TWI538691B - Preparation method of radiation-sensitive copolymer carrier for nanoparticles with chemotherapeutic pharmaceuticals - Google Patents

Description

Method for preparing nanometer drug carrier of radiation-sensitive copolymer

The present invention relates to a method for preparing a radiation-sensitive copolymer nanomedicine carrier, and more particularly to a method for preparing a carrier for a radiation-sensitive copolymer nanomedicine for proton therapy.

According to the statistics of the Department of Health, cancer (malignant tumors) is still the top of the top ten causes of death among Chinese people, with lung cancer and liver cancer ranking first and second. The cause of cancer is caused by human cytopathic diseases. At present, most commonly used treatments in the clinic are surgery, radiation therapy, chemotherapy, target therapy, etc. Different treatment methods have different effects and indications for the lesions. .

The basic principle of radiotherapy is to use radioactivity to block the long strands of DNA double helix in the nucleus of cancer cells, thereby killing cancer cells or inhibiting their growth, and giving tumor cells an ideal dose to achieve therapeutic effects. However, often the y-ray or x-ray produced by traditional photon therapy passes through the human body, and as the radiation enters the depth of the tissue, the relative energy also decays exponentially. Many other normal tissues have been affected before the tumor cells have been destroyed. However, the physical property of protons is that when the proton beam penetrates human tissue, its energy can also increase with distance, and at the end of the range (ie, where the tumor is located), the velocity is reduced, and the maximum energy is instantaneously released, resulting in a Bragg Peak. ), giving cancer cells a high dose of effective treatment without damaging other healthy tissues. Because proton therapy has the characteristics of spread-out Bragg peak (SOBP), in addition to reducing the risk of damage to normal tissues, the side effects of radiation therapy will be reduced to a minimum.

Protons are different from the physical properties of X-rays. X-rays can treat tumors in deep tissues because of their strong penetrating power. However, the shortcoming is that they penetrate into the tumor process. The amount of tissue in front is higher than that of tumors. The tissue still has a considerable residual dose that is likely to damage adjacent normal tissue. Protons, when passing through the tissue, release a small amount of energy, which releases a large amount of energy when it reaches the depth of the tumor to be treated, called the Bragg peak, and the normal tissue behind the tumor is completely absent. Since the single Prague peak is not wide, it is necessary to combine several Prague peaks to expand the size of the tumor to enhance the effect of proton therapy. Proton radiation therapy is the most advanced tumor radiotherapy technology in the world. Proton radiation therapy has much less damage to normal cells around the lesion area, and relatively fewer side effects. It is expected that proton therapy will become more common in the future.

Traditional irradiation therapy uses X-ray to locate and treat cancerous tumors, and it is impossible to accurately control the exact position and dose, and the normal tissue between the body surface and the tumor may also receive the dose and be damaged. Therefore, there is a need for a therapeutic device and method that can accurately position and apply an appropriate dose. At present, the most common method is to use proton therapy. X-ray computed tomography (XCT) imaging system is used to locate the tumor contour and apply the most appropriate dose, but it is difficult to locate using X-ray computed tomography.

U.S. Patent Publication No. US 2007/0031337 A1 discloses that a proton tomographic image can utilize gold nanoparticle particles to have good binding properties to antibodies, thereby attracting antigens in cancer cells to achieve a better therapeutic position. The positioning and control of the applied drug dose. It is expected that the Proton Computed Tomography (PCT) photographic system may become a trend in the future.

The object of the present invention is to provide a method for preparing a radiation-sensitive copolymer nano-medicine carrier, which utilizes diselenide and 3-aminopropylpoly(ethylene glycol) polymer to each other. After the reaction, a diselenide block copolymer (Diselenide Block Co-polymer) is obtained as a nano drug carrier, which itself has a hydrophilic-hydrophobic property, so that a diselenide block copolymer (Diselenide) can be obtained by an emulsification method. Block Co-polymer) self-assembles into nanospheres.

Moreover, during the emulsification process, a bis-stearoylphosphatidylethanolamine-polyethylene glycol-biomarker polymer which is also hydrophobic and hydrophobic is added, so that the hydrophobic end is in the process of self-assembly of the polymer. By arranging and arranging each other by the presence of an organic solvent, and exposing the hydrophilic end to the external aqueous solution, a stable nanosphere structure is formed, so that when the organic solvent is volatilized, the nanosphere particles are due to internal hydrophobicity. Produces a water repellent effect.

