CN109248146B - Oridonin dry suspension for tumor targeting injection and preparation method thereof - Google Patents
Oridonin dry suspension for tumor targeting injection and preparation method thereof Download PDFInfo
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- A61K31/33—Heterocyclic compounds
- A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
- A61K31/35—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom
- A61K31/352—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom condensed with carbocyclic rings, e.g. methantheline
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- A61K47/06—Organic compounds, e.g. natural or synthetic hydrocarbons, polyolefins, mineral oil, petrolatum or ozokerite
- A61K47/22—Heterocyclic compounds, e.g. ascorbic acid, tocopherol or pyrrolidones
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- A61K9/00—Medicinal preparations characterised by special physical form
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- A61K9/0019—Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner
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- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
- A61K9/141—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers
- A61K9/143—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers with inorganic compounds
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- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
- A61K9/141—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers
- A61K9/145—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers with organic compounds
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- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
Abstract
The invention discloses a tumor-targeted oridonin dry suspension for injection and a preparation method thereof. The oridonin dry suspension for tumor targeting injection comprises, by weight, 5-40% of oridonin, 40-60% of 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, 0.1-30% of wetting agent and 1-40% of suspending agent, and is prepared by grinding and mixing the oridonin, the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, the wetting agent and the suspending agent uniformly, sieving, subpackaging in a cillin bottle with the vacuum degree below 10Pa, and sealing. The oridonin dry suspension for tumor targeting injection provided by the invention has the advantages that the tumor targeting property, the anti-tumor effect and the stability of the oridonin are obviously improved by controlling the content of each component.
Description
Technical Field
The invention relates to a tumor-targeted oridonin dry suspension for injection and a preparation method thereof, belonging to the technical field of biomedical engineering.
Background
Oridonin is a natural organic compound of kaurene diterpenes separated from Rabdosia plant of Labiatae, accounts for more than 90% of the effective components of Rabdosia Rubescens (Hemsl.) Hara, is an anti-tumor active component, and is mainly used for treating solid tumors such as primary liver cancer, breast cancer, gastric cancer and esophageal cancer.
Due to good anti-tumor effect and small toxic and side effect, the beta-cyclodextrin inclusion compound, solid lipid nanoparticles, PCL-PEO-PCL drug-loaded nanoparticles, polylactic acid nanoparticles, liposome and other novel dosage forms have been reported. At present, in clinical medication and various main dosage forms of oridonin in a research stage, the oridonin is easy to oxidize, has poor stability, short biological half-life, low targeting property and low bioavailability, and limits the use of the oridonin.
Research results in recent years show that the folate receptor has high expression on certain solid tumors, such as ovarian cancer, endometrial cancer, rectal cancer, breast cancer, lung cancer, liver cancer, neuroendocrine cancer, brain metastasis cancer and the like, and has low expression on normal tissues, and the property provides a chance for using the folate receptor as a drug target.
Disclosure of Invention
The invention aims to provide a tumor-targeted oridonin dry suspension for injection and a preparation method thereof, which improve the tumor targeting property of oridonin on one hand and improve the stability of the oridonin on the other hand.
The technical scheme for realizing the purpose of the invention is as follows:
the oridonin dry suspension for tumor targeting injection comprises the following components in percentage by weight: 5 to 40 percent of oridonin, 40 to 60 percent of 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, 0.1 to 30 percent of wetting agent and 1 to 40 percent of suspending agent.
Preferably, the tumor-targeted oridonin dry suspension for injection comprises the following components in percentage by weight: 5-40% of oridonin, 40-60% of 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, 5-10% of wetting agent and 10-35% of suspending agent.
Further preferably, the rubescensine A accounts for 20-40 wt%, and the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound accounts for 40-50 wt%.
The wetting agent is a conventional wetting agent, and can be poloxamer 188, 237, 338 or 407.
The suspending agent is a conventional suspending agent, and can be methylcellulose, hydroxyethyl cellulose, hydroxypropyl methylcellulose, polyvinylpyrrolidone K15, polyvinylpyrrolidone K30 or polyvinylpyrrolidone K90.
