CN109912607B

CN109912607B - A class of porphyrin-chrysin complexes and their antitumor activity

Info

Publication number: CN109912607B
Application number: CN201811508080.6A
Authority: CN
Inventors: 刘运美; 刘鼎; 张奇志; 李洋; 杨静; 刘振华; 何军
Original assignee: University of South China
Current assignee: University of South China
Priority date: 2018-12-11
Filing date: 2018-12-11
Publication date: 2021-01-22
Anticipated expiration: 2038-12-11
Also published as: CN109912607A

Abstract

The present invention synthesizes a new type of porphyrin-chrysin derivatives, takes porphyrin molecules as carriers, utilizes its tumor tissue aggregation effect, and utilizes its property of producing singlet oxygen to kill tumor cells, combined with chrysin's properties. Natural anti-tumor activity, a new class of anti-tumor compounds is obtained, which provides a new direction for the research of anti-tumor drugs.

Description

Porphyrin-chrysin compound and antitumor activity thereof

Technical Field

The invention relates to the field of medicinal chemistry, in particular to a porphyrin-chrysin compound, a preparation method and application thereof.

Background

In modern society, the incidence of cancer is higher and higher, and the mortality rate is always high. Traditional chemotherapy drugs nonspecifically block cell division to cause cell death, and when they kill cancer cells, they also destroy the growth of normal cells of the human body, bringing about many toxic and side effects.

Photodynamic therapy is a new technology which is applied to tumor treatment only in the late 1970 s, but the development is rapid, and the photodynamic therapy is allowed to enter clinical application by related departments of many countries such as the United states, the United kingdom, Germany, Japan and the like. To date, this therapy has been successfully used to treat a variety of malignancies.

The basic elements of photodynamic therapy can be divided into: photosensitizer, specific exciting light and molecular oxygen. After the patient takes the photosensitizer for several hours, the concentration of the photosensitizer in the tumor tissue is obviously higher than that of the normal tissue, then the focus part is irradiated by the wavelength light with proper energy, and simultaneously, under the condition of the existence of molecular oxygen, the photosensitizer transfers the absorbed energy to the surrounding oxygen molecules, so that the oxygen molecules are excited to obtain excited singlet oxygen (A)¹O₂) And other Reactive Oxygen Species (ROS) to kill tumor cells. Moreover, unlike usual clinical laser treatment, laser irradiation in PDT has a low energy density in order to activate the photosensitizer without causing tissue damage. The first photo-sensitive drug available on the market, Photofri, approved by the FDA in the united states, is the classical porphyrin-structured drug.

Liang Cheng et al synthesized a porphyrin molecule (TCPP-PEG) with a long polyethylene glycol chain, hopefully made the drug effect and the side effect to reach the optimum proportion, through the fluorescence localization analysis, the renal clearance test and the in vivo anti-tumor experiment, obtained better effect.

Hu et al synthesized a novel gallium porphyrin, not only improved the water-solubility of porphyrin macromolecule, but also connected cisplatin and anti-platinum on porphyrin molecule, combined porphyrin's photodynamic curative effect and cisplatin's own anti-tumor therapy effect, obviously improved the anticancer effect of compound.

Gallium porphyrin-cisplatin/anti-platinum complex

Chrysin (5, 7-dihydroxyflavone) is a flavone compound widely existing in the nature, and has wide biological activities of antibiosis, antioxidation, antitumor, anti-inflammation and the like. Recent studies have shown that it can also prevent organ toxicity caused by cisplatin and improve cognitive deficits and brain damage caused by intermittent hypoxia. However, due to its poor water solubility, it is easily absorbed by the intestine and is subject to metabolic inactivation in vivo. In order to improve the pharmacological activity, the structure modification and reconstruction are carried out on the compound, and the compound has important significance for obtaining novel high-efficiency low-toxicity candidate drugs.

In conclusion, the compounds with porphyrin structures are potential antitumor drugs with great research prospects, and scientists have intensively researched and actively developed the compounds from the structures, the singlet oxygen generation capacity, the tumor localization and the antitumor mechanism; the chrysin is a natural flavonoid active substance, has wide activity, has certain research foundation for structural modification and activity research of chrysin, and is worthy of deep exploration in the aspect of anti-tumor cell proliferation.