Therefore, the water molecules will enter the structure and further damage the structure of the nanoparticle. In addition, since the outer surface of the nanosphere is a hydrophilic end, it can be stably present in the aqueous solution, and in the present research, it has developed a radiation-sensitive type. The diselenide block copolymer, so the nanosphere structure prepared by using the diselenide block copolymer can not only stably exist in the aqueous solution, but also can utilize the specific radiation wave to control the disintegration speed of the carrier structure. Therefore, it has the potential to develop into a carrier of radiation drugs.

The following is a preferred embodiment of the preparation method of the radiation-sensitive copolymer nano-medicine carrier of the present invention, which comprises at least the following steps: Step S11: taking 0.05 mol of solid sodium hydroxide (NaOH) 2 g of solution After 25 ml of water, a solution of 3.95 g of 50 mol of selenium powder and 100 mg of cetyltrimethylammonium bromide was added to prepare a selenium (Se) solution. Step S12: Another 6.6 m sodium borohydride (NaBH4) 0.25 g and 0.2 g of solid sodium hydroxide were added, and 5 ml of an aqueous solution was dissolved in an ice bath, and the solution was stirred under a nitrogen atmosphere. While dropping into the selenium solution obtained in the step S1, reacting at room temperature for 1 hour, and then reacting at about 90 ° C for about half an hour to complete the reaction, the obtained reddish brown solution is sodium selenide (Sodium selenide, Na2Se2) alkaline aqueous solution. Step S13: after dissolving the dodecenyl succinic anhydride (2-Dodecen-1-yl-succinic anhydride) in tetrahydrofuran (Tetrehydrofuran, THF for short), and then dissolving with the alkaline aqueous solution of sodium selenide obtained in step S2, After about 12 hours of reaction, the impurities were separated by column chromatography and dried at a high temperature to obtain a product of the product diselenide. Step S14: The diselenide product obtained in the step S3 is dissolved in tetrahydrofuran, and the polyglycol with the amine group is added to the polymer, and the cross-linking agent 1-ethyl-(3-dimethylaminopropane) is added. Base for the reaction of 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) or N-Hydroxysuccinimide (NHS) for about 12 hours, using a tube The impurities are separated by column chromatography and dried at a high temperature to obtain a final product, a diselenide block copolymer (Diselenide Block Co-polymer), for subsequent coating of the radiation nanoparticles or/and a chemotherapeutic drug.

Preparation of nanomedicine coated with radiation nanoparticles or/and chemotherapeutic drugs by using the above dis selenide block copolymer as a nano drug carrier and distearyl phosphatidylethanolamine-polyethylene glycol-biolabel There are three methods, which are described below.

1. Radiation-doped nanoparticles (RNPs): 10 mg of diselenide block copolymer (Diselenide Block Co-polymer) as a main component of the sphere, and an emulsifier that assists in stabilizing the structure. 2 stearoylphosphatidylethanolamine-polyethylene glycol-biomarker (DSPE-PEG-biomarker) 2 mg, dissolved in 5 ml of ultrapure water, and then added with oil phase irradiated nanoparticles ( Radiated nanoparticles) 4 mg of Dichloromethane organic solvent 1 ml, of which diselenide block copolymer: distearoylphosphatidylethanolamine-polyethylene glycol-biological target: oil phase radiation nano The ratio of the particles is 5:1:2, and after the ultrasonic wave is shaken in an ice bath to complete the emulsified state, the organic solvent is removed by heating to about 60 ° C to obtain a radioactive naphthalene having a size of about 100 nm. Radiation-sensitive nanosphere particles (RNPs-Radiation-Sensitive nanoparticles).

2. Coated chemotherapeutic drug (drug): Take 10 mg of diselenide block copolymer and 2 mg of distearoylphosphatidylethanolamine-polyethylene glycol-biological target, and dissolve in 5 ml of ultrapure water. And then add 1 ml of dichloromethane organic solvent containing 4 mg of chemotherapeutic drug Doxorubicin, wherein the diselenide block copolymer: distearoylphosphatidylethanolamine-polyethylene glycol-biological target : The ratio of the chemotherapeutic drug doxorubicin is 5:1:2, and after the ultrasonic wave is shaken in an ice bath to complete the emulsified state, the organic solvent of methylene chloride is removed by heating to about 60 ° C to obtain a final product of about 100 nm. Rice-sized radiation-sensitive nanosphere particles (DOX- Radiation-Sensitive nanoparticles).

3. Coated radiation nanoparticles and chemotherapeutic drugs: 10 mg of diselenide block copolymer and 2 mg of distearoylphosphatidylethanolamine-polyethylene glycol-biological target, dissolved in ultrapure water 5 In the milliliter, add 1 ml of methane chloride organic solvent dissolved in oil phase radiation nanoparticles and 2 mg of chemotherapeutic drug, wherein the diselenide block copolymer: distearoylphosphatidylethanolamine-polyethylene Alcohol-biological target: oil phase radiation nanoparticle: the ratio of the chemotherapy drug doxorubicin is 5:1:1:1, and after ultrasonic wave shock in an ice bath to complete the emulsified state, heat to about 60 ° C. The organic solvent of methyl chloride is removed to obtain a radiation-sensitive nanosphere particle (RNPs/DOX- Radiation-Sensitive nanoparticles) carrying a radiation nanoparticle and doxorubicin having a final product of about 120 nm.

