阅读说明：本技术 一种抗癌药物、抗癌药物组合物及其制备方法和用途 (Anticancer drug, anticancer drug composition, and preparation method and application thereof ) 是由 尤业字 古晗 聂旋 张泽 夏磊 高凡 黄伟强 于 2021-01-22 设计创作，主要内容包括：本发明提供一种抗癌药物、抗癌药物组合物及其制备方法和用途,其中,抗癌药物包括2-氨基-5-[(5-硝基-2-噻唑基)硫代]-1,3,4-噻二唑,抗癌药物组合物包括2-氨基-5-[(5-硝基-2-噻唑基)硫代]-1,3,4-噻二唑和苯硼酸类聚合物。本发明中的2-氨基-5-[(5-硝基-2-噻唑基)硫代]-1,3,4-噻二唑具有广谱的抗癌性,对多种癌症具有明显的抑制作用,同时对正常细胞的毒性低于对癌细胞的毒性；该小分子结构简单,溶血率低。抗癌药物组合物中苯硼酸类聚合物作为辅料,与抗癌药物通过供体-受体相互作用组合,帮助递送药物进入细胞内,苯硼酸对癌细胞中过量产生的活性氧敏感,过量活性氧可以选择性释放药物,从而增强抗癌作用。(The invention provides an anticancer drug, an anticancer drug composition, a preparation method and application thereof, wherein the anticancer drug comprises 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole, and the anticancer drug composition comprises 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and phenylboronic acid polymer. The 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole has broad-spectrum anticancer property, has obvious inhibiting effect on various cancers, and has lower toxicity on normal cells than cancer cells; the small molecule has simple structure and low hemolysis rate. The phenylboronic acid polymer in the anticancer drug composition is used as an auxiliary material and combined with an anticancer drug through donor-receptor interaction to help deliver the drug into cells, the phenylboronic acid is sensitive to active oxygen excessively generated in cancer cells, and the excessive active oxygen can selectively release the drug, so that the anticancer effect is enhanced.)

1. An anticancer drug comprises 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole or a derivative thereof with a biological electron isostere structure, wherein the structure of the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole is shown as the following formula I:

2. an anticancer pharmaceutical composition comprises 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and phenylboronic acid polymer, wherein the amount ratio of the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole to the phenylboronic acid is 2: 1-1: 8, and the phenylboronic acid polymer has a structure shown in the following formula II:

wherein m is an integer of 50 to 200, and n is an integer of 40 to 100.

3. The anticancer pharmaceutical composition according to claim 2, wherein said phenylboronic acid-based polymer and said amino group of 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole have coordination interaction.

4. The anticancer pharmaceutical composition of claim 2, wherein the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole of the phenylboronic acid-based polymer has an attachment rate of 30-80%.

Use of 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole or a derivative thereof having a bioisostere structure for the preparation of a medicament for the prevention and treatment of cancer.

6. Use of a composition comprising 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and a phenylboronic acid polymer for the preparation of a medicament for the prevention and treatment of cancer, wherein the mass ratio of the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole to the phenylboronic acid in the phenylboronic acid polymer is in the range of 2: 1 to 1: 8, and the phenylboronic acid polymer has the following structure represented by formula II:

wherein m is an integer of 50 to 200, and n is an integer of 40 to 100.

7. Use according to claim 5 or 6, wherein the cancer is a solid tumor.

8. The use according to claim 7, wherein the cancer comprises cervical cancer, breast cancer and colon cancer.

9. A method of making a composition comprising 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and a phenylboronic acid-based polymer, the method comprising:

s1, reacting 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and phenylboronic acid in the phenylboronic acid polymer in a first organic solvent according to the mass ratio of 2: 1-1: 8, so that the phenylboronic acid polymer and amino groups in the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole are subjected to coordination interaction;

s2, dialyzing the obtained composition with a second organic solvent to remove the non-loaded 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole.