The structural formula of the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound is shown as a formula (I),
the preparation method comprises the following steps: adding 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid into the graphene oxide dispersion liquid, performing ultrasonic treatment and stirring until the mixture is uniformly mixed, adding N, N-dicyclohexylcarbodiimide and N-hydroxysuccinimide, performing stirring reaction at the temperature of 5-50 ℃, centrifuging, filtering, and drying to obtain the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound.
Preferably, in the preparation method of the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, the ultrasonic treatment and stirring time is 0.5-5 h.
Preferably, in the preparation method of the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, the stirring reaction is carried out for 0.5 to 24 hours.
Preferably, in the preparation method of the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, the centrifugation time is 10min to 120min, and the rotation speed is 500rpm to 15000 rpm.
Preferably, in the preparation method of the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, the drying is vacuum drying, the vacuum degree is-0.05 Mpa to-0.1 Mpa, the temperature is 30 ℃ to 100 ℃, and the drying time is 0.5h to 24 h.
The invention also provides a preparation method of the oridonin dry suspension for tumor targeting injection, which comprises the following steps:
the oridonin, the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, the wetting agent and the suspending agent are ground and mixed uniformly according to the proportion, then the mixture is sieved by a 120-200 mesh sieve, the mixture is packaged in a cillin bottle with the vacuum degree lower than 10Pa, and the mixture is sealed to prepare the oridonin dry suspension for tumor targeting injection.
Compared with the prior art, the invention has the following advantages:
according to the tumor-targeted oridonin dry suspension for injection, the content of each component is controlled, so that the tumor targeting property and the anti-tumor effect of the oridonin are obviously improved on one hand, and the stability of the oridonin is obviously improved on the other hand.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is further described in detail by examples below. It should be understood, however, that the description herein of specific embodiments is only intended to illustrate the invention and not to limit the scope of the invention. The invention is described in more detail in the following examples in which poloxamer 188 is the wetting agent and methylcellulose is the suspending agent, it being understood that other wetting agents such as poloxamer 237, 338 or 407, other suspending agents such as hydroxyethylcellulose, hydroxypropylmethylcellulose, polyvinylpyrrolidone K15, polyvinylpyrrolidone K30 or polyvinylpyrrolidone K90 may also practice the invention.
Comparative example preparation of oridonin injection
The preparation of the oridonin injection comprises the following steps: weighing 40mg of oridonin and 40mg of epigallocatechin gallate, and dissolving in 30mL of 0.5% Tween-80 physiological saline solution to obtain the final product.
Example 1
The preparation method of the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound comprises the following steps:
(1) adding graphene oxide (length: 0.5-3 μm; thickness: 0.55-1.2 nm) into distilled water, and performing ultrasonic treatment and stirring (rotation speed: 50rpm/min) at 5 deg.C for 0.5h to obtain dispersion.
(2) And (4) placing the dispersion liquid into a colloid mill for dispersion and grinding to obtain the graphene oxide dispersion liquid with uniform dispersion.
(3) Adding 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid into the graphene oxide dispersion liquid, and carrying out ultrasonic treatment and stirring for 0.5h to obtain a dispersion liquid mixed by the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid and the graphene oxide.
(4) And (4) adding N, N-dicyclohexylcarbodiimide and N-hydroxysuccinimide into the dispersion liquid obtained in the step (3), and stirring and reacting for 0.5h at the temperature of 5 ℃ to obtain a reaction liquid.
(5) And (4) centrifuging the reaction solution obtained in the step (4) at 500rpm for 10min, and after the centrifugation is finished, discarding the supernatant to obtain a precipitate.
(6) Adding distilled water into the precipitate obtained in the step (5), stirring uniformly, filtering with a microporous filter membrane (aperture: 0.45 μm), and discarding the filtrate to obtain a filter cake.
(7) And (4) placing the filter cake obtained in the step (6) in a vacuum drying oven (vacuum degree: -0.05 Mpa; temperature: 30 ℃) for drying for 0.5h to obtain the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound.
