CN106317040A - Benzo quinazoline tyrosine kinase inhibitor containing thiophene-sulfonamide structure, preparation method and application - Google Patents

Abstract

The present invention relates to the field of tumor diseases. Specifically, the present invention relates to a novel benzoquinazoline tyrosine kinase inhibitor containing a thiophene sulfonamide structure, its preparation method and its application in the preparation and treatment of tumor diseases. Wherein, R is selected from H, C ₁ -C ₈ alkyl, C ₃ -C ₈ cycloalkyl.

Description

Benzoquinazoline tyrosine kinase inhibitor containing thiophene sulfonamide structure, preparation Method and use

technical field

The invention relates to the field of tumor medicine. Specifically, the present invention relates to a class of tyrosine kinase inhibitors of novel benzoquinazoline derivatives containing a thiophene sulfonamide structure that have a therapeutic effect on the above-mentioned diseases, their preparation methods and uses.

Background technique

Tumor is one of the major diseases that seriously threaten human life and quality of life. According to WHO statistics, about 6.9 million patients worldwide die of tumors every year. Due to the change of living environment and living habits, under the influence of adverse environment and some unfavorable factors, the morbidity and mortality of tumors have shown a rapid upward trend in recent years.

Protein kinases constitute one of the largest families of human enzymes and regulate many different signaling processes by adding phosphate groups to proteins. In particular, tyrosine kinases phosphorylate the phenolic moieties of proteins at tyrosine residues. The tyrosine kinase family includes members that control cell growth, migration and differentiation. Aberrant kinase activity has been implicated in many human diseases, including cancer, autoimmune and inflammatory diseases. Since protein kinases are among the key regulators of cellular signaling, they provide targets for modulating cellular functions with small molecule kinase inhibitors and are therefore good targets for drug design. In addition to the treatment of kinase-mediated disease processes, selective and potent inhibitors of kinase activity can be used to study cellular signaling processes and identify other cellular targets of therapeutic interest.

In the past, the treatment of tumors was achieved by finding and destroying tumors. Now, with the deepening of research on cell signal transduction pathways, people have a deeper understanding of the role of oncogenes and anti-cancer genes inside tumor cells. The design of new anti-tumor drugs based on specific molecular targets of tumors has attracted more and more attention and has become a hot research field. As a new treatment method, targeted anti-tumor drugs have also been applied clinically, and in just a few years Significant progress has been made. It is now known that the protein tyrosine kinase (Protein tyrosine kinases, PTK) signaling pathway is closely related to the proliferation, differentiation, migration and apoptosis of tumor cells, and the use of PTK inhibitors to interfere or block the tyrosine kinase pathway can be used for tumor treat. PTK is a member of the oncoprotein and proto-oncoprotein family that plays an important role in normal and abnormal proliferation. It is an enzyme that can selectively phosphorylate tyrosine residues of different substrates. They catalyze The γ-phosphate group of ATP is transferred to the tyrosine residues of many important proteins to phosphorylate the phenolic hydroxyl groups. Protein tyrosine kinases are divided into receptor tyrosine kinases (receptor tyrosine kinase, RTK), non-receptor tyrosine kinases, IR and Janus kinases, etc. (Robinson D.R., et al, Oncogene, 2000, 19, 5548-5557 ), most of which are receptor tyrosine kinases (RTKs). RTKs are a class of endogenous protein tyrosine kinases that participate in the regulation of various cell activities, play an extremely important role in the transmission of mitogenic signals that initiate cell replication, and regulate cell growth and differentiation. All RTKs belong to type I membrane proteins, and their molecules have a similar topology: a large glycosylated extracellular ligand-binding domain, a hydrophobic single-transmembrane domain, and an intracellular tyrosine kinase catalytic Domains and regulatory sequences. Ligand binding (such as the binding of epidermal growth factor (EGF) to EGFR) leads to the activation of receptor kinase activity partially encoded in the receptor cell, which phosphorylates key tyrosines in the target protein, resulting in the transduction of proliferative signals across the plasma membrane. guide.