10. The method of claim 9, wherein the first organic solvent comprises N, N-dimethylformamide, dimethylsulfoxide; the second organic solvent comprises water, a blending solvent of water and methanol.

Technical Field

The invention relates to the technical field of biological medicines, in particular to an anticancer drug, an anticancer drug composition, and a preparation method and application thereof.

Background

Cancer is a serious disease that seriously harms human life in today's society. In recent years, cancer has been treated by a variety of methods, including surgical therapy, radiation therapy, chemotherapy, hormonal therapy, and the like. Although various therapeutic approaches have been shown to be effective in cancer therapy, chemotherapy has gained importance in the treatment of cancer as a systemic treatment. However, chemotherapy drugs used clinically at present still have a plurality of problems, such as rapid removal of the drugs, poor selectivity, reduction of the curative effect of the drugs, systemic toxicity, large toxic and side effects, drug resistance generation after prolonging the use time of the drugs, and the like. Therefore, the finding of the drug with small toxic and side effects, low treatment dosage and selective release is of great significance.

Disclosure of Invention

Technical problem to be solved

Aiming at the problems, the invention provides an anti-cancer drug, an anti-cancer drug composition and application thereof, which are used for at least partially solving the technical problems of poor selectivity, large toxic and side effects and the like of the traditional chemotherapy drugs.

(II) technical scheme

The invention provides an anticancer drug, which comprises 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole or a derivative thereof with a biological electron isostere structure, wherein the structure of the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole is shown as the following formula I:

the invention also provides an anticancer pharmaceutical composition, which comprises 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and phenylboronic acid polymer, wherein the mass ratio of the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole to the phenylboronic acid is in the range of 2: 1 to 1: 8, and the phenylboronic acid polymer has the following structure shown in formula II:

wherein m is an integer of 50 to 200, and n is an integer of 40 to 100.

Furthermore, the coordination interaction occurs between the phenylboronic acid polymer and the amino group in the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole in the anticancer pharmaceutical composition.

Furthermore, the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole of the phenylboronic acid polymer has a loading rate of 30-80%.

In a further aspect, the invention provides the use of 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole or a derivative thereof having a bioisostere structure, or a composition comprising 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and a phenylboronic acid-based polymer, for the manufacture of a medicament for the prevention and treatment of cancer.

Further, the cancer is a solid tumor.

Further, cancers include cervical cancer, breast cancer, and colon cancer.

In a further aspect, the present invention provides a process for the preparation of a composition comprising 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and a phenylboronic acid based polymer, the process comprising: s1, reacting the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and phenylboronic acid in the phenylboronic acid polymer in a first organic solvent according to the mass ratio of 2: 1-1: 8, so that the phenylboronic acid polymer and amino groups in the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole are subjected to coordination interaction; s2, dialyzing the obtained composition with a second organic solvent to remove the non-loaded 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole.

Further, the first organic solvent includes N, N-Dimethylformamide (DMF), dimethyl sulfoxide (DMSO); the second organic solvent comprises water, a blending solvent of water and methanol.

(III) advantageous effects

The anti-cancer drug and the anti-cancer drug composition provided by the invention have an inhibiting effect on cancer, namely inhibiting the growth of cancer cells and inducing the apoptosis of the cancer cells, wherein the anti-cancer drug 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and the derivatives thereof with the bioisostere structure have the following advantages: 1) can effectively kill cancer cells, and has low toxicity to normal cells and small toxic and side effects; 2) the hemolysis rate is low and almost not hemolysis. The anticancer medicine composition has the following advantages: 1) excessive active oxygen in cancer cells can selectively release drugs, thereby enhancing the anticancer effect; 2) the potential of damaged mitochondrial membrane is enhanced, and the pH value is sensitive, and the anticancer effect is enhanced.