The invention effectively combines porphyrin and chrysin antitumor compounds by a proper method to prepare a novel porphyrin-chrysin derivative. The compound of the invention takes porphyrin molecules as a carrier, utilizes the tumor tissue aggregation effect of the porphyrin molecules and the property that the porphyrin molecules can generate singlet oxygen to kill tumor cells, combines the natural antitumor activity of chrysin to obtain a novel antitumor compound, and provides a new direction for the research of antitumor drugs.

Disclosure of Invention

The technical problem to be solved by the invention is as follows: the porphyrin-chrysin compound is prepared with porphyrin molecule as carrier, and through the tumor tissue aggregation effect, the property of singlet oxygen capable of killing tumor cell and the combination of the natural antitumor activity of chrysin, porphyrin-chrysin compound is obtained.

In a first aspect of the present invention, there is provided a compound of formula I and pharmaceutically acceptable salts thereof, having the structure:

formula I;

wherein n is an integer from 2 to 6;

r is selected from OH.

Preferably, n is selected from 2, 4 or 6; more preferably n is selected from 2 or 4.

In another aspect of the invention, there is provided a process for the preparation of a compound of formula I, the synthetic route for which is as follows:

；

wherein n and R are as defined above.

The specific reaction steps are as follows:

porphyrin derivative 1, a base and potassium iodide were added to a 100mL three-necked flask, and DMF was added as a solvent, followed by stirring and refluxing. After 30min, the chrysin derivative 2 is added, the reflux reaction is continued for about 8h, and the reaction is monitored by TLC until the raw material point is unchanged. The cooled reaction solution was poured into a separatory funnel, and the reaction product was extracted with a dichloromethane solvent and washed with water. And (3) after the organic solvent is dewatered, drying the solvent by using a rotary evaporator, separating and purifying the residual solid by using silica gel column chromatography, and recrystallizing the obtained solid again to obtain the compound shown in the formula I.

Preferably, the molar ratio of porphyrin derivative 1 to chrysin derivative 2 is: 1 (1-1.5), preferably 1:1-1.2, more preferably 1:1.2

The base is selected from potassium hydroxide, triethylamine or potassium carbonate, more preferably potassium carbonate.

In another aspect of the present invention, a pharmaceutical composition is provided, which comprises a compound represented by formula I or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier or excipient.

In another aspect, the invention relates to the use of a compound of formula I, and pharmaceutically acceptable salts thereof, or a pharmaceutical composition comprising the same, in the preparation of an anti-cancer medicament;

preferably, the cancer is selected from gastric cancer or cervical cancer; in particular to a human gastric cancer cell strain MGC-803 or a human cervical cancer cell strain Hela.

Compared with the prior art, the invention has the beneficial effects that:

(1) the invention provides a new porphyrin-chrysin compound with anticancer activity, which widens the range of the existing anticancer compounds and can be continuously optimized as a lead compound;

(2) the compound takes porphyrin molecules as a carrier, has targeting effect on tumor cells by utilizing the tumor tissue aggregation effect, and reduces the killing side effect on normal cells;

(3) the periphery of the preferable compound of the invention is provided with a plurality of hydroxyl groups, so that the compound forms a protonation and deprotonation balance in a tumor cell microenvironment, and the compound forms more molecular states under the weak acid environment of tumor cells, and can enter the tumor cells more easily so as to play a cell inhibition effect.

Drawings

FIG. 1 is a graph showing the inhibition of the concentration of Compound 4a on Hela cells.

FIG. 2 is the effect of Compound 4a on the cycle of Hela cells.

FIG. 3 is the effect of Compound 4a on apoptosis of Hela cells.

Detailed Description

The present invention will be described in detail with reference to examples. In the present invention, the following examples are intended to better illustrate the present invention and are not intended to limit the scope of the present invention. Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.