In recent years, people have devoted themselves to inhibiting cell signal transduction pathways to develop novel target anti-tumor drugs. Signal transduction inhibitors down-regulate tumor survival and proliferation signals and promote cell apoptosis rather than through cytotoxicity, so they have higher selectivity and less toxic side effects. At present, more than a dozen signal transduction inhibitors have been used in the clinical treatment of tumors, mainly tyrosine kinase inhibitors, among which 4-(substituted anilino) quinazoline compounds have been developed relatively maturely. Such as small molecule inhibitors gefitinib (Iressa), erlotinib (Tarceva) and lapatinib (Lapatinib) targeting EGFR tyrosine kinase targets.

Gefitinib, trade name Iressa (Iressa), an EGFR tyrosine kinase inhibitor developed by AstraZeneca, is the first epidermal growth factor receptor tyrosine kinase inhibitor to enter clinical research. It was launched in Japan and launched in the United States the following year for the treatment of advanced or metastatic non-small cell lung cancer (NSCLC) who have previously received chemotherapy. Erlotinib, trade name Tarceva (Tarceva), an EGFR tyrosine kinase inhibitor developed by OSI, was acquired by Genentech and Roche. It was launched in the United States in 2004 for the treatment of NSCLC and pancreatic cancer. Belonging to the first generation of aniline quinazoline small-molecule inhibitors for the treatment of NSCLC, it is also the only proven EGFR tyrosine kinase inhibitor that has a survival advantage for advanced non-small cell lung cancer, and is effective for all types of non-small cell lung cancer patients It is effective, well tolerated, without myelosuppression and neurotoxicity, and can significantly prolong the survival period and improve the quality of life of patients.

Small-molecule tyrosine kinase inhibitors, as new targeted anti-tumor drugs, have opened a new window for the treatment and prevention of tumors, and have mild side effects and good tolerance. Although more than 10 small-molecule tyrosine kinase inhibitors have made great contributions to clinical tumor therapy, it is still necessary to find some tyrosine kinase inhibitors with better in vivo activity and/or Additional compounds with improved pharmacological properties. Therefore, the development of new improved or more efficient tyrosine kinase inhibitors, and a deeper understanding of the relationship between such drugs and known target proteins and the mechanism of their anti-tumor effects are of great significance to the clinical treatment of tumors.

The invention discloses a class of novel benzoquinazoline tyrosine kinase inhibitors containing a thiophene sulfonamide structure, and these compounds can be used to prepare tumor therapeutic drugs.

Contents of the invention

It is an object of the present invention to provide a tyrosine kinase inhibitor of general formula I.

Another object of the present invention is to provide a process for the preparation of compounds of general formula I.

Another object of the present invention is to provide the compound containing the general formula I as an active ingredient and its application in treating tumors.

The content of the present invention will now be specifically described in conjunction with the purpose of the present invention.

Compounds of the present invention having general formula I have the following structural formula:

Wherein, R is selected from H, C ₁ -C ₈ alkyl, and C ₃ -C ₈ cycloalkyl.

Compounds of general formula I below are preferred,

Wherein, R is selected from H, C ₁ -C ₃ alkyl, and C ₃ -C ₆ cycloalkyl.

More preferred compounds of general formula I are as follows,

The compound of general formula I of the present invention can be synthesized by the following route:

Compound II is first treated with n-BuLi, and the obtained aryl lithium intermediate is reacted with benzaldehyde III to obtain compound IV; compound IV and thiophenesulfonamide V are heated in the presence of triphenylphosphine and diethyl azodicarboxylate reaction to obtain compound I. R is defined as described above.

The compound of the general formula I of the present invention has tyrosine kinase inhibitory effect and can be used as an active ingredient in the preparation of a drug for treating tumors. The activity of the compound of general formula I in the present invention is verified by the experiments of inhibiting EGFR and HER2 kinase and inhibiting cell proliferation in vitro.

The compounds of general formula I according to the invention are effective over a fairly wide dosage range. For example, the daily dose is about in the range of 1mg-500mg/person, divided into one or several administrations. The actual dosage of the compound of general formula I of the present invention can be determined by a doctor according to relevant conditions.

detailed description

The present invention will be further described below in conjunction with embodiment. It should be noted that the following examples are only for illustration, but not for limiting the present invention. Various changes made by those skilled in the art according to the teaching of the present invention shall be within the scope of protection required by the claims of the present application.