2. The oridonin dry suspension for tumor targeting injection comprises the following components in percentage by weight:
5% of oridonin, 60% of 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, 18810% of wetting agent poloxamer and 25% of suspending agent methylcellulose.
3. The preparation method of the oridonin dry suspension for tumor targeting injection comprises the following steps:
(1) weighing the components in the prescription amount in the oridonin dry suspension for tumor targeting injection, grinding and mixing uniformly, and sieving with a 120-mesh sieve for later use.
(2) And (2) subpackaging the material obtained in the step (1) in a penicillin bottle under the vacuum degree of less than 10Pa, and sealing to obtain the tumor-targeted oridonin dry suspension for injection.
Placing the tumor-targeted oridonin dry suspension for injection at 60 ℃ for 6 months, wherein the content of the oridonin in the tumor-targeted oridonin dry suspension for injection is basically unchanged (the deviation range is less than or equal to 2%). The tumor targeting oridonin dry suspension for injection, which is placed for 6 months at the temperature of 60 ℃, is dispersed in sterilized water for injection and is kept stable within 48 hours. Inoculating HePG-2 cells to subcutaneous tissues of Kunming mice, establishing a mouse liver tumor model, respectively administering a tumor-targeted oridonin dry suspension for injection (dispersed in water for sterile injection before use) and an oridonin injection to tail veins of the mice, killing the mice at different time points (0.05h, 0.25h, 0.5h, 1h, 2h, 4h, 8h, 12h, 24h and 48h), taking tumor blocks, and analyzing the content of the oridonin in tumor tissues by HPLC. The result shows that the relative targeting rate of the oridonin dry suspension for tumor targeting injection in tumor tissues is 7.18.
Inoculating HePG-2 cells to subcutaneous tissues of Kunming mice, establishing a mouse liver tumor model, respectively administering a tumor-targeted oridonin dry suspension for injection (dispersed in water for sterilization injection before use) and an oridonin injection to the tail vein of the mice, treating once a day, killing the nude mice after continuously treating for 10 days, and stripping tumors to calculate the tumor inhibition rate of the nude mice bearing human liver cancer. The results show that the administered dose is 1.68X 10－2g·Kg－1·d－1The tumor inhibition rates of the two drugs (calculated by rubescensin A) are respectively 89.3% and 39.6%.
Example 2
The preparation method of the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound comprises the following steps:
(1) adding graphene oxide (length: 0.5-3 μm; thickness: 0.55-1.2 nm) into distilled water, and performing ultrasonic treatment and stirring (rotation speed: 500rpm) at 50 deg.C for 5h to obtain dispersion.
(2) And (3) placing the dispersion liquid obtained in the step (1) into a colloid mill for dispersion and grinding to obtain the graphene oxide dispersion liquid with uniform dispersion.
(3) And (3) adding 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid into the graphene oxide dispersion liquid obtained in the step (2), and carrying out ultrasonic treatment and stirring for 5 hours to obtain a dispersion liquid mixed by the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid and the graphene oxide.
(4) And (4) adding N, N-dicyclohexylcarbodiimide and N-hydroxysuccinimide into the dispersion liquid obtained in the step (3), and stirring and reacting for 24 hours at 50 ℃ to obtain a reaction liquid.
(5) And (4) centrifuging the reaction solution obtained in the step (4) at 15000rpm for 120min, and after the centrifugation is finished, discarding a supernatant to obtain a precipitate.
(6) Adding distilled water into the precipitate obtained in the step (5), stirring uniformly, filtering with a microporous filter membrane (aperture: 0.45 μm), and discarding the filtrate to obtain a filter cake.
(7) And (4) drying the filter cake obtained in the step (6) in a vacuum drying oven (vacuum degree: -0.1 Mpa; temperature: 100 ℃) for 24 hours to obtain the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound.
2. The oridonin dry suspension for tumor targeting injection comprises the following components in percentage by weight:
the preparation comprises, by weight, 10% of rubescensin, 50% of 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, 18810% of wetting agent poloxamer and 30% of suspending agent methylcellulose.
