Preparation method and application of photosensitizer suitable for proton-induced dynamic radiotherapy

Abstract

The invention belongs to the field of photosensitizers, and particularly relates to a preparation method and application of a photosensitizer suitable for proton-induced dynamic radiotherapy. Comprising the following steps: S1, dissolving zinc acetate dihydrate in ethanol to obtain a reaction solution; s2, adding PEG (Polyethylene Glycol) into the reaction liquid; s3, adding a 25% methanol tetramethylammonium hydroxide solution into the reaction liquid; s4, standing the reaction liquid, and initiating nucleation and growth of ZnO nanoparticles through alkali-catalyzed hydrolysis and condensation reaction; s5, adding the reaction liquid into a normal hexane solution to precipitate the nano-particles; s6, the nanoparticle solution is centrifuged, and supernatant liquid is removed; and S7, suspending the nano-particles obtained by centrifugation in spectral grade ethanol, and washing to obtain PEG (at) ZnO NPs. The invention provides a preparation method of a photosensitizer suitable for proton-induced dynamic radiotherapy, and aims to solve the problems that the photosensitizer in the prior art is poor in radiation resistance and cannot be suitable for a proton irradiation environment.

Description

Technical Field

[0001] The present invention belongs to the field of photosensitizers, and particularly relates to a preparation method and application of a photosensitizer suitable for proton-induced radiotherapy Background Art

[0002] Proton-induced radiotherapy is a cutting-edge tumor treatment method that combines the precision of proton therapy with real-time dynamic adjustment technology. Its core principle is to activate photosensitizers through high-energy protons to achieve proton-induced radiotherapy. Compared with X-ray radiotherapy, the "Bragg peak" characteristic of protons can not only reduce the radiation dose to organs at risk but also achieve the directional activation of photosensitizers in tumor tissues, reduce the toxicity of photosensitizers to normal tissues behind the tumor, and synergistically enhance the protection of normal tissues.

[0003] However, in the prior art, conventional photosensitizers are made of organic nanomaterials and have poor anti-irradiation performance. High-energy proton irradiation may cause the loss or inactivation of the functions of these organic materials, ultimately resulting in unsatisfactory treatment effects of proton-induced radiotherapy. Summary of the Invention

[0004] The present invention provides a preparation method of a photosensitizer suitable for proton-induced radiotherapy, aiming to solve the problem that the photosensitizer in the prior art has poor anti-irradiation performance and cannot be applied to the proton irradiation environment.

[0005] To achieve the above object, the present invention provides a preparation method of a photosensitizer suitable for proton-induced radiotherapy, including the following steps

[0006] S1, Dissolve zinc acetate dihydrate in ethanol to obtain a reaction solution;

[0007] S2, Add PEG to the reaction solution;

[0008] S3, Add 25% methanol tetramethylammonium hydroxide solution to the reaction solution;

[0009] S4, Let the reaction solution stand, and initiate the nucleation and growth of ZnO nanoparticles through base-catalyzed hydrolysis and condensation reactions;

[0010] S5, Take the reaction solution and add it to the n-hexane solution to precipitate the nanoparticles;

[0011] S6, Centrifuge the nanoparticle solution and remove the supernatant;

[0012] S7, Suspend the centrifuged nanoparticles in spectroscopic-grade ethanol, wash, and obtain PEG@ZnO NPs.

[0013] This solution discloses a preparation method of a photosensitizer, and finally PEG@ZnO NPs are prepared. This kind of photosensitizer has strong anti-irradiation performance, can meet the use requirements of proton dynamic radiotherapy, and solves the technical shortcomings of conventional photosensitizers in the prior art.

[0014] Preferably, in the step S1, the concentration of the prepared reaction solution is 4 mol / L.

[0015] Preferably, in order to make the zinc acetate dihydrate dissolve more thoroughly, in the step S1 of this solution, the reaction solution is heated and stirred. The heating and stirring process helps the zinc acetate dihydrate dissolve.

[0016] Preferably, in the step S2, the added PEG has a molecular weight of 2000-6000, and the molar ratio of PEG to Zn²⁺ is 0.2-0.5.

[0017] Preferably, in the step S3, 10vol%-20vol% of 25% methanol tetramethylammonium hydroxide solution is added.

[0018] Preferably, in the step S4, the reaction solution is left to stand for 300-400 min.

[0019] Preferably, in the step S5, the volume ratio of the added reaction solution to the solution in n-hexane is 1:3-5.

[0020] Preferably, in the step S6, the centrifugation speed is 2000-5000 r / min, and the centrifugation is carried out for 15-20 minutes.