When proton therapy is carried out using the nanomedicine prepared by the nanoparticle drug carrier of the radiation-sensitive copolymer of the present invention, when high-energy protons are injected into the body, the proton energy decreases with penetration depth. When just touching a tumor, the proton energy may have been reduced to 1/3-1/4 of the initial energy, depending on the incident energy and penetration depth. When protons hit 238U on the tumor, if the proton energy is 10–1000 MeV, there will be a chance to cause 238U to produce a nuclear fission reaction. The first picture is a proton (high-energy proton incident energy of 100 MeV) impinges on uranium. 238 produces a splitting reaction, and the proportion of occurrence of the cleavage product is distributed as a function of the mass number. These cleavage products are usually unstable nuclear species and will continue to undergo decay reactions.

Table 1 lists the nuclear species names and their associated decay reaction data of higher proportion of split products, including protons (high-energy proton incident energy of 10~250 MeV), which impinge on uranium-238 to produce a splitting reaction, and a higher proportion of split products The name of the nuclear species and its associated decay response data. These cleavage products emit high-energy electrons during the decay process, so that after the proton therapy is completed, the patients can still use the electrons generated by these decays to destroy the cancer cells of the tumor and enhance the therapeutic effect.

Table 1         <TABLE style="BORDER-BOTTOM: medium none; BORDER-LEFT: medium none; BORDER-COLLAPSE: collapse; BORDER-TOP: medium none; BORDER-RIGHT: medium none; mso-border-alt: double windowtext 1.5pt; Mso-yfti-tbllook: 1184; mso-padding-alt: 0cm 5.4pt 0cm 5.4pt; mso-border-insideh: 1.5pt double windowtext; mso-border-insidev: 1.5pt double windowtext"_0004"MsoTableGrid" border=" 1" cellSpacing="0" cellPadding="0"><TBODY><tr><td></td><td> Yield (%) </td><td> Half-life </td><td> Decay Mode </td><td> decay energy y (keV) (strength%) </td></tr><tr><td>^135碲</td><td> 23.0 < /td><td> 19.0 s </td><td> %Beta-=100 </td><td> Beta-: 5960 (50%), 5356 (25%), 5090 (%16) </td ></tr><tr><td>⁹¹<b>锶</b></td><td> 10.8 </td><td> 9.63 h </td><td > %Beta-=100 </td><td> Beta-: 2707 (29%), 1402 (25%), 1127 (%34.7) </td></tr><tr><td>⁹⁹molybdenum</td><td> 9.9 </td><td> 65.94 h </td><td> %Beta-=100 </td><td> Beta-: 1214 (82.4%) ,437 (16.4%) </td></tr><tr><td>¹⁴⁰镧</td><td> 7.3 </td><td> 1.6781 d </td> <td> %Beta-=100 </td><td> Beta-: 1679 (19.2%), 1246 (5.7%), 1240 (%10.9) </td></tr><tr><td>< Sup>132碲</sup></td><td> 5.5 </td><td> 3.204 d </td><td> %Beta-=100 </td><td> Beta-: 215(100 %) </td></tr><tr><td>¹⁴⁵镧</td><td> 4.8 </td><td> 24.8 s </td><td> % Beta-=100 </td><td> Beta-: 4110 (15%), 4040 (19%), 3992 (%15) </td></tr><tr><td><sup>141< /sup><b>钡</b></td><td> 4.8 </td><td> 18.27 m </td><td> %Beta-=100 </td><td> Beta-: 3023 (7%), 2746 (18%), 2565 (%25) </td></tr><tr><td>¹⁴⁶镧</td><td> 4.3 </ Td><td> 6.27 s </td><td> %Beta-=100 </td><td> Beta-:6530 (18%), 6272 (28%), 5603 (%5.5) </td> </tr><tr><td>¹³⁵<b>iodine</b></td><td> 4.2 </td><td> 6.57 h </td><td> %Beta-=100 </td><td> Beta-:1388 (23.8%), 1083 (8%), 970 (%21.9) </td></tr></TBODY></TABLE>

As can be seen from the above, the method for producing a radiation-sensitive copolymer nano-medicine and a carrier thereof of the present invention has the advantages of improving the dose of the cancer cell, effectively improving the accuracy of applying the drug, and has an industry. Utilization, novelty and progress.