3. The preparation method of the oridonin dry suspension for tumor targeting injection comprises the following steps:
(1) weighing the components in the prescription amount in the oridonin dry suspension for tumor targeting injection, grinding and mixing uniformly, and sieving with a 200-mesh sieve for later use.
(2) And (2) subpackaging the material obtained in the step (1) in a penicillin bottle under the vacuum degree of less than 10Pa, and sealing to obtain the tumor-targeted oridonin dry suspension for injection.
Placing the tumor-targeted oridonin dry suspension for injection at 60 ℃ for 6 months, wherein the content of the oridonin in the tumor-targeted oridonin dry suspension for injection is basically unchanged (the deviation range is less than or equal to 2%). The tumor targeting oridonin dry suspension for injection, which is placed for 6 months at the temperature of 60 ℃, is dispersed in sterilized water for injection and is kept stable within 48 hours. Inoculating HePG-2 cells to subcutaneous tissues of Kunming mice, establishing a mouse liver tumor model, respectively administering a tumor-targeted oridonin dry suspension for injection (dispersed in water for sterile injection before use) and an oridonin injection to tail veins of the mice, killing the mice at different time points (0.05h, 0.25h, 0.5h, 1h, 2h, 4h, 8h, 12h, 24h and 48h), taking tumor blocks, and analyzing the content of the oridonin in tumor tissues by HPLC. The result shows that the relative targeting rate of the oridonin dry suspension for tumor targeting injection in tumor tissues is 7.28.
Inoculating HePG-2 cells to subcutaneous tissues of Kunming mice, establishing a mouse liver tumor model, respectively administering a tumor-targeted oridonin dry suspension for injection (dispersed in water for sterilization injection before use) and an oridonin injection to the tail vein of the mice, treating once a day, killing the nude mice after continuously treating for 10 days, and stripping tumors to calculate the tumor inhibition rate of the nude mice bearing human liver cancer. The results show that the administered dose is 1.68X 10－2g·Kg－1·d－1The tumor inhibition rates of the two drugs are respectively 90.8 percent and 39.6 percent (calculated by oridonin).
Example 3
The preparation method of the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound comprises the following steps:
(1) adding graphene oxide (length: 0.5-3 μm; thickness: 0.55-1.2 nm) into distilled water, and performing ultrasonic treatment and stirring (rotation speed: 300rpm) at 20 deg.C for 2h to obtain dispersion.
(2) And (3) further dispersing and grinding the dispersion liquid obtained in the step (1) in a colloid mill to obtain a uniformly dispersed graphene oxide dispersion liquid.
(3) Adding 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid into the graphene oxide dispersion liquid obtained in the step (2), and carrying out ultrasonic treatment and stirring for 2 hours to obtain a dispersion liquid mixed by the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid and the graphene oxide.
(4) And (4) adding N, N-dicyclohexylcarbodiimide and N-hydroxysuccinimide into the dispersion liquid obtained in the step (3), and stirring and reacting for 10 hours at the temperature of 20 ℃ to obtain a reaction liquid.
(5) And (4) placing the reaction solution obtained in the step (4) into a high-speed centrifuge for centrifugation for 60min at the rotating speed of 2000rpm/min, and after the centrifugation is finished, removing the supernatant to obtain a precipitate.
(6) Adding distilled water into the precipitate obtained in the step (5), stirring uniformly, filtering with a microporous filter membrane (aperture: 0.45 μm), and discarding the filtrate to obtain a filter cake.
(7) And (4) drying the filter cake obtained in the step (6) in a vacuum drying oven (vacuum degree: -0.08 Mpa; temperature: 60 ℃) for 12 hours to obtain the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound.
2. The oridonin dry suspension for tumor targeting injection comprises the following components in percentage by weight:
20% of oridonin, 40% of 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, 1885% of wetting agent poloxamer and 35% of suspending agent methyl cellulose.
3. The preparation method of the oridonin dry suspension for tumor targeting injection comprises the following steps:
(1) weighing the components in the prescription amount in the oridonin dry suspension for tumor targeting injection, grinding and mixing uniformly, and sieving with a 160-mesh sieve for later use.
