selected from different tissues for in vitro culture, and experiments demonstrated that the compounds were cytotoxic to different normal cell lines, including rat schwann cells, human immortal epidermal cells, human normal lung epithelial cells, immortal cell lines, and human normal hepatocytes. Cells were seeded on 96-well clear plates and cultured with increasing compound concentrations using DMSO-treated cells as a control. The concentration of DMSO was kept below 0.5%. After 48 hours of incubation, thiazolyl tetrazolium bromide (Heowns, tianjin) was added according to the manufacturer's protocol and the plates were incubated for an additional 4 hours (37 ℃,5% co 2). Absorbance was measured at 570nm wavelength using a microplate reader (MolecularDevice SpectraMax ID). Data are presented as different cell viability relative to PBS-treated control cells. Data points represent mean ± SD values of three independent experiments. Half maximal effective concentrations (EC 50 values) were calculated from the dose-response curve using the OriginPro 2020 (OriginLab) software. And screening the potent compounds with in vitro nonspecific toxicity through an MTT test. Representative compound data for IV series and V series are shown in table 2.
TABLE 2 cytotoxicity of different Compounds
The results show that the IV series and V series compounds have no obvious in vitro toxicity.
The foregoing is merely a preferred embodiment of the present invention and it should be noted that modifications and adaptations to those skilled in the art may be made without departing from the principles of the present invention, which are intended to be comprehended within the scope of the present invention.

Claims (10)

1. An isoindoline dichlorobenzene derivative has a structure shown in a formula 1:
In the formula I, R 1 is
L 1 is H or an amino acid residue which is the residual structure of the carboxyl group of the amino acid after removal of one-OH.
2. The isoindoline dichlorobenzene derivative according to claim 1, wherein the amino acid is a natural amino acid or a non-natural amino acid;
the natural amino acid is glycine, alanine, valine, leucine, isoleucine, phenylalanine, proline, tryptophan, serine, tyrosine, cysteine, methionine, asparagine, glutamine, threonine, aspartic acid, glutamic acid, lysine, arginine or histidine;
the unnatural amino acids comprise a variety of derivatives of 20 natural amino acids in chemical structure.
3. The isoindoline dichlorobenzene derivative according to claim 1 or 2, which has a structure represented by any one of formulas IV-1 to IV-3 or V-1 to V-7:
4. a process for producing isoindoline dichlorobenzene derivatives according to any of the claims 1-3, characterized in that,
When L 1 is H, the preparation method comprises the following steps:
(1) Under the action of a catalyst, carrying out a first substitution reaction on a compound with a structure shown in a formula I and pinacol biborate to obtain a compound with a structure shown in a formula II;
(2) Under the action of a catalyst, the compound with the structure shown in the formula II and the compound with the structure shown in the formula III undergo a second substitution reaction, and the isoindoline dichlorobenzene derivative with the structure shown in the formula IV is obtained after deprotection;
When L 1 is an amino acid residue, the isoindoline dichlorobenzene derivative with the structure shown in the formula IV and amino acid undergo a third substitution reaction to obtain the isoindoline dichlorobenzene derivative with L 1 as the amino acid residue.
5. The method according to claim 4, wherein in the step (1), the catalyst is a Pd (II) catalyst;
the first substitution reaction is carried out in the presence of potassium acetate, the temperature of the first substitution reaction is 80-110 ℃, and the time is 8-10 h;
In the step (2), the catalyst is Pd (II) catalyst;
the second substitution reaction is carried out in the presence of potassium acetate, the temperature of the second substitution reaction is 80-110 ℃, and the time is 8-10 h.
6. The method according to claim 4, wherein the amino acid is an amino acid after activation of carboxyl group;
the temperature of the third substitution reaction is room temperature and the time is 8-14 h.
7. The use of the isoindoline dichlorobenzene derivative according to any one of claims 1-3 or the isoindoline dichlorobenzene derivative prepared by the preparation method according to any one of claims 4-6 in the preparation of a PD-1/PD-L1 inhibitor.
8. A PD-1/PD-L1 inhibitor comprises an active ingredient and pharmaceutically acceptable auxiliary materials;
The active ingredient comprises the isoindoline dichlorobenzene derivative according to any one of claims 1-3 or the isoindoline dichlorobenzene derivative prepared by the preparation method according to any one of claims 4-6, and stereoisomers, pharmaceutically acceptable salts, prodrugs and hydrates or solvates thereof.
9. The use of the isoindoline dichlorobenzene derivatives according to any one of claims 1 to 3 or the isoindoline dichlorobenzene derivatives prepared by the preparation method according to any one of claims 4 to 6 in the preparation of antitumor drugs.
10. An antitumor drug comprises active ingredients and pharmaceutically acceptable auxiliary materials;
The active ingredient comprises the isoindoline dichlorobenzene derivative according to any one of claims 1-3 or the isoindoline dichlorobenzene derivative prepared by the preparation method according to any one of claims 4-6, and stereoisomers, pharmaceutically acceptable salts, prodrugs and hydrates or solvates thereof.
CN202410069645.4A 2024-01-17 2024-01-17 Isoindoline dichlorobenzene derivative and preparation method and application thereof Pending CN117924147A (en)

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