Drawings

FIG. 1 schematically shows the in vitro anticancer effects of an anticancer drug and an anticancer drug composition according to example 1 of the present invention;

FIG. 2 schematically shows the in vitro anticancer effects of the anticancer drug according to example 2 of the present invention at different pH;

FIG. 3 schematically shows the results of mitochondrial membrane potential change analysis of anticancer drugs and anticancer drug compositions according to example 3 of the present invention;

FIG. 4 is a schematic view showing hemolytic effects of the anticancer drug and the anticancer drug composition according to example 4 of the present invention.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is described in further detail below with reference to specific embodiments and the accompanying drawings.

The embodiment of the present disclosure provides an anticancer drug, including 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole or a derivative thereof having a bioisostere structure, wherein the structure of 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole is shown as the following formula I:

the anti-cancer drug disclosed by the disclosure has a broad spectrum and has a killing effect on various cancer cells, such as cervical cancer cells, breast cancer cells and drug-resistant cells resistant to other drugs. Meanwhile, the compound has low killing effect on normal cells, small toxic and side effects and better anticancer effect compared with other similar derivatives. The cell membrane is a selective functional barrier, important for protons and other ions, and the ion gradient can be used in the secondary transport process to drive the selective uptake of substrates and the excretion of metabolites. The proton kinetic potential (Δ p) consists of a pH gradient (Δ pH) and a transmembrane potential (Δ ψ), is an energy intermediate of cells, and can be used for driving processes of solute transport, ATP synthesis, and the like. The proton motive force is connected to each other by special transport proteins, also by H⁺The phosphoric acid potentials are connected. An increase in the passive flux of protons or ions through the membrane may lead to a decrease in Δ p (or sodium kinetics) that impairs the normal function of the membrane in energy transduction and screening of the cytoplasm from the environment. The membrane potential analysis shows that the 2-amino-5- [ (5-nitro-2-thiazolyl) thio]-1, 3, 4-thiadiazole and compositions thereof cause damage to mitochondrial membranes and compositions thereof exhibit enhanced damage to mitochondrial membranes, indicating their ability to dissipate a proton motive potential across membrane potential. While 2-amino-5- [ (5-nitro-2-thiazolyl) thio]The cell killing effect of 1, 3, 4-thiadiazole and compositions decreases with increasing pH, and the compounds may also have the ability to dissipate pH gradients in proton motive force. At the same time, the anticancer drug 2-amino-5- [ (5-nitro-2-thiazolyl) thio]The-1, 3, 4-thiadiazole has low hemolytic property and reaches the contentLess than 5% at 25. mu.g/mL, is considered to be non-hemolytic. 2-amino-5- [ (5-nitro-2-thiazolyl) thio]The-1, 3, 4-thiadiazole molecule has a simple structure, can kill cancer cells at a lower concentration, has weaker killing capability on normal cells, and has selectivity and high curative effect. Compared with the traditional antitumor drugs, the anticancer drug can be used for treating cancer without using a carrier, and has a great clinical application prospect.

Another embodiment of the present disclosure provides an anticancer pharmaceutical composition, comprising 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and phenylboronic acid polymer, wherein the mass ratio of the drug to the phenylboronic acid is in the range of 2: 1 to 1: 8, the phenylboronic acid polymer has the following structure represented by formula II, wherein m is an integer from 50 to 200, n is an integer from 40 to 100,

an anticancer composition comprising a phenylboronic acid structure capable of binding to the amino group of 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole via a donor-acceptor interaction means that is responsive to Reactive Oxygen Species (ROS), wherein excess ROS in cancer cells selectively releases the drug, thereby enhancing the anticancer effect, and wherein excess ROS is not present in normal cells and is poorly released in normal cells. Meanwhile, the anti-cancer composition has enhanced damage capability to mitochondrial membrane potential and enhanced sensitivity to pH value, and has stronger anti-cancer effect than 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole.