(2) And (2) subpackaging the material obtained in the step (1) in a penicillin bottle under the vacuum degree of less than 10Pa, and sealing to obtain the tumor-targeted oridonin dry suspension for injection.
Placing the tumor-targeted oridonin dry suspension for injection at 60 ℃ for 6 months, wherein the content of the oridonin in the tumor-targeted oridonin dry suspension for injection is basically unchanged (the deviation range is less than or equal to 2%). The tumor targeting oridonin dry suspension for injection, which is placed for 6 months at the temperature of 60 ℃, is dispersed in sterilized water for injection and is kept stable within 48 hours. Inoculating HePG-2 cells to subcutaneous tissues of Kunming mice, establishing a mouse liver tumor model, respectively administering a tumor-targeted oridonin dry suspension for injection (dispersed in water for sterile injection before use) and an oridonin injection to tail veins of the mice, killing the mice at different time points (0.05h, 0.25h, 0.5h, 1h, 2h, 4h, 8h, 12h, 24h and 48h), taking tumor blocks, and analyzing the content of the oridonin in tumor tissues by HPLC. The result shows that the relative targeting rate of the oridonin dry suspension for tumor targeting injection in tumor tissues is 7.82.
Inoculating HePG-2 cells into subcutaneous tissue of Kunming mouse, establishing mouse liver tumor model, and respectively mixing tumor-targeted oridonin dry suspension (dispersed in sterilized water for injection before use) and oridonin dry suspensionThe Lingcao A injection is administrated through tail vein of mouse, and is used for treating once a day, and the nude mouse is killed after continuous treatment for 10 days, and the tumor inhibition rate of the nude mouse with human liver cancer is calculated by stripping tumor. The results show that the administered dose is 1.68X 10－2g·Kg－1·d－1The tumor inhibition rates of the two drugs (calculated by rubescensin A) are 91.6 percent and 39.6 percent respectively.
Example 4
The preparation method of the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound comprises the following steps:
(1) adding graphene oxide (length: 0.5-3 μm; thickness: 0.55-1.2 nm) into distilled water, and performing ultrasonic treatment at 25 deg.C and stirring (rotation speed: 500rpm) for 5 hr to obtain dispersion.
(2) And (3) further dispersing and grinding the dispersion liquid obtained in the step (1) in a colloid mill to obtain a uniformly dispersed graphene oxide dispersion liquid.
(3) Adding 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid into the graphene oxide dispersion liquid obtained in the step (2), and carrying out ultrasonic treatment and stirring for 2 hours to obtain a dispersion liquid mixed by the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid and the graphene oxide.
(4) And (4) adding N, N-dicyclohexylcarbodiimide and N-hydroxysuccinimide into the dispersion liquid obtained in the step (3), and stirring and reacting for 5 hours at the temperature of 30 ℃ to obtain a reaction liquid.
(5) And (4) placing the reaction solution obtained in the step (4) in a high-speed centrifuge for centrifugation for 30min at the rotating speed of 2000rpm/min, and after the centrifugation is finished, discarding the supernatant to obtain a precipitate.
(6) Adding distilled water into the precipitate obtained in the step (5), stirring uniformly, filtering with a microporous filter membrane (aperture: 0.45 μm), and discarding the filtrate to obtain a filter cake.
(7) And (4) drying the filter cake obtained in the step (6) in a vacuum drying oven (vacuum degree: -0.08 Mpa; temperature: 50 ℃) for 10 hours to obtain the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound.
2. The oridonin dry suspension for tumor targeting injection comprises the following components in percentage by weight:
40% of oridonin, 40% of 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, 18810% of wetting agent poloxamer and 10% of suspending agent methyl cellulose.
3. The method for preparing the oridonin dry suspension for tumor targeted injection comprises the following steps:
(1) weighing the components in the prescription amount in the oridonin dry suspension for tumor targeting injection, grinding and mixing uniformly, and sieving with a 160-mesh sieve for later use.
(2) And (2) subpackaging the material obtained in the step (1) in a penicillin bottle under the vacuum degree of less than 10Pa, and sealing to obtain the tumor-targeted oridonin dry suspension for injection.