Example 1 Synthesis of porphyrin-chrysin complexes

。

A100 mL three-necked flask was charged with 2 (110.1 mg)/3 (118.1 mg)/4 (101.8 mg), (0.15 mmol), and the appropriate amounts of baked potassium carbonate and potassium iodide were added as catalysts, and 40 mL of DMF was added as a solvent, and the reaction was stirred under reflux at 80 deg.C (65 deg.C, 4). After 30min chrysin derivative 1a (1 b, 1c, 1d, 1 e) (0.18 mmol) was added and the reaction was continued at reflux for about 8h, monitored by TLC until no further change in the feed point. The cooled reaction solution was poured into a separatory funnel, and the reaction product was extracted with a dichloromethane solvent, and washed with water several times to remove a DMF solvent and impurities such as potassium carbonate and potassium iodide. And (3) after the organic solvent is dewatered, drying the solvent by using a rotary evaporator, separating and purifying the residual solid by using silica gel column chromatography, and recrystallizing the obtained solid again to obtain the mauve/reddish brown solid.

The yields and characterization data for the corresponding compounds are as follows:

2a (23.3 mg, 0.023 mmol), yield 15.3%. ¹H NMR (400 MHz, CDCl₃) δ 12.80 (s, 1H), 8.90 (d, J = 4.7 Hz, 2H), 8.84 (s, 6H), 8.14 (t, J = 8.2 Hz, 8H), 7.91 (d, J = 7.0 Hz, 2H), 7.75 (d, J = 8.2 Hz, 6H), 7.59 - 7.50 (m, 3H), 7.34 (d, J = 8.3 Hz, 2H), 6.71 (s, 1H), 6.68 (s, 1H), 6.55 (s, 1H), 4.65 (s, 2H), 4.60 (s, 2H), -2.83 (s, 2H); MS (MALDI-TOF) m/z: calcd for C₆₁H₃₉Cl₃N₄O₅ ⁺1012.20 [M + H]⁺, found 1012.12.

2b (27.1 mg, 0.026 mmol), yield 17.6%. ¹H NMR (500 MHz, CDCl₃) δ 12.76 (s, 1H), 8.89 (d, J = 4.2 Hz, 2H), 8.86 - 8.75 (m, 6H), 8.12 (t, J = 8.5 Hz, 8H), 7.91 (d, J = 6.5 Hz, 2H), 7.74 (d, J = 7.1 Hz, 6H), 7.53 (d, J = 7.1 Hz, 3H), 7.31 (d, J = 8.2 Hz, 2H), 6.69 (s, 1H), 6.64 (s, 1H), 6.50 (s, 1H), 4.47 (t, J = 5.7 Hz, 2H), 4.43 (t, J = 5.9 Hz, 2H), 2.55 - 2.45 (m, 2H), -2.83 (s, 2H).

2c (28.8 mg, 0.028 mmol), yield 18.4%. ¹H NMR (500 MHz, CDCl₃) δ 12.76 (s, 1H), 8.91 (s, 2H), 8.83(s, 6H), 8.13 (t, J = 8.6 Hz, 8H), 7.89 (d, J = 7.2 Hz, 2H), 7.74 (d, J = 7.9 Hz, 6H), 7.50 (t, J = 7.5 Hz, 3H), 7.30 (d, J = 7.9 Hz, 2H), 6.69 (s, 1H), 6.59 (s, 1H), 6.46 (s, 1H), 4.36 (s, 2H), 4.26 (s, 2H), 2.21 (s, 4H), -2.84 (s, 2H).

2d (40.4 mg, 0.038 mmol), yield 25.5%. ¹H NMR (500 MHz, CDCl₃) δ 12.75 (s, 1H), 8.91 (d, J = 4.2 Hz, 2H), 8.83 (s, 6H), 8.12 (dd, J = 13.4, 8.3 Hz, 8H), 7.86 (d, J = 6.7 Hz, 2H), 7.74 (d, J = 8.0 Hz, 6H), 7.50 (dt, J = 14.2, 7.5 Hz, 3H), 7.29 (d, J = 8.4 Hz, 2H), 6.65 (s, 1H), 6.56 (d, J = 2.0 Hz, 1H), 6.44 (d, J = 2.0 Hz, 1H), 4.30 (t, J = 6.1 Hz, 2H), 4.17 (t, J = 6.3 Hz, 2H), 2.14 - 1.97 (m, 4H), 1.86 (dd, J = 15.0, 8.0 Hz, 2H), -2.84 (s, 2H).

