CN108619511B - Preparation method and application of metal-organic framework drug delivery system based on cytarabine small-molecule prodrug - Google Patents
Preparation method and application of metal-organic framework drug delivery system based on cytarabine small-molecule prodrug Download PDFInfo
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Abstract
The invention discloses a preparation method and application of a metal organic framework drug-loading system of a cytarabine prodrug. Taking the synthesized Ara-IR820@ ZIF-8 as an example, the in vivo antitumor effect results of mice show that the HA/Ara-IR820@ ZIF-8 nano aqueous solution HAs good antitumor effect and lower systemic toxicity. The intravenous injection preparation has high drug loading, simple and economic preparation process, batch production, low toxicity and good safety, and provides wide application prospect for realizing high drug loading of the cytarabine in a metal organic framework and treating solid tumors by intravenous injection administration.
Description
Technical Field
The invention relates to the technical field of medicines, in particular to design and synthesis of a novel cytarabine small molecule prodrug, and preparation and evaluation of a metal organic framework drug delivery system based on the drug small molecule prodrug.
Background
With the rising morbidity and mortality, cancer has become the leading factor threatening public health, and the treatment of cancer is still slow. Chemotherapy is a means of inhibiting tumor cell proliferation and promoting tumor cell apoptosis by using chemical drugs, and is the most widely used treatment means at present. Cytarabine, a typical pyrimidine antimetabolite, is one of the most common chemotherapeutic drugs in clinic and is widely used for treating various acute and chronic leukemias. However, the molecular polarity is very large, the fat solubility is low, and the membrane permeability is poor; furthermore, NH on the pyrimidine ring2Is easy to deaminate and inactivate by deaminase to form inactive arabinouracil, so that it has low oral bioavailability and very short plasma half-life after intravenous injection. In order to achieve effective blood concentration, a continuous intravenous drip mode is mainly adopted clinically; moreover, the clinical application of cytarabine is greatly limited due to the poor effect of cytarabine on the treatment of solid tumors reported by the literature.
For improving the fat solubility and the biological membrane permeability of the cytarabine and protecting NH on a pyrimidine ring of a medicament2Without metabolic inactivation by deaminase, hydrophobic chains of different chain length (e.g. C)6-C20Aliphatic chain, etc.) covalent modification at cytarabine NH2The prodrug small molecule is obtained. At present, the satisfactory effect of the monotherapy on the tumor cannot be achieved, and clinically, in order to obtain a better treatment effect and improve the tumor cure rate, a combined means of two or more treatment modes is adopted for treatment. By mixing drug molecules (such as camptothecin, doxorubicin hydrochloride, paclitaxel, etc.), photothermal therapy photosensitizer (such as indocyanine green, neoindocyanine green, etc.)IR780, etc.), photodynamic therapy photosensitizer (hypocrellin, chlorin, porphyrin derivative, etc.) covalently modified in cytarabine NH2The multifunctional prodrug micromolecules can be prepared, wherein chemotherapy, photothermal therapy and photodynamic therapy have respective action mechanisms and cytotoxicity, usually generate synergistic effect, can weaken side effect generated in the treatment process and obviously enhance the anti-tumor curative effect. In the current research on small molecule prodrugs, limited drug grafts can realize drug molecule self-assembly, or prepare free prodrug aqueous solutions, or prepare precipitates with larger particle sizes. The water solution of the free prodrug has the problem of non-targeted distribution, so that a large dose is required to meet the treatment effect of the drug on tumor parts, and the systemic toxicity is increased. Larger, irregular particle sizes can limit the route of administration of the formulation: the preparation process of the oral administration dosage form is complicated, and the production cost is high; intratumoral injection is not favorable for practical clinical operation.