Placing the tumor-targeted oridonin dry suspension for injection at 60 ℃ for 6 months, wherein the content of the oridonin in the tumor-targeted oridonin dry suspension for injection is basically unchanged (the deviation range is less than or equal to 2%). The tumor targeting oridonin dry suspension for injection, which is placed for 6 months at the temperature of 60 ℃, is dispersed in sterilized water for injection and is kept stable within 48 hours. Inoculating HePG-2 cells to subcutaneous tissues of Kunming mice, establishing a mouse liver tumor model, respectively administering a tumor-targeted oridonin dry suspension for injection (dispersed in water for sterile injection before use) and an oridonin injection to tail veins of the mice, killing the mice at different time points (0.05h, 0.25h, 0.5h, 1h, 2h, 4h, 8h, 12h, 24h and 48h), taking tumor blocks, and analyzing the content of the oridonin in tumor tissues by HPLC. The result shows that the relative targeting rate of the oridonin dry suspension for tumor targeting injection in tumor tissues is 8.31.
Inoculating HePG-2 cells to subcutaneous tissues of Kunming mice, establishing a mouse liver tumor model, respectively administering a tumor-targeted oridonin dry suspension for injection (dispersed in water for sterilization injection before use) and an oridonin injection to the tail vein of the mice, treating once a day, killing the nude mice after continuously treating for 10 days, and stripping tumors to calculate the tumor inhibition rate of the nude mice bearing human liver cancer. The results show that the administered dose is 1.68X 10－2g·Kg－1·d－1The tumor inhibition rates of the two drugs (calculated by rubescensin A) are respectively 92.5% and 39.6%.
Comparative example 1
The comparative example is basically the same as the example 4, except that the oridonin dry suspension for tumor targeting injection comprises the following components in percentage by weight:
3% of oridonin, 30% of 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, 18817% of wetting agent poloxamer and 50% of suspending agent methyl cellulose.
Placing the tumor-targeted oridonin dry suspension for injection at 60 ℃ for 6 months, wherein the content of the oridonin in the tumor-targeted oridonin dry suspension for injection is basically unchanged (the deviation range is less than or equal to 2%). The tumor targeting oridonin dry suspension for injection, which is placed for 6 months at the temperature of 60 ℃, is dispersed in sterilized water for injection and is kept stable within 48 hours. Inoculating HePG-2 cells to subcutaneous tissues of Kunming mice, establishing a mouse liver tumor model, respectively administering a tumor-targeted oridonin dry suspension for injection (dispersed in water for sterile injection before use) and an oridonin injection to tail veins of the mice, killing the mice at different time points (0.05h, 0.25h, 0.5h, 1h, 2h, 4h, 8h, 12h, 24h and 48h), taking tumor blocks, and analyzing the content of the oridonin in tumor tissues by HPLC. The result shows that the relative targeting rate of the oridonin dry suspension for tumor targeting injection in tumor tissues is 3.56.
Inoculating HePG-2 cells to subcutaneous tissues of Kunming mice, establishing a mouse liver tumor model, respectively administering a tumor-targeted oridonin dry suspension for injection (dispersed in water for sterilization injection before use) and an oridonin injection to the tail vein of the mice, treating once a day, killing the nude mice after continuously treating for 10 days, and stripping tumors to calculate the tumor inhibition rate of the nude mice bearing human liver cancer. The results show that the administered dose is 1.68X 10－2g·Kg－1·d－1The tumor inhibition rates of the two drugs (calculated by rubescensin A) are respectively 60.1% and 39.6%.
Comparative example 2
The comparative example is basically the same as the example 4, except that the oridonin dry suspension for tumor targeting injection comprises the following components in percentage by weight:
50% of oridonin, 30% of 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, 18810% of wetting agent poloxamer and 10% of suspending agent methyl cellulose.