2e (41.4 mg, 0.038 mmol), yield 25.8%. ¹H NMR (500 MHz, CDCl₃) δ 12.74 (s, 1H), 8.91 (s, 2H), 8.84 (s, 6H), 8.12 (dd, J = 14.5, 8.1 Hz, 8H), 7.86 (d, J = 6.9 Hz, 2H), 7.74 (d, J = 7.8 Hz, 6H), 7.48 (d, J = 7.5 Hz, 3H), 7.29 (d, J = 8.2 Hz, 2H), 6.65 (s, 1H), 6.54 (s, 1H), 6.42 (s, 1H), 4.27 (t, J = 5.9 Hz, 2H), 4.13 (t, J = 6.1 Hz, 2H), 2.04 - 2.02 (m, 2H), 2.00 - 1.93 (m, 2H), 1.79 - 1.65 (m, 4H), -2.83 (s, 2H).

3a (25.5 mg, 0.025 mmol), yield 16.9%. ¹H NMR (500 MHz, CDCl₃) δ 12.79 (s, 1H), 8.97 - 8.69 (m, 8H), 8.12 (d, J = 8.3 Hz, 8H), 7.92 (d, J = 6.5 Hz, 2H), 7.57 - 7.50 (m, 3H), 7.28 (d, J = 8.4 Hz, 8H), 6.71 (s, 1H), 6.65 (d, J= 1.7 Hz, 1H), 6.53 (s, 1H), 4.53 (s, 4H), 4.09 (s, 9H), -2.73 (s, 2H); MS (MALDI-TOF) m/z: calcd for C₆₄H₄₈N₄O₈ ⁺ 1000.35 [M + H]⁺, found 1000.31.

3b (26.5 mg, 0.026 mmol), yield 17.4%. ¹H NMR (500 MHz, CDCl₃) δ 12.76 (s, 1H), 8.86 (d, J = 5.3 Hz, 8H), 8.12 (d, J = 7.2 Hz, 8H), 7.91 (dd, J = 7.5, 1.6 Hz, 2H), 7.52 (d, J = 6.9 Hz, 3H), 7.29 (t, J = 7.2 Hz, 8H), 6.70 (s, 1H), 6.65 (d, J = 2.0 Hz, 1H), 6.51 (d, J = 2.0 Hz, 1H), 4.60 - 4.34 (m, 4H), 4.10 (s, 9H), 2.69 - 2.26 (m, 2H), -2.75 (s, 2H).

3c (35.2 mg, 0.034 mmol), yield 22.8%. ¹H NMR (500 MHz, CDCl₃) δ 12.77 (s, 1H), 8.87 (s, 8H), 8.12 (d, J = 8.3 Hz, 8H), 7.91 - 7.76 (m, 2H), 7.57 - 7.41 (m, 3H), 7.28 (d, J = 8.4 Hz, 8H), 6.68 (s, 1H), 6.59 (d, J = 2.0 Hz, 1H), 6.46 (d, J = 2.1 Hz, 1H), 4.32 (s, 2H), 4.24 (d, J = 5.2 Hz, 2H), 4.09 (s, 9H), 2.19 (s, 4H), -2.75 (s, 2H).

3d (43.4 mg, 0.042 mmol), yield 27.7%. ¹H NMR (500 MHz, CDCl₃) δ 12.74 (s, 1H), 8.86 (s, 8H), 8.12 (d, J = 8.0 Hz, 8H), 7.87 (d, J = 6.6 Hz, 2H), 7.49 (t, J = 7.4 Hz, 3H), 7.29 (d, J = 8.3 Hz, 8H), 6.66 (s, 1H), 6.57 (d, J= 2.0 Hz, 1H), 6.44 (d, J = 1.9 Hz, 1H), 4.30 (t, J = 6.1 Hz, 2H), 4.17 (t, J= 6.3 Hz, 2H), 4.10 (s, 9H), 2.13 - 1.98 (m, 4H), 1.91 - 1.78 (m, 2H), -2.75 (s, 2H).