The mass ratio of the substances is that the ratio of the phenylboronic acid in the compound of formula II to the anticancer drug is 2: 1-1: 8, and the loading rate of the phenylboronic acid polymer to the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole is 30% -80%. In the polymer, a chain segment where phenylboronic acid is located is a hydrophobic chain segment, and a chain segment where ethylene glycol is located is a hydrophilic chain segment, and the chain segment can form a micelle structure in water. In the invention, the phenylboronic acid is firstly coordinated with the medicament and then dispersed in water to form micelles. The ratio of the phenylboronic acid to the drug is 1: 1, so that the ratio is 1: 1, which is the most efficient, and 1: 1 is also used in the examples.

Based on the above examples, the combination of the anticancer pharmaceutical composition is such that the phenylboronic acid polymer and the amino group of 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole have specific coordination interaction.

The anti-cancer pharmaceutical composition forms boron-nitrogen coordination by boron in phenylboronic acid and nitrogen in 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole, and the coordination action enables the composition to have responsiveness to ROS, can effectively kill cancer cells, and has small toxicity to normal cells and small toxic and side effects.

The polymerization degree of the chain segment containing phenylboronic acid in the phenylboronic acid polymer is 50-200, and the polymerization degree of the chain segment containing ethylene glycol is 40-100. The phenylboronic acid polymer within the polymerization degree range can be used for carrying more anticancer drugs and cannot be difficult to dissolve in water due to overlarge molecular weight.

The present disclosure also provides a method of preparing a composition based on coordination interactions of phenylboronic acid with an amino group, the method comprising:

s1, reacting 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and phenylboronic acid compounds in an organic solvent N, N-Dimethylformamide (DMF) or dimethyl sulfoxide (DMSO) to obtain a coordination interaction structure.

S2, putting the obtained composition into water or a blending solvent of water and methanol to dialyze excessive un-bonded 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole.

Embodiments of the present disclosure also provide an anticancer pharmaceutical composition comprising 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and a phenylboronic acid-based polymer according to the foregoing, for use in preventing and treating cancer.

2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole is combined with an auxiliary material phenylboronic acid polymer, so that the anticancer effect is enhanced, and the damage to normal cells is reduced; meanwhile, the anti-cancer composition has enhanced damage capability to mitochondrial membrane potential and enhanced sensitivity to pH value, and has stronger anti-cancer effect than 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole.

The present invention is further illustrated by the following specific embodiments. The reagents used in the examples were purchased from national pharmaceutical group chemical reagents, Inc. and Shanghai Biyun Biotechnology, Inc., and the cells were purchased from Shanghai cell Bank of Chinese academy.

Example 1

In vitro anticancer evaluation of 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and its composition with phenylboronic acid polymer

The in vitro anticancer effect is measured by adopting an MTT method: adding cancer cells (HeLa) into 96-well plate, incubating in incubator for 24 hr, adding 100 μ L cell liquid per well, and inoculating number of 6 × 10³To 8X 10³And the number of inoculations per well was about the same. 2-amino-5- [ (5-nitro-2-thiazolyl) thio]-1, 3, 4-thiadiazole (formula I) and composition are formulated in a concentration of 0.5mg/mL, wherein DMF is used as solvent for obtaining a structure of coordination interaction, and wherein 2-amino-5- [ (5-nitro-2-thiazolyl) thio is present in the anticancer pharmaceutical composition]The ratio of the amount of the phenylboronic acid substance in the-1, 3, 4-thiadiazole to the amount of the phenylboronic acid polymer is 1: 1, the degree of polymerization of the phenylboronic acid in the phenylboronic acid polymer is 100, and the degree of polymerization of the ethylene glycol in the segment is 44. Sucking the culture medium, adding the prepared culture medium containing the anticancer drugs and the composition with different concentrations (the concentration of the composition is calculated according to the content of the anticancer drugs inoculated into the culture medium), wherein the concentration of the anticancer drugs is 0.5 mu g/mL to 10 mu g/mL, and incubating for 24h in an incubator. After the medium was aspirated and 100. mu.L of thiazole blue (MTT) solution was added to each well for incubation for 4 hours, the MTT solution was aspirated and 120. mu.L of dimethyl sulfoxide (DMSO) was added to each well. Measuring the light absorption value at 490nm with enzyme-linked immunosorbent assay device, and drawing cell survival rate curve according to the comparison with the light absorption value of control group (adding no anticancer drug and only culture medium in the drug adding step). Meanwhile, in mouse colon cancer cell CT26, humanSimilar in vitro anti-tumor tests were performed in breast cancer cells MDA-MB-231 and mouse fibroblast (normal cell) L929. Fig. 1 shows that the anticancer drug and the composition have significant cytotoxic effects on various cancer cells, and have less toxic effects on normal cells, and the anticancer drug composition has stronger killing effect on cancer cells at a lower concentration. In addition, the cells incubated with the anticancer drug were observed with an optical microscope, and it was found that when the anticancer drug concentration was higher, the cells became rounded, apoptotic, and finally became dead cells in suspension, indicating that the anticancer drug would alter the cell morphology and cause apoptosis.