The main problems that pharmaceutical preparations are now required to solve are to enhance the tumor-inhibiting effect of the drug and to reduce its toxicity: the medicine is delivered to the tumor part to increase the curative effect of the medicine, avoid the whole body distribution of the medicine, reduce the damage of the medicine to normal tissues and enhance the killing effect of the medicine to tumor cells. Therefore, the development of a drug nano delivery system with high drug loading, safety, effectiveness and targeting distribution function has important research significance and clinical application value. The existing drug carrier materials are mainly divided into inorganic materials and organic materials: the inorganic material is used as a carrier, the material has certain toxicity, and the biodegradability is poor due to the over-stable structure of the material, so that the material cannot be effectively degraded by organisms and discharged out of the body. Although the organic material has good biocompatibility and can be subjected to post-modification on the surface of the material, the drug-loading rate of the material is limited; moreover, due to its weak thermal and chemical stability, the drug may have a burst release phenomenon upon release. Therefore, the development of new effective drug carriers is of great importance. Metal Organic Frameworks (MOFs) (e.g., ZIF series, MIL series, UiO series, IRMOF series, etc.) are coordination complex crystal materials with periodic network structures, which are self-assembled by using Metal ions as hybridization centers and oxygen-containing or nitrogen-containing Organic compounds as ligands. The MOF has the structural characteristics of pore channel adjustability, ultrahigh specific surface area, framework composition diversity, composition designability, excellent surface post-modification and the like, shows huge application prospect in the field of drug delivery, and is expected to become an excellent drug carrier. However, because of the stability of the MOF material structure and the specificity of different drug molecular structures, the currently known drug types that can be loaded by the MOF material are very limited, and the anticancer drug cytarabine of the present invention is easy to leak from the MOF pore size due to the small molecular structure, and is difficult to implement loading, therefore, the development of a metal organic framework drug-loaded delivery system of a high-loading small-molecule drug is urgently needed.
Disclosure of Invention
In combination with the above problems, the present invention was first developed for MOF formulations carrying small molecule prodrugs, in particular cytarabine small molecule prodrugs. According to the invention, an anticancer drug cytarabine is combined with a hydrophobic chain, a drug molecule, a photo-thermal treatment photosensitizer and a photodynamic treatment photosensitizer, and carboxyl groups, sulfonic acid groups and ester groups in the hydrophobic chain, the drug molecule, the photo-thermal treatment photosensitizer and the photodynamic treatment photosensitizer are coordinated with metal in a metal organic framework, so that the cytarabine is dragged into an MOF cavity, the co-loading of two compounds with equal molar ratio is realized, and the characteristic of higher drug loading is further realized. For illustration, the invention selects and utilizes near-infrared dye new indocyanine green (IR820), and the coordination of the sulfonic acid group of the indocyanine green and the metal in the metal organic framework zeolite imidazolate framework material (ZIF-8) to realize the effective entrapment of the MOF on the small-molecule anticancer drug cytarabine.
In order to achieve the above purpose, the present invention specifically relates to the following technical solutions:
the invention synthesizes a novel cytarabine and novel indocyanine green small molecule prodrug (Ara-IR820), the IR820 and chemotherapy drug cytarabine are reacted through amide to obtain the prodrug Ara-IR820, the Ara-IR820 not only improves the curative effect of cytarabine on solid tumors, but also improves the stability of cytarabine, in addition, the synthesized Ara-IR820 prodrug has larger structural polarity, can be dispersed in water to prepare high-concentration aqueous solution, and lays a foundation for the subsequent realization of a drug delivery system with high drug loading capacity.
In order to achieve the purpose, the invention specifically relates to the following technical scheme:
the invention provides a novel cytarabine and novel indocyanine green small molecule prodrug (Ara-IR820), which has the following structural formula:
the invention also provides a preparation method of the cytarabine and novel indocyanine green small molecule prodrug (Ara-IR820), which comprises the following steps:
reacting the new indocyanine green with 6-aminocaproic acid, modifying carboxyl on the new indocyanine green, separating and purifying the product, and then carrying out amide reaction with cytarabine to obtain the product.
Preferably, the reaction process of the novel indocyanine green and the cytarabine is as follows: reacting the new indocyanine green with 6-aminocaproic acid under the condition of adding triethylamine to generate a first-step product; and (3) separating and purifying the product, and reacting the product with cytarabine under the catalysis of triethylamine and ethyl chloroformate to generate the cytarabine and the new indocyanine green micromolecule prodrug (Ara-IR 820).
The specific reaction steps comprise: (1) dissolving a certain amount of new indocyanine green in anhydrous N, N-Dimethylformamide (DMF), adding triethylamine, dissolving a certain amount of 6-aminocaproic acid in anhydrous DMF, adding the obtained solution into the reaction solution, and reacting under the conditions of inert gas protection and 85 ℃ oil bath to obtain IR 820-COOH.
The novel indocyanine green: 6-aminocaproic acid: the molar ratio of triethylamine is 1:5: 5.
In a preferred embodiment, a step (2) of IR820-COOH purification is included thereafter.
