CN111329857A - A kind of fluorobenzamide derivative injection and its anti-breast cancer application - Google Patents

Abstract

The invention provides a fluorobenzamide derivative injection and its anti-breast cancer application, belonging to the technical field of medicine. The test found that the fluorobenzamide derivative BOF has a strong cytotoxic effect on MCF-7 breast cancer cells, but has no toxicity on MCF-10A normal breast epithelial cells. In addition, the present invention greatly improves the water solubility of the fluorobenzamide derivative BOF by adding glycine as a cosolvent, so that a high-concentration anti-breast cancer injection can be prepared.

Description

A kind of fluorobenzamide derivative injection and its anti-breast cancer application

technical field

The invention belongs to the technical field of medicine, and relates to a new anticancer application of a compound, more particularly, to a fluorobenzamide derivative injection and the application of the fluorobenzamide derivative in the preparation of an anti-breast cancer drug.

Background technique

According to the statistics of the World Health Organization, since 2012, the number of cancer patients has been increasing at an annual rate of 350,000, and it is about to become the biggest threat to human health. Especially for women, breast cancer has become one of the common cancers. However, in economically developed regions such as Europe and the Americas, cancer prevention is relatively emphasized. The promotion of early screening enables many breast cancer patients to be detected and treated at an early stage, and the probability of recovery is relatively high. However, my country's current attitude towards breast cancer is not as good as that of European and American countries in terms of national policies, public awareness and medical level. Therefore, the detection of breast cancer is basically at an advanced stage, so the cure rate of breast cancer in my country is low.

Although significant breakthroughs have been made in the clinical treatment of breast cancer, the recurrence and metastasis rates are still increasing. At present, the treatment of breast cancer is mainly through surgery, radiotherapy and chemotherapy. However, its development has encountered a bottleneck. For example, it is difficult to completely eliminate tumor tissue by surgery, and radiotherapy causes great damage to normal tissue. Chemotherapy drugs such as paclitaxel have significant anti-tumor effects and have become the most commonly used drugs for the treatment of breast cancer. However, long-term use of it can cause toxic side effects and develop drug resistance, leading to an increased risk of metastasis. Therefore, medical workers all over the world have been working on research and development of new chemotherapy drugs to make up for the disadvantages of existing drugs.

The breast cancer MCF-7 cell line was isolated and established from the pleural effusion of a 69-year-old Caucasian female breast cancer patient. It is also the most studied human breast cancer cell line in the world and has been promoted by multiple research groups for over 40 years [Lee Adrian V., Oesterreich Steffi, Davidson Nancy E. MCF-7Cells—Changing the Course of Breast Cancer Research and Care for 45Years.Journal of the National Cancer Institute. 2015.107(7):1-4]. Therefore, the results of this cell line have great implications for breast cancer research and patient prognosis.

CN110818672A discloses that novel cyclobutenone compounds are obtained through efficient competitive [2+2] cycloaddition reaction, and the target compound has good antioxidative effect through antioxidative detection by DPPH method. However, by searching the prior art at home and abroad, there is no literature that this cyclobutenone compound has anti-cancer activity, and there is no literature report that the compound is used for anti-breast cancer.

SUMMARY OF THE INVENTION

The inventors discovered for the first time that the fluorobenzamide derivative BOF has the biological activity of inhibiting the proliferation of breast cancer MCF-7, and the discovery can be used to prepare an anti-breast cancer drug. Therefore, the first object of the present invention is to provide a pharmaceutical use of a fluorobenzamide derivative, namely: the application of a fluorobenzamide derivative in the preparation of a medicine for inhibiting the proliferation of breast cancer cells; and the fluorobenzamide derivative The application of the compound in the preparation of anti-breast cancer drugs.

