A quinazoline small molecule compound and a preparation method and application thereof

Abstract

The application belongs to the technical field of medicine, and particularly relates to a quinazoline small-molecule compound and a preparation method and application thereof. 25 H 19 N3O, a structural formula of which is: which is obtained by reacting 6-methoxy-2-naphthaldehyde and 2-(2-aminophenyl-)-1H-benzimidazole under a heated condition. The quinazoline small-molecule compound shows good selective inhibitory effect on PTP1B, and has a strong inhibitory effect on HepG2 hepatoma cells at a cell level, and can be applied to the preparation of tumor drugs.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to a quinazoline small molecule compound, its preparation method, and its application. Background Technology

[0002] Protein tyrosine phosphatases (PTPs) are widely distributed non-metallic enzymes within cells. Together with protein tyrosine kinases, they regulate the phosphorylation level of tyrosine within cells. They are expressed in various tissues of the human body and play important roles in a variety of physiological processes. Abnormal expression of PTPs can lead to excessive or insufficient tyrosine dephosphorylation, resulting in many diseases related to signal transduction disorders, including cancer, diabetes, obesity, and immune disorders. Currently, numerous genetic and pharmacological studies have shown that many PTPs are closely related to human diseases, and some of these PTPs hold promise as therapeutic targets for human diseases.

[0003] Among the vast family of PTPs, PTP1B has attracted the most attention in terms of research and application. PTP1B was isolated from human placental tissue in 1988, becoming the first PTP to be successfully isolated. Its crystal structure was first confirmed in 1994. PTP1B contains 435 amino acid residues, of which 30-278 amino acids constitute the catalytic functional region of the enzyme, and it is located on the endoplasmic reticulum via a C-terminus of 35 proline residues. The main structural features of PTP1B are three highly conserved ring structures: a catalytically active region containing Cys215, a WPD ring, and a second binding site. Recent literature reports overexpression of PTP1B in breast cancer and liver cancer tissues, suggesting that PTP1B inhibitors may be potential therapeutic agents.

[0004] Currently, although a number of small molecule inhibitors of PTP1B have been reported, due to the high homology of the PTP family, especially the 74% sequence homology between TCPTP and PTP1B, there is still a lack of inhibitors with high activity and high selectivity. Therefore, the development of highly effective and specific inhibitors for the treatment of PTP1B-related diseases remains of practical significance. Summary of the Invention

[0005] In view of this, the purpose of this invention is to overcome the shortcomings of the prior art and provide a quinazoline small molecule compound, its preparation method and application.

[0006] To achieve the above objectives, the technical solution of the present invention is as follows:

[0007] A quinazoline small molecule compound with the molecular formula: C 25 H 19 N3O, its structural formula is:

[0008]

[0009] The quinazoline small molecule compound belongs to the monoclinic crystal system, with space group P21 / n and unit cell parameters as follows: α=90°, β=94.986(5), γ=90°.

[0010] The preparation method of the quinazoline small molecule compound as described above includes the following steps: dissolving 6-methoxy-2-naphthaldehyde and 2-(2-aminophenyl-)-1H-benzimidazole in methanol, and heating under reflux to obtain the quinazoline small molecule compound.

[0011] Furthermore, the molar ratio of 6-methoxy-2-naphthaldehyde to 2-(2-aminophenyl-)-1H-benzimidazole is 1:1.

[0012] Furthermore, the heating reflux temperature is 60°C, and the time is 3 hours.

[0013] The asymmetric unit of the quinazoline small molecule compound of the present invention comprises one molecule. During the preparation process, under the action of heating, 6-methoxy-2-naphthaldehyde undergoes cyclization with 2-(2-aminophenyl-)-1H-benzimidazole, and the ligand is transformed from a Schiff base compound into a 6-(6-methoxynaphth-2-yl)-12,12a-dihydrobenzo[4,5]imidazo[1,2-c]quinazoline small molecule compound. At the same time, the C-H···N in one molecule forms a weak molecular interaction with the C-H···N in another molecule, forming a one-dimensional chain structure.

[0014] The application of quinazoline small molecule compounds as PTP1B inhibitors, as described above.

[0015] A PTP1B inhibitor comprising a quinazoline small molecule compound as described above.

[0016] The application of quinazoline small molecule compounds or PTP1B inhibitors as described above in the preparation of oncology drugs.

[0017] Furthermore, the tumor includes liver cancer.

[0018] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0019] The quinazoline small molecule compound of the present invention is obtained by reacting 6-methoxy-2-naphthaldehyde with 2-(2-aminophenyl-)-1H-benzimidazole under heating conditions. The preparation method is simple, the product has high purity and a crystal structure, and the yield is high, reaching 70%.