Example 2

Proton motive force related evaluation of 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole

Proton kinetic potential related experiments were evaluated by the MTT method. Taking 5 centrifuge tubes, adding 10mL culture medium with serum, adjusting pH value with 1M hydrochloric acid and 1M sodium hydroxide to 5.4, 6.4, 7.4, 8.4 and 9.4 respectively for standby. Adding cancer cells (HeLa) into 96-well plate, incubating in incubator for 24 hr, adding 100 μ L cell liquid per well, and inoculating number of 6 × 10³To 8X 10³And the number of inoculations per well was about the same. 2-amino-5- [ (5-nitro-2-thiazolyl) thio]The 1, 3, 4-thiadiazole (formula I) is prepared at a concentration of 0.5 mg/mL. Taking culture media with different pH values, respectively preparing culture media containing anticancer drugs with different concentrations, wherein the concentration of the anticancer drugs is 0.25 mu g/mL to 10 mu g/mL, sucking the original culture media, adding the newly-prepared culture media containing the anticancer drugs with different pH values, and incubating for 24h in an incubator. After the medium was aspirated and 100. mu.L of MTT solution was added to each well for incubation for 4 hours, the MTT solution was aspirated and 120. mu.L of dimethyl sulfoxide (DMSO) was added to each well. Measuring the light absorption value at 490nm with enzyme linked immunosorbent assay device, and drawing the cell survival rate curve at different pH values according to the comparison with the light absorption value of control group (adding no anticancer drug and only adding culture medium in the drug adding step). FIG. 2 shows that the anticancer drug is highly toxic at low pH, and the toxicity of the anticancer drug to cells is gradually reduced as the pH is gradually increased.

Example 3

Evaluation of Membrane potential Change in 2-amino-5- [ (5-Nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and composition

