CN107802836B

CN107802836B - Tumor targeted photo-thermal medicament, preparation method and application

Info

Publication number: CN107802836B
Application number: CN201711311833.XA
Authority: CN
Inventors: 张先正; 马宁; 张明康; 曾旋
Original assignee: Wuhan University WHU
Current assignee: Wuhan University WHU
Priority date: 2017-12-11
Filing date: 2017-12-11
Publication date: 2020-09-08
Anticipated expiration: 2037-12-11
Also published as: CN107802836A

Abstract

The invention discloses a tumor targeted photo-thermal medicament, a preparation method and application thereof. The invention synthesizes the medicament with the photothermal antitumor effect by carrying out hydrophilic modification on molybdenum telluride through PEG with a targeting group modified by sulfydryl. The medicament has the characteristic of tumor cell targeting, and can realize the curative effect of inhibiting the proliferation of tumor cells through the photothermal effect.

Description

Tumor targeted photo-thermal medicament, preparation method and application

Technical Field

The invention belongs to the technical field of nano materials, and particularly relates to a tumor-targeted photo-thermal medicament, and a preparation method and application thereof.

Background

Photothermal therapy is an emerging therapeutic approach based on the generation of far infrared waves from light-activated materials. With the development of photothermal reagents such as conjugated organic groups, carbon nanotubes, gold nanorods, etc., photothermal therapy has been widely used for tumor imaging and treatment. The specificity and selectivity of photothermal agents plays a key role in diagnostic and therapeutic proceduresThe function of the bond. The geometrical structure of the nano material has great influence on the photo-thermal effect of the nano material. In recent years, MoS with nanometer-scale thickness₂，MoSe₂，WS₂，WSe₂Isotransition metal disulfides have attracted considerable attention. Due to the unique physical and chemical properties, the transition metal dichalcogenide has strong application potential in electronic components and parts, photocatalysis and energy storage. Inspired by electrochemical applications, more and more researchers are working on the applications of transition metal dichalcogenides with excellent optical and electronic properties for cancer imaging and therapy. For example, Liu [ 2 ] has been studied in a recent study of a combination therapy of photothermal and chemotherapyAdvanced materials2014, 26, 3433.]With his colleagues prepared a MoS₂And (4) nano flakes.

Molybdenum ditelluride is a gray hexagonal powdery solid. Can be decomposed in alkali, is insoluble in water, is soluble in nitric acid and is stable in air. Can be used as a novel semiconductor material instead of silicon. At present, MoTe is not seen₂Can be used for multifunctional therapy of cancer.

Disclosure of Invention

The invention solves the first technical problem by providing a preparation method of a tumor-targeted photothermal medicament. The method uses molybdenum ditelluride for multifunctional therapy of cancer, and prepares MoTe by mechanical ball milling method₂The nano-flake is functionalized by thiol reaction, so that the tumor targeting property and biocompatibility are endowed. The invention provides MoTe with excellent biocompatibility₂Nanoflakes exhibit strong near-infrared absorption, low toxicity and in vivo degradability properties, making them a promising agent for photothermal therapy.

The second technical problem to be solved by the invention is to provide the application of the tumor-targeted photothermal medicine prepared by the method.

In order to solve the first technical problem, the invention provides a preparation method of a tumor-targeted photothermal medicament, which comprises the following steps:

a) ball-milling the molybdenum telluride crystal in a high-speed ball mill for 24-36 h at the rotation speed of 200-800 rpm, uniformly dispersing the ball-milled powder in deionized water in an ice bath by using a sound wave degradation method,

b) mixing and stirring molybdenum telluride dispersed in water and sulfhydryl modified PEG with targeting groups according to the mass ratio of 1:1 for reaction for 48 h, and then centrifugally separating at 8000 rpm to obtain MoTe with excellent biocompatibility₂And (4) nano flakes.

The sound wave degradation method comprises the following steps: 0-10^oAnd C, stripping the ball-milled powder under the action of a cell crusher, wherein the crushing time is 6-12 h, and the obtained nano-sheet layer is 1-10 molecular layers.

Preferably, in step b), the targeting group is RGD, cRGD, HA, PAA.

Preferably, in step b), the molecular weight of PEG may be 200-.

Further, the invention provides the tumor-targeted photothermal agent prepared by the method.

In order to solve the second technical problem, the invention provides an application of the tumor-targeted photothermal agent in preparing an anti-tumor drug.

The starting materials of the present invention are commercially available, unless otherwise specified, and the equipment used in the present invention may be any equipment conventionally used in the art or may be any equipment known in the art.

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

the invention selects the molybdenum telluride nano material with high photothermal conversion rate as the photothermal reagent, avoids the use of heavy metal and high-toxicity aromatic compounds, and has low cost and fast in vivo metabolism compared with similar products in the market. The novel tumor-targeted photo-thermal medicament has strong absorption on near infrared light; has good inhibition effect on the growth of tumor cells and higher lethality on the tumor cells; the photo-thermal conversion efficiency is high; can induce apoptosis.

Drawings

FIG. 1: TEM images of molybdenum telluride photothermal antitumor agents.

FIG. 2: absorption of ultraviolet light by the antitumor agent.

FIG. 3: a. experiment of inhibiting growth of mouse breast cancer cell by using the anti-tumor photothermal agent. b. Experiment on growth inhibition of mouse breast cancer cells by the anti-tumor photothermal medicament after being irradiated by far infrared rays.

FIG. 4: the temperature distribution of the anti-tumor photo-thermal medicament under far infrared irradiation to the temperature of a water system.

FIG. 5: the influence of the far infrared radiation anti-tumor photo-thermal medicament on the system temperature changes along with the time and the influence of water on the system temperature is compared.