[0020] The quinazoline small molecule compound provided by this invention can effectively inhibit PTP1B and has approximately 5 times the selectivity compared to its homologous TCPTP, while also exhibiting strong inhibitory activity against HepG2 human liver cancer cells. This quinazoline small molecule compound is also a potential candidate for treating PTP1B-related diseases. Attached Figure Description

[0021] Figure 1 Synthetic route of the quinazoline small molecule compound of this invention

[0022] Figure 2 The asymmetric structural unit of the quinazoline small molecule compound of the present invention

[0023] Figure 3 One-dimensional chain diagram of the quinazoline small molecule compound of this invention

[0024] Figure 4 Electrospray mass spectra of the quinazoline small molecule compounds of this invention

[0025] Figure 5 The IC50 of the quinazoline small molecule compound of this invention inhibits PTP1B activity 50 Value measurement curve

[0026] Figure 6 The IC50 of the quinazoline small molecule compound of this invention inhibits TCPTP activity 50 Value measurement curve

[0027] Figure 7 Effects of the quinazoline small molecule compound of this invention on the proliferation of HepG2 cells Detailed Implementation

[0028] To facilitate understanding of the present invention, a more comprehensive description will be given below. However, the present invention can be implemented in many different forms and is not limited to the embodiments described herein. Rather, these embodiments are provided to provide a thorough and complete understanding of the disclosure of the present invention.

[0029] Example 1: Preparation of small molecule quinazoline compounds

[0030] Synthetic routes for quinazoline small molecule compounds are as follows: Figure 1The specific process is as follows: 0.186 g (1 mmol) of 6-methoxy-2-naphthaldehyde was added to 10 mL of methanol and heated and stirred until completely dissolved. Then, 0.209 g (1 mmol) of 2-(2-aminophenyl-)-1H-benzimidazole was dissolved in 10 mL of methanol. The dissolved benzyl ether was added dropwise to the methanol solution, and stirring was started. The mixture was heated under reflux at 60 °C for 3 hours to form a colorless mixture. After the colorless mixture cooled to room temperature, it was filtered. The filtrate was placed in a clean glass vial and allowed to stand at room temperature. After about 10 days, colorless blocky crystals precipitated on the vial wall, yielding the quinazoline small molecule compound with a yield of 72%. Elemental analysis: C 25 H 19 N3O: Theoretical values: C, 79.55, H, 5.07, N, 11.13; Experimental values: C, 79.26, H, 5.23, N, 11.04.

[0031] Example 2: Crystal structure analysis of quinazoline small molecule compounds

[0032] After trying various methods for long crystallization, single crystals of the quinazoline small molecule compound were finally obtained through room-temperature volatilization. The structure of this compound was tested by selecting regularly shaped, relatively transparent bulk single crystals under a microscope, attaching them to glass fibers, and then performing the tests using a Bruker APEX-Ⅱ CCD diffractometer at the Beijing Synchrotron Radiation Facility (BSRF) 3W1A beamline. A graphite monochromator was used. X-ray diffraction data were collected at room temperature using ω-2θ scanning. The unit cell parameters were determined using the SMART program, and the original data were restored and corrected using the SAINTPLUS program. The final data were analyzed using the SHELXS-97 package via a direct method. Finally, the data were refined using the SHELXL197 (Sheldrick, 2008) package. The obtained unit cell parameters and hydrogen bond information are shown in Tables 1 and 2. The crystal analysis results show that the small molecule compound prepared in this invention belongs to the monoclinic crystal system, space group P21 / n, and its unit cell parameters are: α = 90°, β = 94.986(5), γ = 90°. Its asymmetric unit contains a molecule (such as...). Figure 2 Simultaneously, weak intermolecular interactions form one-dimensional chain structures (such as...). Figure 3 ).

[0033] Table 1. Crystallographic data of quinazoline small molecule compounds

[0034]

[0035] Table 2 Hydrogen bond information for quinazoline small molecule compounds

[0036] D—H···A D—H H···A D···A D—H···A <![CDATA[C14—H14···N1 i ]]> 0.98 2.60 3.569(4) 169

[0037] Example 3: Electrospray mass spectrometry testing of quinazoline small molecule compounds

[0038] To investigate the existence of the quinazoline small molecule compound of the present invention in solution, a small amount of the compound crystals were dissolved in DMSO, the supernatant was collected, and the sample was loaded onto an electrospray mass spectrometer. The electrospray ion source was used to detect and record the data in a positive ion mode. Figure 4 Table 3 shows the positive ion electrospray mass spectrum of this compound, in which the molecular ion peak can be observed. The results indicate that the experimental values ​​are consistent with the theoretical values, suggesting that the compound exists stably in the synthesized form.