The synthesis of embodiment 1 compound I-1

Step 1. Synthesis of compound IV-1

Compound II (2.25g, 10mmol) was dissolved in 25mL of dry THF, stirred under nitrogen protection, cooled to -78°C with liquid nitrogen-ethanol, and 1.6M n-BuLi n-hexane solution (6.25 mL), after the dropwise addition, the reaction mixture was continued to stir at this temperature for 1 h, and then a solution of III-1 (1.24 g, 10 mmol) dissolved in 3 mL of dry THF was slowly added dropwise with a syringe. After the dropwise addition, the reaction mixture was stirred at room temperature for 3 hours, and TLC showed that the reaction was complete. The reaction mixture was carefully poured into 200 mL of ice water, stirred, extracted with 50 mL×3CH ₂ Cl ₂ , the combined extracts were washed with brine (100 mL), and dried over anhydrous sodium sulfate. The desiccant was removed by suction filtration, and the filtrate was evaporated to dryness on a rotary evaporator to obtain compound IV-1 as a white solid. ESI-MS, m/z=315 ([M+H] ⁺ ).

Step 2. Synthesis of Compound I-1

Diethyl azodicarboxylate (DEAD, 1.74g, 10mmol) was dissolved in 20mL of dry THF, stirred, and slowly added dropwise with triphenylphosphine (2.62g, 10mmol) dissolved in 5mL of dry THF under ice-water bath cooling The prepared solution was then added with compound V (1.06 g, 6 mmol), the mixture was stirred at this temperature for 1 hour, and compound IV-1 (1.89 g, 6 mmol) was added. The reaction mixture was reacted overnight at room temperature and then refluxed for 12 hours. TLC showed the reaction was complete. The reaction mixture was carefully poured into 200 mL of ice water, stirred, extracted with 50 mL×3CH ₂ Cl ₂ , the combined extracts were washed with brine (100 mL), and dried over anhydrous sodium sulfate. The desiccant was removed by suction filtration, and the filtrate was evaporated to dryness on a rotary evaporator to obtain compound I-1 as a white solid. ESI-MS, m/z=474 ([M+H] ⁺ ).

The synthesis of embodiment 2 compound 1-2

Step 1. Synthesis of compound IV-2

Compound II (2.25g, 10mmol) was dissolved in 25mL of dry THF, stirred under nitrogen protection, cooled to -78°C with liquid nitrogen-ethanol, and slowly added 1.6M n-BuLi n-hexane solution (6.25 mL), after the dropwise addition, the reaction mixture was continued to stir at this temperature for 1 h, and then a solution of III-2 (1.38 g, 10 mmol) dissolved in 3 mL of dry THF was slowly added dropwise with a syringe. After the dropwise addition, the reaction mixture was stirred at room temperature for 3 hours, and TLC showed that the reaction was complete. The reaction mixture was carefully poured into 200 mL of ice water, stirred, extracted with 50 mL×3CH ₂ Cl ₂ , the combined extracts were washed with brine (100 mL), and dried over anhydrous sodium sulfate. The desiccant was removed by suction filtration, and the filtrate was evaporated to dryness on a rotary evaporator to obtain compound IV-2 as a white solid. ESI-MS, m/z=329 ([M+H] ⁺ ).

Step 2. Synthesis of Compound 1-2

Diethyl azodicarboxylate (DEAD, 1.74g, 10mmol) was dissolved in 20mL of dry THF, stirred, and slowly added dropwise with triphenylphosphine (2.62g, 10mmol) dissolved in 5mL of dry THF under ice-water bath cooling The resulting solution was then added with compound V (1.06 g, 6 mmol), the mixture was stirred at this temperature for 1 hour, and compound IV-2 (1.97 g, 6 mmol) was added. The reaction mixture was reacted overnight at room temperature and then refluxed for 12 hours. TLC showed the reaction was complete. The reaction mixture was carefully poured into 200 mL of ice water, stirred, extracted with 50 mL×3CH ₂ Cl ₂ , the combined extracts were washed with brine (100 mL), and dried over anhydrous sodium sulfate. The desiccant was removed by suction filtration, and the filtrate was evaporated to dryness on a rotary evaporator to obtain compound I-2 as a white solid. ESI-MS, m/z=488 ([M+H] ⁺ ).

Example 3-6

Referring to the operation steps of Example 1, the compounds listed in the following table were synthesized.