FIG. 6: analysis of apoptosis of cancer cells under different conditions of action.

Detailed Description

The features and advantages of the present invention will be further understood from the following detailed description taken in conjunction with the accompanying drawings. The examples provided are merely illustrative of the method of the present invention and do not limit the remainder of the disclosure in any way.

Example 1 synthesis of tumor-targeting photothermal agent 1:

(1) and ball-milling the molybdenum telluride crystals in a high-speed ball mill for 48 hours at the rotating speed of 400 rmp.

(2) Uniformly dispersing the ball-milled powder in deionized water by using a cell crusher, wherein the crushing time is 12 hours, and the temperature is 0-4 DEG^oC。

(3) Mixing molybdenum telluride dispersed in water with HS-PEG-cRGD (Mn = 25.7 kg/mol) according to the mass ratio of 1:1, stirring and reacting for 48 h, and then centrifuging at 8000 rpm to obtain MoTe with excellent biocompatibility₂And (4) nano flakes.

(4) The material structure was characterized by a projection electron microscope (fig. 1) and the obtained product was stored at-18 ℃ protected from light.

[ example 2 ] ultraviolet-visible absorption spectrum of tumor-targeted photothermal agent 1

MoTe₂The UV-visible absorption spectrum of the aqueous nanoflake solution was measured by a UV-Vis spectrophotometer (Thermoscientific, Waltham, Mass.). The results are shown in FIG. 2, MoTe₂The nanoplatelets absorb light in the ultraviolet and visible rangeThe spectrum shows strong near infrared absorption at 800-950 nm, thus proving MoTe₂The nano-flake is a compound with photo-thermal conversion potential, and the optimal absorption wavelength range is 800-950 nm.

[ example 3 ] tumor-targeting photothermal agent 1 acting on mouse breast cancer cells

Mouse breast cancer cells were seeded in 2 96-well plates at a density of 6000 cells/well, and cultured with 0.1 ml of a medium for 24 hours. Then, the culture medium is prepared into MoTe with different concentrations₂Nanoflake PBS solution was added to each well separately. The culture was maintained at 37 ℃ for 48 hours in the absence of light. The dark toxicity group was cultured for 24 hours under dark conditions. To evaluate the phototoxicity of the material, the material that has not been endocytosed was replaced with fresh medium after 6 hours of incubation and then irradiated with NIR laser. 808 nm near infrared light is used for each hole, and the power is 1W cm^-1Laser irradiation of (2) for 1 minute. The culture was continued for 24 hours. Then, 20. mu.l of MTT (dissolved in 5 mg/ml PBS buffer) was added to each well. After a total incubation time of 4 hours, the medium was aspirated and 150. mu.l of dimethyl sulfoxide was added. And (3) measuring the light absorption value at 570 nm in each hole by using a microplate reader, and calculating the cell survival rate to further obtain the phototoxicity and dark toxicity of the material obtained in the example 1 to the mouse breast cancer cells.

The results are shown in fig. 3, the material after illumination has great toxicity to mouse breast cancer cells, thereby proving that the photothermal anti-tumor compound has the characteristic of selectively killing tumor cells.

Example 4 testing of photo-thermal Properties of hydrophilic molybdenum telluride nanoflakes

In order to test the photo-thermal performance of the hydrophilic molybdenum telluride nanoflakes, the hydrophilic nanoflakes and the blank control were irradiated with 808 nm far infrared laser. The course of laser irradiation was recorded with an infrared thermal imager.

As a result, as shown in fig. 4 and 5, the temperature of the molybdenum telluride flake group was significantly increased with the increase of the irradiation time, and the blank group had almost no temperature change.

[ example 5 ] tumor-targeting photothermal agent 1 induces apoptosis of tumor cells

Mouse breast cancer cells were treated with 1 × 10⁵The cells were cultured in six-well plates at a density of one cell/well and in 2 ml of medium at 37 ℃. After 24 hours, medium with dissolved material and blank medium were added to the six-well plate. After further incubation for 6 hours, the cells were irradiated with a laser at 808 nm. In contrast to irradiation with laser light alone or addition of material alone without any treatment. After further incubation for 18 hours, all cells were trypsinized and harvested by centrifugation. After washing with PBS and annexin buffer, all cells were stained with FITC for 30 min and assayed by flow cytometry.

The results are shown in fig. 6, the material group after far infrared irradiation shows obvious apoptosis, which well verifies that the material has excellent photothermal effect.

Claims

1. The preparation method of the tumor-targeted photothermal agent is characterized by comprising the following steps:

a）MoTe₂preparing nano flakes: ball-milling the molybdenum telluride crystal in a high-speed ball mill for 24-36 h at a rotation speed of 200-800 rpm, then stripping the ball-milled powder under the action of a cell crusher to obtain a molecular layer composition with 1-10 nanosheets, uniformly dispersing the molecular layer composition in deionized water, wherein the crushing time is 6-12 h and the temperature is 0-10^oC；

b) Preparing a tumor-targeted photothermal medicament: MoTe to be dispersed in Water₂Mixing and stirring the nano-sheets and PEG modified by sulfydryl and having a targeting group according to the mass ratio of 1:1 for reaction for 48 hours, and then centrifugally separating at 8000 rpm to obtain MoTe with excellent biocompatibility₂And (4) nano flakes.

2. The method of claim 1, wherein in step b), the targeting group is RGD, cRGD, HA, PAA.

3. The method as claimed in claim 1, wherein the molecular weight of PEG in step b) is 200-5000.

4. A tumor-targeted photothermal agent prepared by the method of claim 1.

5. The use of the tumor-targeted photothermal agent of claim 4 in the preparation of an antitumor drug.