[0039] Table 3 Electrospray mass spectra of quinazoline derivatives

[0040] Quinazoline small molecule compounds Species Experimental values Theoretical value <![CDATA[C 25 H 19 N3O]]> <![CDATA[[(1)+H] + ]]> 378.07 378.45

[0041] Example 4: IC50 of the inhibitory effect of quinazoline small molecule compounds on PTP1B and TCPTP activity 50 Value determination

[0042] IC 50 Assay Method: The inhibitory effects of PTP1B and TCPTP activity were determined using a previously reported method, with p-nitrophenol phosphate (pNPP) as the substrate for the enzyme inhibition assay. The inhibitor was dissolved in DMSO and serially diluted. The experiment was conducted in clear 96-well plates. First, 83 μL of enzyme-containing MOPS buffer (pH = 7.24) was added to the first three rows of the plate, along with a blank control group containing enzyme-free MOPS buffer. 10 μL of DMSO solution was added to the first column, and the second to eighth columns were treated as experimental groups, with inhibitors added at varying concentrations (from lowest to highest). After addition, the plates were thoroughly mixed. The plates were incubated at 37°C for 30 min, and then 2 μL of 0.1 M pNPP solution was added to initiate the reaction. After approximately 10 min, the solution in the 96-well plate turned yellow. Then, 5 μL of 2 M NaOH was added to terminate the reaction. The absorbance at substrate decomposition was measured (λ = 405 nm). Finally, the data were processed to obtain the IC50 value. 50 Values. The inhibitor solutions for each concentration gradient were freshly prepared before each experiment. This experiment needs to be repeated three times or more to ensure the accuracy and reliability of the experimental data. The experimental results are as follows: Figure 5 and 6 As shown. The half-maximal inhibitory concentration (IC50) of the quinazoline small molecule compound of the present invention against PTP1B is... 50 The half-maximal inhibitory concentration (IC50) for TCPTP was 0.69 μM. 50 The concentration is 3.20 μM, which is a highly selective inhibitor.

[0043] Example 5: Study on the effect of quinazoline small molecule compound on the proliferation of HepG2 cells

[0044] Overexpression of PTP1B was detected in HepG2 cells. To investigate whether the quinazoline small molecule compound inhibited PTP1B expression in cells, HepG2 cells were used for this study. All HepG2 cells were cultured in DMEM medium. When the cells reached approximately 80% confluence, the old medium was discarded, and the cells were washed twice with 0.85% NaCl. Residual liquid was removed with a pipette, and an appropriate amount of trypsin was added to digest the cells. After centrifugation, the supernatant was collected, and 1 mL of fresh medium was added and mixed thoroughly. An appropriate amount of cell suspension was added to a culture flask containing 5-6 mL of medium, and the flask was shaken in an "∞" shape before being placed in an incubator for further incubation.

[0045] Once the cells reached the logarithmic growth phase, they were washed twice with 0.85% NaCl, digested with trypsin, centrifuged, and then resuspended in 1 mL of fresh culture medium. Depending on the culture time, 50 μL of different concentrations of cell suspension were seeded into each well of a 96-well plate. After cell attachment, quinazoline small molecule compound at final concentrations of 0, 1, 5, 10, 25, and 50 μM was added for 24 h, 48 h, and 72 h, respectively. After incubation with MTT for 4 h, the supernatant was discarded, and 150 μL LDMSO was added. The cells were shaken for approximately 10 min, and the absorbance was measured using a microplate reader. The experimental results are shown below. Figure 7 As shown in the figure, the quinazoline small molecule compound significantly reduced the survival rate of HepG2 cells, and the cell survival rate was positively correlated with the treatment time and concentration. After treatment with 50 μM quinazoline small molecule compound for 24 h, 48 h, and 72 h, the cell survival rates of HepG2 cells were 87.35%, 67.73%, and 45.61%, respectively. It is preliminarily speculated that the quinazoline small molecule compound forms hydrogen bonds with certain intracellular proteins, enhancing the interaction between the complex and the protein, thereby exhibiting better antitumor activity.

[0046] The above description is only for better explaining the embodiments of the present invention and is not intended to limit them. Any modifications or equivalent substitutions that do not depart from the spirit and scope of the present invention shall fall within the scope of the present invention.

Claims

1. A quinazoline small molecule compound, characterized in that, Its molecular formula is: C 25 H 19 N3O, structural formula: 。 2. The quinazoline small molecule compound according to claim 1, characterized in that, The quinazoline small molecule compound belongs to the monoclinic crystal system, with space group P21 / n and unit cell parameters as follows: a =10.6210(10)Å, b =10.2037(8)Å, c =17.5041(14)Å, α =90°, β =94.986(5), γ =90°.

3. The method for preparing the quinazoline small molecule compound according to claim 1 or 2, characterized in that, Includes the following steps: The quinazoline small molecule compound was prepared by dissolving 6-methoxy-2-naphthaldehyde and 2-(2-aminophenyl-)-1H-benzimidazole in methanol and heating under reflux.

4. The method for preparing the quinazoline small molecule compound according to claim 3, characterized in that, The molar ratio of 6-methoxy-2-naphthaldehyde to 2-(2-aminophenyl-)-1H-benzimidazole is 1:

1.

5. The method for preparing the quinazoline small molecule compound according to claim 3, characterized in that, The heating reflux temperature is 60 °C, and the time is 3 h.

6. The use of the quinazoline small molecule compound according to claim 1 or 2 in the preparation of PTP1B inhibitors.

7. A PTP1B inhibitor, characterized in that, Including the quinazoline small molecule compound as described in claim 1 or 2.

8. The use of the quinazoline small molecule compound of claim 1 or 2 or the PTP1B inhibitor of claim 7 in the preparation of liver cancer drugs.

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