3e (43.9 mg, 0.041 mmol), yield 27.6%. ¹H NMR (500 MHz, CDCl₃) δ 12.75 (s, 1H), 8.88 (s, 8H), 8.12 (t, J = 7.6 Hz, 8H), 7.86 (d, J = 6.7 Hz, 2H), 7.47 (d, J = 7.2 Hz, 3H), 7.28 (d, J = 8.3 Hz, 8H), 6.66 (s, 1H), 6.55 (s, 1H), 6.43 (s, 1H), 4.24 (t, J = 6.2 Hz, 2H), 4.12 (t, J = 6.4 Hz, 2H), 4.08 (s, 9H), 2.04 - 1.99 (m, 2H), 1.98 - 1.90 (m, 2H), 1.75 - 1.64 (m, 4H), -2.74 (s, 2H).

4a (23.0 mg, 0.024 mmol), yield 15.7%. ¹H NMR (500 MHz, DMSO) δ 12.88 (s, 1H), 9.98 (s, 3H), 8.86 (d, J = 11.9 Hz, 8H), 8.14 (d, J = 6.3 Hz, 4H), 8.00 (d, J = 7.9 Hz, 6H), 7.71 - 7.56 (m, 3H), 7.43 (d, J = 8.2 Hz, 2H), 7.21 (d, J = 8.1 Hz, 6H), 7.10 (s, 1H), 7.02 (s, 1H), 6.59 (s, 1H), 4.65 (t, J = 6.2 Hz, 2H), 4.56 (t, J = 6.2 Hz, 2H), -2.90 (s, 2H); MS (MALDI-TOF) m/z: calcd for C₆₁H₄₂N₄O₈ ⁺ 958.30 [M + H]⁺, found 958.17.

4b (23.7 mg, 0.024 mmol), yield 16.3%. ¹H NMR (400 MHz, dmso) δ 12.83 (s, 1H), 9.98 (s, 3H), 8.87 (s, 6H), 8.81 (s, 2H), 8.10 (d, J = 8.1 Hz, 4H), 8.03 - 7.97 (m, 6H), 7.57 (d, J = 7.4 Hz, 3H), 7.38 (d, J = 8.6 Hz, 2H), 7.20 (d, J = 8.2 Hz, 6H), 7.04 (s, 1H), 6.94 (s, 1H), 6.50 (s, 1H), 4.16 (t, J = 5.9 Hz, 2H), 3.54 (t, J = 6.4 Hz, 2H), 2.18 - 1.94 (m, 4H), -2.90 (s, 2H).

4c (27.4 mg, 0.027 mmol), yield 17.8%. ¹H NMR (500 MHz, DMSO) δ 12.81 (s, 1H), 9.98 (s, 3H), 8.87 (s, 6H), 8.83 (s, 2H), 8.09 (d, J = 8.2 Hz, 2H), 8.05 (d, J = 7.5 Hz, 2H), 8.02 - 7.98 (m, 6H), 7.59 - 7.48 (m, 3H), 7.35 (d, J = 8.3 Hz, 2H), 7.20 (d, J = 8.1 Hz, 6H), 7.00 (s, 1H), 6.86 (s, 1H), 6.43 (s, 1H), 4.26 (t, J = 6.1 Hz, 2H), 4.17 (t, J = 6.3 Hz, 2H), 1.96 - 1.89 (m, 2H), 1.88 - 1.80 (m, 2H), 1.61 (dd, J = 15.4, 9.9 Hz, 4H), -2.91 (s, 2H).

example 2 detection of antitumor cell Activity in vitro by MTT colorimetric method

Inoculating cells: separately digesting the gastric cancer cell MGC-803 and cervical cancer cell Hela in monolayer culture with 0.25% trypsin, and adding 10% calf serum (containing 1 × 10)⁵ U·L^-1Penicillin and 1X 10⁵ U·L^-1Streptomycin) RPMI-1640 medium, cultured at 5X 10 per well in 96-well plates³And inoculating each cell to prepare a cell suspension, wherein the volume of each cell is 150 mu L. The blank group was supplemented with only the same amount of culture medium. And a blank PBS solution is added to the outermost circle of the inoculated plate. Culturing the cells: the seeded 96-well plates were transferred to 37 ℃ with 5% CO₂The incubator continues to incubate for about 48 hours until the cell monolayer has spread to the bottom of the well.