The above description is only a preferred embodiment of the invention and is not intended to limit the scope of the invention. That is, the equivalent changes and modifications made by the scope of the patent application of the present invention are covered by the scope of the invention.

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Figure 1 shows that the protons of the present invention (high-energy proton incident energy is 100 MeV) collide with uranium-238 to produce a splitting reaction, and the proportion of split products is distributed as a function of mass.

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Claims (7)

A method for synthesizing a composite nanomedicine carrier for proton therapy comprises at least the following steps: Step S11: after dissolving solid sodium hydroxide in water, adding selenium powder and cetyltrimethylammonium bromide, and arranging Selenium solution; Step S12: Another sodium borohydride and solid sodium hydroxide are taken, dissolved in an aqueous solution under an ice bath, and the solution is added to the selenium solution obtained in the step S11 while stirring under a nitrogen atmosphere. After reacting at room temperature for 1 hour, and then reacting at about 90 ° C for about half an hour to complete the reaction, the obtained reddish brown solution is an alkaline aqueous solution of sodium selenide; Step S13: taking dodecenyl succinic anhydride After being dissolved in tetrahydrofuran, and then being mutually miscible with the alkaline aqueous solution of sodium selenide obtained in the step S12, after reacting for about 12 hours, the impurities are separated by column chromatography, and dried at a high temperature to obtain a product diselenide product; Step S14 : the product obtained in the step S13 is dissolved in tetrahydrofuran, and the polyglycol with the amine group is added to the polymer, and the cross-linking agent 1-ethyl-(3-dimethylaminopropyl)carbonium is added. Diimine or N-hydroxybutyl After the imide is reacted for about 12 hours, the impurities are separated by column chromatography, and dried at a high temperature to obtain a final product diselenide block copolymer for subsequent coating of radiation nanoparticles or/and chemotherapy drugs. . A method for synthesizing a composite nanomedicine for proton therapy using the diselenide block copolymer prepared in claim 1, comprising at least: taking the diselenide block copolymer and a distearyl phosphatidylethanolamine-polyethylene glycol-biological target, which is dissolved in ultrapure water, and then added to a dichloromethane organic solvent in which oil phase radiation nanoparticles are dissolved, wherein the diselenide is embedded Segment Copolymer: distearoylphosphatidylethanolamine-polyethylene glycol-biological target: oil phase The ratio of the radioactive nanoparticles is 5:1:2, and after the ultrasonic wave is shaken in an ice bath to complete the emulsified state, the organic solvent is removed by heating to about 60 ° C to obtain a size of about 100 nm. Radiation-sensitive nanosphere particles carrying radioactive nanoparticles. A method for preparing a proton-treated composite nanomedicine for coating a chemotherapeutic drug by using the diselenide block copolymer of claim 1, comprising at least: taking the diselenide block copolymer and the second hard a fatty acylphosphatidylethanolamine-polyethylene glycol-biological target, dissolved in ultrapure water, and then added to a dichloromethane organic solvent in which a chemotherapeutic drug is dissolved, wherein the diselenide block copolymer: distearyl Acylphosphatidylethanolamine-polyethylene glycol-biological target: the ratio of chemotherapeutic drugs is 5:1:2, and after ultrasonic wave shock in an ice bath to complete the emulsified state, the organic acid is heated to about 60 ° C. The solvent is removed to obtain a radiation-sensitive nanosphere particle of a final product of about 100 nanometers in size. A method for synthesizing a composite nanomedicine for proton therapy using the diselenide block copolymer of claim 1 to prepare a coated nanoparticle and a chemotherapeutic drug, comprising at least: taking the diselenide block The polymer and distearoylphosphatidylethanolamine-polyethylene glycol-biological target are mutually dissolved in ultrapure water, and then added with a dichloromethane organic solvent in which oil phase radiation nanoparticles and chemotherapeutic drugs are dissolved, wherein Selenide block copolymer: distearoylphosphatidylethanolamine-polyethylene glycol-biological target: oil phase radiation nanoparticles: the ratio of chemotherapy drugs is 5:1:1:1, and under ice bath After the ultrasonic vibration is completed and the emulsified state is completed, the dichloromethane organic solvent is removed by heating to about 60 ° C to obtain radiation-sensitive nanosphere particles carrying the radiation nanoparticles and the chemotherapeutic drug having a final product of about 120 nm. . The synthesis method according to claim 2, wherein the radiation nanoparticle is uranium 238. The synthetic method according to claim 3, wherein the chemotherapeutic drug is doxorubicin. The method according to claim 1, wherein the radiation nanoparticle is uranium 238, and the chemotherapeutic drug is doxorubicin.