It should be noted that the molecular formula of the above-mentioned fluorobenzamide derivative BOF is C ₁₈ H ₁₂ FNO ₄ (molecular weight 325), and the chemical name is N-[2-(2H-1,3-benzodioxol-5-yl)-4 -oxocyclobut-1-en-1-yl]-4-fluorobenzamide, its hydrogen spectrum data are: ¹ H NMR (400MHz, DMSO-d ₆ ): δ10.68 (s, 1H), 7.89 (d, J=4.0 Hz, 2H), 7.70(d, J＝4.0Hz, 2H), 7.52-7.39(m, 2H), 6.93(d, J＝8.0Hz, 1H), 6.11(s, 2H), 4.35(s, 2H) ). Its chemical structure is as follows:

In addition, it was found in the experiment that the fluorobenzamide derivative BOF has poor water solubility, and its solubility in water is less than 1 mg/ml, which is not conducive to the preparation of injection. For this reason, the inventors have generally estimated the addition of solubilizers such as propylene glycol, Tween, poloxamer, PEG, polyoxyethylene castor oil, etc., but the result still does not improve the solubility to more than 2 mg/ml. In a further test, the inventor unexpectedly found that a certain amount of glycine as a co-solvent can greatly enhance the water solubility of BOF, so that a high-concentration BOF injection can be prepared. Therefore, the second object of the present invention is to provide a kind of fluorobenzamide derivative injection, and the concrete technical scheme is summarized as follows:

A fluorobenzamide derivative injection, the injection contains a fluorobenzamide derivative, a cosolvent and an isotonicity regulator, and the cosolvent is glycine.

Further preferably, the above-mentioned fluorobenzamide derivative injection, wherein the concentration of each component in the injection is:

Fluorobenzamide derivatives 2-10mg/ml

Glycine 0.5-5mg/ml

Isotonicity modifier 3-9mg/ml.

Still further preferably, the above-mentioned fluorobenzamide derivative injection, wherein the concentration of each component in the injection is:

Fluorobenzamide derivatives 5-10mg/ml

Glycine 1-4mg/ml

Isotonicity modifier 6-9mg/ml.

Still further preferably, in the above-mentioned fluorobenzamide derivative injection, the isotonicity modifier is selected from any of the following: sodium chloride, glucose, mannitol, and sorbitol. The isotonicity regulators are most commonly used for the effect of sodium chloride or glucose.

The beneficial effect of the present invention is that: the fluorobenzamide derivative BOF has a strong cytotoxic effect on MCF-7 breast cancer cells, but has basically no toxicity on MCF-10A normal breast epithelial cells. In addition, the present invention greatly improves the water solubility of the fluorobenzamide derivative BOF by adding glycine as a cosolvent, so that a high-concentration anti-breast cancer injection can be prepared.

Description of drawings

Figure 1: Effects of different drug concentrations on the survival of human breast cancer cells.

Detailed ways

The present invention will be further described in detail below through specific embodiments. However, those skilled in the art will understand that the following examples are only used to illustrate the present invention, and should not be construed as limiting the scope of the present invention. In addition, if the specific technical operation steps or conditions are not indicated in the examples, all are carried out in accordance with the techniques or conditions described in the literature in this field or in accordance with the product specification. The reagents or instruments used without the manufacturer's indication are conventional products that can be obtained from the market.

Example 1 Screening experiment of fluorobenzamide derivatives (BOF) cosolvents

Take three 100ml beakers, add 50ml of distilled water, and then add 1g, 2g, and 4g of propylene glycol respectively. After stirring evenly, add 0.5g of BOF to the three beakers, and stir repeatedly with a glass rod for 2min to observe the dissolution of BOF. The results showed that the solutions in the three beakers were all turbid, and insoluble particles appeared at the bottom of the beakers after being placed for 1 hour.

According to the above method, propylene glycol was replaced with Tween 80, poloxamer, propylene glycol, PEG400, polyoxyethylene castor oil, and glycine, respectively. The results showed that only glycine could promote the dissolution of BOF, and other cosolvents could not dissolve BOF. The solubility of glycine increased to above 2 mg/ml, which may be due to the formation of a soluble complex between glycine and BOF.