Specifically, the purification step (2) is as follows: carrying out reduced pressure rotary evaporation to remove the anhydrous DMF (dimethyl formamide) serving as a reaction solvent, and carrying out vacuum drying to obtain a crude product; the crude product was dissolved in methanol and purified by silica gel column chromatography eluting with ethyl acetate and methanol in a gradient (10:1-3:1) to give pure IR820-COOH as a blue solid.
(3) Dissolving a certain amount of IR820-COOH in anhydrous DMF, adding triethylamine and ethyl chloroformate, and reacting under the conditions of inert gas protection and ice bath;
(4) and (3) dissolving a certain amount of cytarabine in anhydrous DMF, and adding the dissolved cytarabine and the reaction solution in the step (3) to prepare the small molecule prodrug of the cytarabine and the new indocyanine green (Ara-IR 820).
The IR 820-COOH: cytarabine: triethylamine: the molar ratio of ethyl chloroformate was 1:3:3: 3.
In a preferred embodiment, the method further comprises a step (5) of Ara-IR820 purification.
Specifically, the purification step (5) is as follows: carrying out reduced pressure rotary evaporation to remove the anhydrous DMF (dimethyl formamide) serving as a reaction solvent, and carrying out vacuum drying to obtain a crude product; the crude product was dissolved in methanol and purified by silica gel column chromatography eluting with a gradient of ethyl acetate and methanol (4:1-2.5:1) to give Ara-IR820 as a pure blue solid.
The invention also provides the application of the cytarabine and the novel indocyanine green small molecule prodrug (Ara-IR820) in resisting tumors, which can be used for treating or relieving the cancers of certain tissues or organs, wherein the cancers include but are not limited to leukemia, solid tumors and the like.
The invention also synthesizes a metal organic framework drug delivery system with an active targeting function based on the cytarabine small molecule prodrug.
In order to achieve the purpose, the invention specifically relates to the following technical scheme:
the invention discloses a preparation method of a metal organic framework drug delivery system with an active targeting function based on a cytarabine small molecule prodrug, which comprises the following steps: dissolving the synthesized Ara prodrug in water, dropwise adding the synthesized Ara prodrug into an aqueous solution of metal salt ions under the stirring condition, and dropwise adding the mixed solution into an aqueous solution of a ligand under the stirring condition; centrifuging the formed suspension, washing with water, and drying to obtain a solid, namely the metal organic framework drug delivery system encapsulating the cytarabine small molecule prodrug;
and uniformly dispersing the metal organic framework encapsulating the cytarabine small-molecule prodrug in a ligand aqueous solution with a tumor active targeting function, stirring at room temperature, centrifuging the formed suspension, washing with water, and drying to obtain a solid, namely the metal organic framework drug delivery system with the active targeting function based on the cytarabine small-molecule prodrug.
The characteristic that sulfonic acid groups, carboxylic acid groups and ester groups in a part of compounds can be coordinated with metal in a metal organic framework so as to realize high drug loading is utilized, related compounds and anticancer drug cytarabine are covalently combined through amide bonds so as to drag the cytarabine into an MOF cavity, and two molecules are loaded together in an equal molar ratio, so that the construction of a metal organic framework drug delivery system with a targeting function of encapsulating small-molecule prodrugs is realized.
Specifically, taking cytarabine and a novel indocyanine green small molecule prodrug (Ara-IR820) and a ZIF-8 metal organic framework as examples, the preparation method comprises the following steps:
dissolving Ara-IR820 prodrug in water, and adding dropwise zinc nitrate hexahydrate (Zn (NO)3)2·6H2O) adding the mixed solution into the aqueous solution of 2-methylimidazole dropwise under the condition of stirring; centrifuging, washing and drying the formed suspension to obtain a blue solid, namely a metal organic framework drug delivery system Ara-IR820@ ZIF-8 based on cytarabine and the new indocyanine green small molecule prodrug;
Ara-IR820@ ZIF-8 is uniformly dispersed in an aqueous solution of Hyaluronic Acid (HA), stirred at room temperature, and the formed suspension is centrifuged, washed and dried to obtain a blue solid, namely a metal organic framework drug delivery system HA/Ara-IR820@ ZIF-8 with an active targeting function based on cytarabine and a new indocyanine green small molecule prodrug (Ara-IR 820).