Example 7 Compound Inhibits EGFR and HER2 Analysis in Vitro

The following experiments can be used to determine the inhibitory effect of the compounds of the present invention on the activity of erbB family tyrosine kinases (EGFR and HER2) in vitro.

Kinase analysis in vitro was detected with HTScan EGF Receptor Kinase Assay Kit and HTScan HER2/ErbB2 Kinase Assay Kit from Cell Signaling Technology Company. The operation steps refer to the instructions of the kit, and the method detects in vitro the inhibitory effect of the test compound on the phosphorylation of the substrate peptide by EGFR or Her2 receptor tyrosine kinase. Incubate ATP, substrate peptide, and test compound in kinase reaction buffer at room temperature. After incubation for a period of time, stop the reaction by adding stop solution and transfer the sample to a streptavidin-coated 96-well plate, wash the plate and wash with HRP A labeled anti-substrate phosphorylation antibody detects the level of phosphorylation on the substrate peptide, developed with TMB, and stopped with 2M sulfuric acid. Detect the absorption wavelength at 450nm, and calculate the IC ₅₀ value (nM). The results are shown in the table below.

compound EGFR _IC50 (nM) HER2IC ₅₀ (nM) Compound I-1 54 31 Compound I-2 3.7 9.2 Compound I-3 8.2 13.1 Compound I-4 11.5 14.9 Compound I-5 23.7 12.5 Compound I-6 28.1 21.6

It can be seen from the results in the above table that the compound of the present invention has a strong inhibitory effect on EGFR and HER2, and can be used as an anti-tumor drug.

The inhibitory effect of embodiment 8 compound on cell proliferation

The cell proliferation inhibition test used human breast cancer cell line BT474, human gastric cancer cell line NCI-N87, human lung cancer cell line Calu-3 and human skin cancer cell line A431, in which BT474 highly expressed Her2 receptors, and N87 highly expressed EGFR and Her2 receptors. Culture cells in Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum, 2 mM glutamine, and non-essential amino acids in a 37°C, 5% _CO2 cell incubator with trypsin/ethylenediaminetetra Acetic acid (EDTA) Harvested cells from cell culture flasks. Add 4000 cells/well (0.1mL culture medium) to 96-well cell culture plate to adhere to the wall overnight, add 0.1mL dilution of the compound to be tested, the final concentration of DMSO is 0.25%, and place the cell culture plate at 37°C in 5% Incubate for 72 h under CO ₂ conditions. Then the changes of cell morphology were observed under a microscope, and then 50 μL of 50% (mass/volume) trichloroacetic acid (TCA) was added to each well to fix the cells. The final concentration of TCA was 10%. After standing for 5 minutes, place it in a refrigerator at 4°C for 1 hour. Rinse each well of the culture plate with deionized water 5 times to remove TCA, shake it dry, and air-dry until there is no wet trace. Add 100 μL of 0.4% (mass/volume) SRB to each well, let it stand at room temperature for 10 minutes, discard the liquid in each well, rinse with 1% acetic acid for 5 times, air-dry and rinse with pH 10.5, 10mM Tris (trishydroxymethylaminomethane ) 150 μL extraction, detection of absorption wavelength of 540nm. Results IC ₅₀ values (nM) are shown in the table below.

It can be seen from the above table that the compound of the present invention has high inhibitory activity on tumor cells with high expression of EGFR and HER2, and can be used as an anti-tumor drug.

Claims (5)

1. general formula I compound,

Wherein, R is selected from H, C₁-C₈Alkyl, C₃-C₈Cycloalkyl.

2. general formula I compound defined in claim 1,

Wherein, R is selected from H, C₁-C₃Alkyl, C₃-C₆Cycloalkyl.

3. compound of Formula I defined in claim 2, is selected from,

4. the method for compound of Formula I defined in synthesis any one of claim 1-3:

Compound II is first with n-BuLi process, and the aryl lithium intermediate obtained reacts with benzaldehyde III, obtains compound IV；Change Compound IV and thiophene sulfonamide V reacting by heating in the presence of triphenylphosphine and diethylazodicarboxylate, obtain compound I；R's Definition is as described in any one of claim 1-3.

5. the application in terms of preparation treatment tumor disease medicine of the compound of Formula I defined in any one of claim 1-3.