Cells are dyed by a mitochondrial membrane potential detection kit JC-1, color change is observed by a fluorescence microscope, and membrane potential is evaluated, wherein the kit JC-1 is purchased from Shanghai Biyunnan biotechnology limited and has the model of C2006. Adding cancer cells (HeLa) into 24-well plate, incubating in incubator for 24 hr, adding 400 μ L cell liquid per well, and inoculating 2 × 10 cells per well⁴To 4X 10⁴And the number of inoculations per well was about the same. Sucking out the original culture medium, adding the prepared 2-amino-5- [ (5-nitro-2-thiazolyl) thio]Medium with 1, 3, 4-thiadiazole (formula I) concentration of 10. mu.g/mL, incubated for 4h in the incubator. An appropriate amount of JC-1(200X) was taken and JC-1 was diluted at a rate of adding 8mL of ultrapure water to 50. mu.L of JC-1 (200X). Vigorous vortexing to dissolve and mix JC-1 well. Then adding 2mL JC-1 staining buffer solution (5X) and mixing uniformly to obtain the JC-1 staining working solution. The medium was aspirated, 400. mu.L JC-1 staining solution was added to each well, and mixed well. The cells were incubated at 37 ℃ for 20 minutes in a cell incubator. During the incubation, an appropriate amount of JC-1 staining buffer (1X) was prepared at a rate of 4mL of distilled water per 1mL of JC-1 staining buffer (5X) and placed in an ice bath. After incubation at 37 ℃ was complete, the supernatant was aspirated and washed 2 times with JC-1 staining buffer (1X). 1mL of serum-containing medium was added, observed under a fluorescence microscope, and photographs were taken on both red and green channels (red channel: λ ex 543nm, green channel: λ ex 488 nm). Calculating average fluorescence intensity of red and green channels by using Image J software, and calculating lambda_543nm/λ_488nmThe ratio of (a) to (b). When the mitochondrial membrane potential is higher, JC-1 is gathered in a mitochondrial matrix to form a polymer, and red fluorescence is generated; when the mitochondrial membrane potential is low, JC-1 cannot aggregate in a mitochondrial matrix, and JC-1 can generate green fluorescence as a monomer. For anticancer pharmaceutical composition (wherein 2-amino-5- [ (5-nitro-2-thiazolyl) thio in anticancer pharmaceutical composition]Similar operations were carried out with a ratio of 1: 1 of-1, 3, 4-thiadiazole to the amount of phenylboronic acid species in the phenylboronic acid-based polymer. FIG. 3 shows the combination of the control group (medium without anticancer drug added during the dosing step) with the anticancer drug group and the anticancer drugResults of analysis of the mitochondrial membrane potential of the group. Lambda [ alpha ]_543nm/λ_488nmA smaller ratio of (A) indicates a higher degree of membrane destruction, a higher degree of transmembrane potential reduction, and a higher degree of (. DELTA.. psi.) dissipation. From the analysis results, it can be seen that the destructive effect of the composition on the membrane potential is strongest.

Example 4

Evaluation of hemolysis of 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole and composition

The hemolytic properties of the anti-cancer drugs and compositions were evaluated by whether the drugs rupture red blood cells at various concentrations. About 1mL of blood is taken from the eye socket of an ICR mouse, an appropriate amount of PBS is added to the blood and mixed evenly, the mixture is placed into a centrifuge to be centrifuged for 3min at 800r/min, the supernatant is discarded, and the lower layer of red blood cells are left. The cells were washed twice more with PBS and the supernatant was clarified to leave the lower layer of red blood cells. 0.8mL of red blood cells were added to PBS to prepare a 0.2% red blood cell suspension. 900 μ L of erythrocyte suspension is taken out by a 1mL centrifuge tube, 5 μ g/mL to 50 μ g/mL of 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole (formula I) and a composition (the concentration of the composition is calculated by the content of the anticancer drug which is inoculated in the composition) are respectively added into the erythrocyte suspension, wherein the ratio of the 2-amino-5- [ (5-nitro-2-thiazolyl) thio ] -1, 3, 4-thiadiazole in the anticancer drug composition to the amount of the phenylboronic acid substance in the phenylboronic acid polymer is 1: 1, and the erythrocyte suspension is incubated for 1 hour in an incubator at 37 ℃ after being uniformly mixed. After incubation, the cells were centrifuged at 800r/min for 3min in a centrifuge, 100. mu.L of the supernatant was transferred to a 96-well plate and the light absorption was measured at 576nm using an ELISA. The control groups of the experiment were PBS group and triton group, PBS group was negative control, and triton group was positive control. FIG. 4 shows the hemolysis rate of the anti-cancer drugs and the composition, and the hemolysis rate is still lower than 10% when the drug content is 50 μ g/mL, which proves that the drugs have no hemolysis and can be well applied in the organism without the influence of hemolysis.

The above-mentioned embodiments are intended to illustrate the objects, technical solutions and advantages of the present invention in further detail, and it should be understood that the above-mentioned embodiments are only exemplary embodiments of the present invention, and are not intended to limit the present invention, and any modifications, equivalents, improvements and the like made within the spirit and principle of the present invention should be included in the protection scope of the present invention.