[0021] The second aspect of the present invention discloses a photosensitizer prepared by using the above-mentioned photosensitizer preparation method applicable to proton-induced dynamic radiotherapy.

[0022] The third aspect of the present invention discloses the application of the above-mentioned photosensitizer in the field of enhancing ROS generation.

[0023] The beneficial effects of the present invention are as follows: First, this solution discloses a preparation method of a photosensitizer, and finally PEG@ZnO NPs are prepared. This kind of photosensitizer has strong anti-irradiation performance, can meet the use requirements of proton dynamic radiotherapy, and solves the technical shortcomings of conventional photosensitizers in the prior art.

[0024] Second, the disclosed PEG@ZnO NPs in this solution rely on the passivation effect of PEG to modify the surface defects of nano-ZnO. After passivation, the surface defects of nano-ZnO lose the ability to capture electrons and holes, can significantly inhibit the recombination of electron-hole pairs of nano-ZnO at the defects, improve the utilization rate of electrons and holes, thereby improving the catalytic cracking of water by the photosensitizer material to generate more ROS, reducing the oxygen dependence of tumor treatment, and enhancing the killing effect on tumor cells. Brief Description of the Drawings

[0025] Figure 1 TEM image of the PEG@ZnO NPs composite nanomaterial prepared in Example 1.

[0026] Figure 2 Emission spectra of PEG@ZnO NPs, PEG, and ZnO under proton irradiation.

[0027] Figure 3 Emission spectrum of PEG under proton irradiation.

[0028] Figure 4 Evolution diagram of tumor cells with the concentration of PEG@ZnO NPs composite nanomaterial and irradiation dose.

[0029] Figure 5 XRD pattern of the PEG@ZnO NPs composite nanomaterial.

[0030] Figure 6 FTIR spectrum of the PEG@ZnO NPs composite nanomaterial.

[0031] Figure 7 FTIR spectra of PEG with a molecular weight of 4000 and different molar ratios of PEG to Zn²⁺. Detailed Description of the Invention

[0032] In order to make the objectives, technical solutions, and advantages of the embodiments clearer, the present invention will be further described in detail below in conjunction with the accompanying drawings and embodiments. When the following description refers to the accompanying drawings, unless otherwise indicated, the same numbers in different drawings represent the same or similar elements. The embodiments described in the following exemplary embodiments do not represent all embodiments consistent with the present disclosure. On the contrary, they are merely examples of devices and methods consistent with some aspects of the present disclosure as detailed in the appended claims.

[0033] In the present disclosure, unless otherwise stated, the orientation terms such as "inside, outside" are defined according to the contour of the corresponding component itself. The terms such as "first, second" used in the present disclosure are used to distinguish one element from another and do not have sequential and importance.

[0034] Example 1

[0035] A method for preparing a photosensitizer applicable to proton-induced dynamic radiotherapy, comprising the following steps:

[0036] S1. Dissolve zinc acetate dihydrate in ethanol to form a reaction solution. The concentration of the reaction solution is 4 mol / L. Meanwhile, to completely dissolve zinc acetate dihydrate in ethanol, in this embodiment, the reaction solution is heated and stirred for 30 minutes, and the heating temperature is 65°C.

[0037] S2. Add PEG to the reaction solution and stir during the addition process. The added PEG has a molecular weight of 4000, and in this embodiment, the molar ratio of PEG to Zn²⁺ is 0.4.

[0038] S3. Add a 10 vol% 25% methanol tetramethylammonium hydroxide (TMAOH) solution to the reaction solution.

[0039] S4. Let the reaction solution stand for 300 min - 400 min to initiate the nucleation and growth of ZnO nanoparticles through base-catalyzed hydrolysis and condensation reactions.

[0040] S5. Take 10 mL of the reaction solution and add it to 40 mL of a n-hexane solution to precipitate the ZnO nanoparticles.

[0041] S6. Place the nanoparticle solution into a centrifuge with a rotation speed of 5000 r / min and centrifuge for 18 min. Remove the supernatant to obtain the precipitated nanoparticles.

[0042] S7. Suspend the nanoparticles obtained by centrifugation in spectroscopic-grade ethanol and wash them 4 - 6 times to obtain PEG@ZnONPs.

[0043] The TEM image of the prepared PEG@ZnO NPs is as Figure 1 shown. It can be seen from the image that the diameter of the PEG@ZnO NPs is 5 - 20 nm. The small diameter of the nanoparticles is beneficial for the nanoparticles to penetrate from the blood into the tumor tissue.