And carefully absorbing the supernatant in the holes of the 96-hole plate by using a pipette gun, and adding 150 mu L of the compound solution to be detected, which is prepared in advance, into each hole. In this experiment, experimental group 1 and experimental group 2 (porphyrin-chrysin derivative), a positive control group (5-fluorouracil) and a blank control group were designed. 150 muL of drugs with different concentration gradients are added into the experimental groups 1 and 2 and the positive control group respectively. Drug (including positive control) concentration gradients were finally determined to be 16, 32, 64, 128 μ M. Each compound concentration was repeated in 3 wells. Blank control 150. mu.L of RPMI 1640 containing 10% fetal bovine serum was added to each well for cultureAnd (4) liquid. All plates were placed at 37 ℃ and 5% CO₂The incubator of (1) was incubated for 48 h, wherein after the first 4 hours, the experimental group 2 was taken out, the upper layer of the culture solution was carefully aspirated by a pipette and the culture solution containing no drug was added again, and the incubation was continued for 10 min with an LED violet lamp with a power of 12W 20 cm from a 96-well plate, after which the incubation was continued in the incubator.

The plate supernatant was discarded, and 20. mu.L of MTT solution (5 mg. multidot.mL) was added to each well^-1) And mixing them. Continuing culturing for 4 hr, removing supernatant with pipette gun, adding 100 μ L dimethyl sulfoxide into each well, shaking at constant speed for 10 min, zeroing with microplate reader, measuring absorbance (OD) at 490 nm wavelength in each well, and calculating IC of each drug₅₀Values, results were calculated for 3 replicates for each drug (including experimental groups 1, 2) and control.

Data on antitumor cell activity of compound

As can be seen from the above table, in the porphyrin-chrysin series derivatives, the antitumor activity of compound 4 series was the best, the antitumor activity of compound 2 series was the next to the antitumor activity of compound 3 series was the worst. The 4a, 4b compound has the best anti-tumor cell proliferation activity, and the activity of acting on Hela of human cervical carcinoma cells is better than that of chrysin and 5-FU positive control drug. In particular compound 4a, which is active against Hela cell proliferation IC₅₀The average value reaches 26.51 mu M. The compound 2a, 2c and 2e has higher MGC-803 cell inhibition rate under the illumination condition than chrysin and is similar to 5-FU. Whereas the activity of the whole compound 3 series was poorly inhibitory to both tumor cells.

In addition, the photodynamic therapy effect of porphyrin is simulated in the MTT experimental process in an overlapping mode, and the data show that the cytotoxicity of the compound in the illumination group is obviously stronger than that in the non-illumination group. The 4a, 4b, 2a, 2c and 2e compounds have higher antitumor activity under the illumination condition, and the illumination condition of the activity of the 4a compound is 5 times that of the activity of the 4a compound under the non-illumination condition. This demonstrates that the compound is produced¹O₂Has inhibitory effect on tumor cell proliferation.

Hela cells were treated with compound 4a at concentrations of 1, 2, 4, 8, 16, 32, 64, and 128. mu.M, respectively, and the log of the concentration was taken to obtain a graph of concentration versus cell viability (see FIG. 1). As can be seen from the attached figure 1, activity screening is performed on Hela cells according to 8 concentration gradients, the inhibition rate of the compound 4a on the Hela cells is obviously concentration-related, and the survival rate of the Hela cells is only 25% when the concentration is 128 mu M.

Example 3 cell cycle distribution experiment

Adding medicine: hela cells were seeded in 6-well plates at about one hundred thousand cells per well and placed in an incubator for 24 h. 2 secondary holes are arranged at each concentration by 0 (Control), 20, 40 and 60 mu M drug (4 a) concentration gradient, and the culture is continued for 48 h after the drug adding treatment. Wherein, after the first 4 hours, the pore plates with 4 concentrations are taken out, an LED purple lamp with the power of 12W is used for continuously irradiating for 10 min at a distance of 20 cm from the 6 pore plates, and after the irradiation is finished, the culture is put into an incubator for continuous culture.

Fixing: the drug-containing medium was aspirated and washed with PBS, the cells were digested with trypsin, the cell suspension was centrifuged and washed three times with PBS. 1 mL of 70% ethanol at 4 ℃ was added to each sample, and after resuspending the cells, the lid was closed and placed in a 4 ℃ environment for 12 h.

Dyeing: after 12 h, the cells were again washed twice by centrifugation in the same manner. 0.5 mL of stain solution (buffer + PI stain + RNase A) was added to each sample to resuspend the cells. Incubating the mixture at 37 ℃ for 30min in a dark environment and then carrying out flow detection.