The screening experiment of embodiment 2 glycine dosage

Take five 100ml beakers, add 50ml distilled water respectively, then add 0.05g, 0.1g, 0.2g, 0.4g, 0.8g glycine, stir evenly, add 0.5g BOF to each of the five beakers, and stir repeatedly with a glass rod for 2min , observe the dissolution of BOF. The results showed that the solution in the beaker to which 0.05 g of glycine was added was turbid; the solution in the other beakers was clear, and no insoluble particles appeared after being placed for 1 hour.

Example 3 Culture of human breast cancer MCF-7 cells

RPMI1640 medium containing 10% FBS was used as complete medium. First, revive the MCF-7 cells, place the culture flask in an incubator with a temperature of 37°C and 5% CO ₂ to incubate, when the cell density is about 80%, discard the waste liquid, wash it, and then use 0.25% pancreatic After enzymatic digestion, the cells were carefully observed under a microscope to form a spherical shape, and the digestion process was stopped by adding serum-containing medium to the culture flask. Then carefully blow off the cells and add them to a 15mL centrifuge tube, centrifuge (1000r/min, 5min), discard the supernatant after centrifugation, and use a pipette to pipette an appropriate amount of the complete culture-based centrifuge tube. Then a little of the suspension was added to a new flask and placed in a cell incubator for further cultivation.

Example 4 Experiment on the effect of fluorobenzamide derivatives (BOF) on the survival rate of breast cancer MCF-7 cells

MCF-7 cells in log phase were collected, pipetted into a single cell suspension with a culture medium with a concentration of 10% FBS, counted with a counting plate, and adjusted the cell concentration to the desired concentration, 5 × 10 ³ cells per well. /100μL was inoculated in a 96-well plate and cultured in an incubator. After the cells adhered to the wall, 100 μL of drug-containing medium was added to each well (a blank control was set), and 3 duplicate wells were set for each drug concentration. After culturing for 24 hours, add 10 μL MTT to each well, incubate for 4 hours, discard the supernatant, add 100 μL DMSO to each well, put it on a shaker at room temperature, shake for 10 minutes, at a wavelength of 490 nm The absorbance value (OD value) of the 96-well plate was measured using a microplate reader, and the cell viability was calculated according to the absorbance value.

Figure 1 shows the survival rate of human breast cancer cells measured by MTT method. It can be seen from the figure that when the drug concentration is not less than 130mg/L, the survival rate of human breast cancer cells is much lower after BOF acts on human breast cancer cells. In the control group, there is a certain dose-effect relationship, which indicates that BOF can inhibit the growth of breast cancer cells.

Claims (8)

1. the application of fluorobenzamide derivative in the preparation of the medicine that suppresses breast cancer cell proliferation, and the chemical structure of described fluorobenzamide derivative is as follows:

2. The application of the fluorobenzamide derivative according to claim 1 in the preparation of a drug for inhibiting the proliferation of breast cancer cells, wherein the breast cancer cells are human breast cancer MCF-7 cell line. 3. the application of fluorobenzamide derivative in the preparation of anti-breast cancer medicine, the chemical structure of described fluorobenzamide derivative is as follows:

4. a fluorobenzamide derivative injection, is characterized in that, described injection contains fluorobenzamide derivative, cosolvent and isotonicity regulator, and described cosolvent is glycine, and described fluorine The chemical structures of benzamide derivatives are as follows:

5. according to the described fluorobenzamide derivative injection of claim 4, it is characterized in that, the concentration of each component in described injection is: Fluorobenzamide derivatives 2-10mg/ml Glycine 0.5-5mg/ml Isotonicity modifier 3-9mg/ml. 6. according to the described fluorobenzamide derivative injection of claim 5, it is characterized in that, the concentration of each component in described injection is: Fluorobenzamide derivatives 5-10mg/ml Glycine 1-4mg/ml Isotonicity modifier 6-9mg/ml. 7. according to the described fluorobenzamide derivative injection of any one of claim 4-6, it is characterized in that, described isotonicity regulator is selected from following any one: sodium chloride, glucose, mannitol, Sorbitol. 8. The fluorobenzamide derivative injection according to claim 7, wherein the isotonicity regulator is sodium chloride or glucose.

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