[0044] Irradiate PEG@ZnO NPs, PGE, and ZnO with a 2 MeV proton beam, and collect the emission spectrum in real time during irradiation. The detection results are as Figure 2As shown, the intensity of the defect luminescence center of ZnO after PEG group modification is significantly reduced, which is mainly due to the passivation effect of PEG modification on the surface defects of nano-ZnO. The surface defects of the passivated nano-ZnO lose the ability to capture electrons and holes, which can significantly inhibit the recombination of electron-hole pairs of nano-ZnO at the defects, thereby improving the utilization rate of electrons and holes, increasing the catalytic water splitting of the photosensitizer material to generate more ROS, reducing the oxygen dependence of tumor treatment, and enhancing the killing effect on tumor cells. At the same time, PEG@ZnO NPs can still maintain their good electron and hole yields under MeV-level proton irradiation, and can be applied to the medical high-energy proton irradiation environment, solving the problem of poor anti-irradiation performance of existing photosensitizers, and at the same time can significantly reduce the oxygen dependence of tumor tissues.

[0045] At the same time, as Figure 3 shown, MeV-level proton irradiation of PEG alone proves that the PEG group shows no obvious inactivation under MeV-level proton irradiation and can be used as a functional modification material to reduce the biological toxicity of nano-inorganic photosensitizer materials and reduce the toxic side effects of inorganic materials on normal human tissues.

[0046] Perform proton-induced radiotherapy cell experiments on the PEG@ZnO NPs prepared in this example. HepG2 liver cancer cells were used as the cells for testing the toxicity of the photosensitizer material, and standard medium (DMEM) and 37°C, 5% CO2 were used as the cell culture conditions. The cells were seeded in 96-well plates (about 5×10³~1×10 4 cells / well) and cultured for 24 hours until they adhered to the wall.

[0047] The test results are as Figure 4 shown. The results show that the PEG@ZnO NPs composite nanomaterial has the effect of killing tumor cells under proton irradiation. At the same time, as the proton irradiation dose increases, the survival rate of tumor cells further decreases. When the concentration of PEG@ZnO NPs is 3 μg / mL and the radiation dose is 6 Gy, the survival rate of tumor cells is the lowest, proving that the photosensitizer prepared in this study has an obvious tumor cell killing effect under proton induction.

[0048] Example 2

[0049] The difference between this example and Example 1 is that the added PEG has a molecular weight of 2000.

[0050] The remaining steps are the same as those in Example 1 to prepare PEG@ZnO NPs.

[0051] Example 3

[0052] The difference between this example and Example 1 is that the added PEG has a molecular weight of 6000.

[0053] The remaining steps are the same as those in Example 1, and PEG@ZnO NPs are prepared.

[0054] The XRD images of the PEG@ZnO NPs obtained in Example 1, Example 2, and Example 3 are as Figure 5 shown. It can be seen from Figure 5 that the prepared PEG@ZnO NPs are in the ZnO phase, and the modification of the PEG group does not change the phase of the ZnO material.

[0055] The FTIR images of the PEG@ZnO NPs obtained in Example 1, Example 2, and Example 3 are as Figure 6 shown. Among them, the characteristic peaks within the dashed box correspond to the chemical bonds formed by the indirect grafting of ZnO and PEG. The test results show that the PEG groups in the three examples have been modified on the surface of nano-ZnO, and even if the PEG molecular weights are different, they can be anchored on the ZnO surface through bonding. In addition, the intensity of the characteristic peaks has a saturation concentration with the PEG molecular weight. When the PEG molecular weight exceeds 4000, the intensity of the characteristic peaks no longer increases. Therefore, PEG with a molecular weight of 4000 is preferably used, which combines modification efficiency and economy.

[0056] Example 4

[0057] The difference between this example and Example 1 is that in step S3 of this example, 20 vol% of a 25% methanol solution of tetramethylammonium hydroxide (TMAOH) is added to the reaction solution.

[0058] The remaining steps are the same as those in Example 1, and PEG@ZnO NPs are prepared.

[0059] Example 5

[0060] The difference between this example and Example 1 is that in step S5 of this example, 10 mL of the reaction solution is taken and added to 30 mL of a n-hexane solution to precipitate the ZnO nanoparticles.

[0061] The remaining steps are the same as those in Example 1, and PEG@ZnO NPs are prepared.

[0062] Example 6

[0063] The difference between this example and Example 1 is that in step S5 of this example, 10 mL of the reaction solution is taken and added to 50 mL of a n-hexane solution to precipitate the ZnO nanoparticles.

[0064] The remaining steps are the same as those in Example 1, and PEG@ZnO NPs are prepared.

[0065] Example 7

[0066] The difference between this example and Example 1 is that in S2 of this example, the added PEG has a molecular weight of 4000, and at the same time, the molar ratio of PEG to Zn²⁺ in this example is 0.5.