Placing the tumor-targeted oridonin dry suspension for injection at 60 ℃ for 6 months, wherein the content of the oridonin in the tumor-targeted oridonin dry suspension for injection is basically unchanged (the deviation range is less than or equal to 2%). The tumor targeting oridonin dry suspension for injection, which is placed for 6 months at the temperature of 60 ℃, is dispersed in sterilized water for injection and is kept stable within 48 hours. Inoculating HePG-2 cells to subcutaneous tissues of Kunming mice, establishing a mouse liver tumor model, respectively administering a tumor-targeted oridonin dry suspension for injection (dispersed in water for sterile injection before use) and an oridonin injection to tail veins of the mice, killing the mice at different time points (0.05h, 0.25h, 0.5h, 1h, 2h, 4h, 8h, 12h, 24h and 48h), taking tumor blocks, and analyzing the content of the oridonin in tumor tissues by HPLC. The result shows that the relative targeting rate of the oridonin dry suspension for tumor targeting injection in tumor tissues is 4.21.
Inoculating HePG-2 cells to subcutaneous tissues of Kunming mice, establishing a mouse liver tumor model, respectively administering a tumor-targeted oridonin dry suspension for injection (dispersed in water for sterilization injection before use) and an oridonin injection to the tail vein of the mice, treating once a day, killing the nude mice after continuously treating for 10 days, and stripping tumors to calculate the tumor inhibition rate of the nude mice bearing human liver cancer. The results show that the administered dose is 1.68X 10－2g·Kg－1·d－1The tumor inhibition rates of the two drugs (calculated by oridonin) are respectively 64.3 percent and 39.6 percent.
Claims (9)
1. The oridonin dry suspension for tumor targeting injection is characterized by comprising the following components in percentage by weight: 5-40% of oridonin, 40-60% of 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, 0.1-30% of wetting agent and 1-40% of suspending agent, wherein the wetting agent is selected from poloxamer 188, 237, 338 or 407, and the suspending agent is selected from methyl cellulose, hydroxyethyl cellulose, hydroxypropyl methyl cellulose, polyvinylpyrrolidone K15, polyvinylpyrrolidone K30 or polyvinylpyrrolidone K90.
2. The tumor-targeted oridonin dry suspension for injection according to claim 1, which comprises the following components in percentage by weight: 5-40% of oridonin, 40-60% of 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, 5-10% of wetting agent and 10-35% of suspending agent.
3. The tumor-targeted oridonin dry suspension for injection according to claim 2, wherein the oridonin is 20-40 wt% and the 2-amino-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound is 40-50 wt%.
4. The tumor-targeted oridonin dry suspension for injection according to any one of claims 1 to 3, wherein the structural formula of the 2-amino-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound is shown as the formula (I),
5. the tumor-targeted oridonin dry suspension for injection according to claim 4, wherein the preparation method of the 2-amino-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound comprises the following steps: adding 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid into the graphene oxide dispersion liquid, performing ultrasonic treatment and stirring until the mixture is uniformly mixed, adding N, N-dicyclohexylcarbodiimide and N-hydroxysuccinimide, performing stirring reaction at the temperature of 5-50 ℃, centrifuging, filtering, and drying to obtain the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound.
6. The oridonin dry suspension for tumor targeting injection according to claim 5, wherein the ultrasound and stirring time is 0.5-5 h, and the stirring reaction is 0.5-24 h.
7. The tumor-targeted oridonin dry suspension for injection according to claim 5, wherein the centrifugation time is 10min to 120min, and the rotation speed is 500rpm to 15000 rpm.
8. The tumor-targeted oridonin dry suspension for injection according to claim 5, wherein the drying is vacuum drying, the vacuum degree is-0.05 Mpa to-0.1 Mpa, the temperature is 30 ℃ to 100 ℃, and the drying time is 0.5h to 24 h.
9. The tumor-targeted oridonin dry suspension for injection according to any one of claims 1 to 3, wherein the preparation method comprises the following steps: the oridonin, the 2-ammonia-4-hydroxy-7, 8-dihydropteroic acid coupled graphene compound, the wetting agent and the suspending agent are ground and mixed uniformly according to the proportion, then the mixture is sieved by a 120-200 mesh sieve, the mixture is packaged in a cillin bottle with the vacuum degree lower than 10Pa, and the mixture is sealed to prepare the oridonin dry suspension for tumor targeting injection.
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