The experimental results are shown in fig. 2, and it can be seen from the figure that, as the concentration of the drug increases (0, 20, 40, 60 μ M respectively), the percentage of Hela cells in the G1 phase gradually increases (69.67%, 72.64%, 75.77%, 78.85%), while the percentage of cells in the S phase and the G2 phase gradually decreases, so we can conclude that the compound 4a can effectively inhibit the proliferation of human cervical cancer cells Hela, and mainly inhibits the G1 phase of cell proliferation and is concentration-dependent.

Example 4 apoptosis assay

Hela cells were plated at 1X 10 per well⁵The cells were seeded in 6-well plates and placed at 37 ℃ in 5% CO₂The incubator is 24 h. Drug (4 a) was applied to the wells in a concentration gradient of 0, 20, 40, 60 μ M, 0 μ M being the control, plus PBS. The culture was continued for 48 h. After the first 4 hours, the experimental group is taken out, an LED purple lamp with the power of 12W is used for continuously irradiating for 10 min at a distance of 20 cm from a 6-hole plate, and the experimental group is placed into an incubator for continuous culture after the irradiation is finished. Cells were digested by adding trypsin, and then Annexin-V-FITC and 10. mu.L PI were added to the cell suspension, respectively. Incubation was continued for 10 min at room temperature in the absence of light. And finally, detecting by using a flow cytometer to obtain a result.

The experimental results are shown in fig. 3, in the four-quadrant graph shown in the figure, Q1 represents mechanically damaged necrotic cells, Q2 represents late apoptotic cells, Q3 represents early apoptotic cells, and Q4 represents normal cells. We can find that in the control group (0 μ M), normal cells account for 82.6% of the total cell number and apoptotic cells (Q2 + Q3) are 15.53%. When the drug concentration is 60 mu M, the normal cells are only 10.3%, and the apoptotic cell increase is 88.58%. Therefore, the compound 4a can remarkably induce Hela cell apoptosis and shows remarkable concentration dependence (20 mu M, 34.31%; 40 mu M, 66.89%; 60 mu M, 88.58%).

Claims

1. a compound shown in formula I and a pharmaceutically acceptable salt thereof, it has the following structure:

Among them, n=2;

R is selected from OH.

2. a method for preparing the compound of formula I as claimed in claim 1, is characterized in that: reaction scheme is as follows:

Wherein, the definition of n, R is as described in claim 1;

The specific reaction steps are as follows:

Porphyrin derivative 1, alkali and potassium iodide were added to a 100 mL three-necked flask, and DMF was added as a solvent, and the reaction was stirred and refluxed. After 30 min, chrysin derivative 2 was added, and the reflux reaction was continued for about 8 hours. No change, pour the cooled reaction solution into a separatory funnel, extract the reaction product with dichloromethane solvent, wash with water, remove water from the organic solvent, spin dry the solvent with a rotary evaporator, and separate and purify the remaining solid by silica gel column chromatography, The resulting solid is recrystallized again to give the compound of formula I.

3. preparation method according to claim 2 is characterized in that: the mol ratio of porphyrin derivative 1 and chrysin derivative 2 is: 1:(1-1.5).

4. The preparation method according to claim 3, wherein the molar ratio of the porphyrin derivative 1 to the chrysin derivative 2 is 1:1-1.2.

5. preparation method according to claim 4 is characterized in that: the molar ratio of porphyrin derivative 1 and chrysin derivative 2 is 1:1.2.

6. The preparation method according to any one of claims 2-5, wherein the base is selected from potassium hydroxide, triethylamine or potassium carbonate.

7. preparation method according to claim 6 is characterized in that: described alkali is potassium carbonate.

8. A pharmaceutical composition comprising the compound of formula I according to claim 1 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

9. Use of the compound of claim 1 or a pharmaceutically acceptable salt thereof or the pharmaceutical composition of claim 8 in the preparation of a medicament for the treatment of cancer, wherein the cancer is gastric cancer or cervical cancer.

10. The use according to claim 9, wherein the cancer is cancer caused by human gastric cancer cell MGC-803 and cancer caused by human cervical cancer cell Hela.