[0067] The remaining steps are the same as those in Example 1, and PEG@ZnO NPs are prepared.

[0068] Example 8

[0069] The difference between this example and Example 1 is that in S2 of this example, the added PEG has a molecular weight of 4000, and at the same time, the molar ratio of PEG to Zn²⁺ in this example is 0.3.

[0070] The remaining steps are the same as those in Example 1, and PEG@ZnO NPs are prepared.

[0071] Example 9

[0072] The difference between this example and Example 1 is that in S2 of this example, the added PEG has a molecular weight of 4000, and at the same time, the molar ratio of PEG to Zn²⁺ in this example is 0.2.

[0073] The remaining steps are the same as those in Example 1, and PEG@ZnO NPs are prepared.

[0074] The FTIR spectra of the PEG@ZnO NPs prepared in Example 1, Example 7, Example 8, and Example 9 are as Figure 7 shown, as shown in the figure.

[0075] When the PEG molecular weight is 4000, Figure 7 the characteristic peaks appearing in the range of 3000 - 2700 cm -1 correspond to the chemical bonds formed by the grafting of ZnO and PEG molecules. When PEG:Zn²⁺ is 20 mol% (i.e., 0.2) and 30 mol% (i.e., 0.3), the characteristic peak intensities are lower, indicating that the lower PEG content results in fewer formed chemical bonds. Subsequently, as the PEG content increases, the characteristic peak intensities gradually increase, and the characteristic peak intensities formed by PEG with 40 mol% (i.e., 0.4) and 50 mol% (i.e., 0.5) contents are similar, indicating that there is a saturation content for PEG molecule modification of nano-ZnO, that is, when the content is 40 mol%, the characteristic peak intensity reaches the highest. Therefore, it is more preferable that the molar ratio of PEG to Zn²⁺ is 0.4. At this content, the PEG modification efficiency is the highest, while avoiding the waste of PEG.

[0076] The above are only embodiments of the present invention, and common knowledge such as specific structures and characteristics known in the art are not described in detail herein. It should be noted that for those skilled in the art, without departing from the structure of the present invention, several modifications and improvements can be made, which should also be regarded as the protection scope of the present invention, and these will not affect the implementation effect of the present invention and the practicability of the patent. The protection scope claimed in this application shall be subject to the content of its claims, and the specific implementation manners and the like recorded in the specification can be used to interpret the content of the claims.

Claims

1. A preparation method of photosensitizer applicable to proton-induced dynamic radiotherapy, characterized in that: Including the following steps S1. Dissolve zinc acetate dihydrate in ethanol to obtain a reaction solution; S2. Add PEG to the reaction solution; S3. Add a 25% methanol solution of tetramethylammonium hydroxide to the reaction solution; S4. Let the reaction solution stand, and initiate the nucleation and growth of ZnO nanoparticles through base-catalyzed hydrolysis and condensation reactions; S5. Take the reaction solution and add it to the n-hexane solution to precipitate the nanoparticles; S6. Centrifuge the nanoparticle solution and remove the supernatant; S7. Suspend the centrifuged nanoparticles in spectroscopic-grade ethanol, wash them to obtain PEG@ZnO NPs.

2. The method for preparing a photosensitizer according to claim 1, wherein: In S1, the concentration of the obtained reaction solution is 4 mol / L.

3. The method for preparing a photosensitizer according to claim 1, wherein: In S1, heat and stir the reaction solution.

4. The method for preparing a photosensitizer according to claim 1, characterized in that: In S2, the added PEG has a molecular weight of 2000 - 6000, and the molar ratio of PEG to Zn²⁺ is 0.2 - 0.

5.

5. The method for preparing a photosensitizer according to claim 1, characterized in that: In S3, add 10 vol% - 20 vol% of the 25% methanol solution of tetramethylammonium hydroxide.

6. The method for preparing a photosensitizer according to claim 1, characterized in that: In S4, let the reaction solution stand for 300 - 400 min.

7. The method for preparing a photosensitizer according to claim 1, wherein: In S5, the volume ratio of the added reaction solution to the n-hexane solution is 1:3 - 5.

8. The method for preparing a photosensitizer according to claim 1, wherein: In S6, the centrifugation speed is 2000 - 5000 r / min, and centrifuge for 15 - 20 minutes.

9. A photosensitizer, characterized in that: Prepared by the method for preparing a photosensitizer applicable to proton-induced dynamic radiotherapy according to any one of claims 1 to 8.

10. Use of a photosensitizer as claimed in claim 9 in the field of enhancing ROS generation.

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