Experiments show that the concentration of the formed stable suspension of the nano aggregate intravenous injection preparation can reach more than 10 mg/mL.
The metal organic framework drug delivery system HA/Ara-IR820@ ZIF-8 with the active targeting function obtained by the preparation method is also in the protection scope of the invention.
The invention also discloses application of the metal organic framework drug delivery system with the active targeting function based on the cytarabine small molecule prodrug, which can be used for treating or relieving cancers of a certain tissue or organ, wherein the cancers include but are not limited to leukemia, solid tumors and the like.
Preferably, the application is that a metal organic framework drug delivery system with an active targeting function based on the cytarabine small molecule prodrug is ultrasonically and uniformly dispersed in water to prepare a target intravenous injection preparation. The intravenous injection preparation is obtained by a simple, rapid and green method, has good stability, is easy to store, and has excellent in-vivo anti-tumor effect.
The invention achieves the following beneficial effects:
(1) the invention synthesizes cytarabine and new indocyanine green micromolecule prodrug Ara-IR820 for the first time, the prodrug molecule not only overcomes the defects that Ara is easy to be metabolized and inactivated in vivo and the like, but also improves the toxicity of the drug to solid tumor, improves the Ara stability and expands the anti-tumor application range of Ara.
(2) According to the invention, Ara-IR820 is loaded into the pore cavity of the metal organic framework ZIF-8 to prepare the nanoparticle by a simple, convenient and rapid method, active targeted HA modification is realized, the method is economical and environment-friendly, and can be used for mass production, and the solid is easy to store and transport, so that the possibility is provided for industrial production.
(3) The invention firstly generates the prodrug through the grafting reaction with the compound, encapsulates the cytarabine into the pore cavity of the metal organic framework ZIF-8 and prepares the metal organic framework drug delivery system with the active targeting function, thereby laying a foundation for researching the metal organic framework drug delivery system based on the Ara micromolecule prodrug.
(4) The HA/Ara-IR820@ ZIF-8 nanoparticle intravenous injection preparation HAs low toxicity, good safety and high solid tumor inhibition rate, and provides possibility for realizing cytarabine intravenous injection administration for treating solid tumors.
Drawings
FIG. 1 NMR spectra of IR820-COOH and Ara-IR 820;
FIG. 2HA/Ara-IR820@ ZIF-8 nanoparticle Transmission Electron Microscope (TEM) image;
FIG. 3 shows the results of tumor volume, mouse body weight change curve and tumor inhibition rate in the pharmacodynamic experiment of solid tumor treatment by HA/Ara-IR820@ ZIF-8 nanoparticle intravenous injection.
Detailed Description
The present invention is further illustrated by reference to specific examples, which are intended to be illustrative only and not limiting. If the experimental conditions specified in the examples are not indicated, they are generally according to the conventional conditions, or according to the conditions recommended by the reagents company; reagents, consumables and the like used in the following examples are commercially available unless otherwise specified.
EXAMPLE 1Ara-IR820 prodrug molecular Synthesis
Accurately weighing a certain amount of IR820 by an analytical balance, dissolving the IR820 in anhydrous DMF, placing the solution in a round bottom flask, accurately weighing a certain amount of 6-aminocaproic acid by the analytical balance, dissolving the 6-aminocaproic acid in the anhydrous DMF, adding the DMF solution of the IR820 under the stirring condition, adding triethylamine under the stirring condition, reacting for 4 hours under the protection of nitrogen and oil bath at 85 ℃. Wherein the molar weight is IR 820: triethylamine: 6-aminocaproic acid 1:5:5, the progress of the reaction was monitored using thin layer plates. After the reaction is finished, the anhydrous DMF is removed by reduced pressure rotary evaporation, and the crude product is obtained by vacuum drying overnight. The crude product was dissolved in methanol, stirred over a small amount of column chromatography silica gel, purified by column chromatography on silica gel, and eluted with ethyl acetate and methanol in a gradient (10:1-3:1) to give IR820-COOH as a pure blue solid in 48.8% yield.
Accurately weighing a certain amount of IR820-COOH on an analytical balance, dissolving in anhydrous DMF, placing in a round bottom flask, adding triethylamine and ethyl chloroformate under stirring, and reacting for 20min under the conditions of nitrogen protection and ice bath. Weighing a certain amount of cytarabine, dissolving the cytarabine in anhydrous DMF, and slowly dripping the cytarabine into the reaction solution under the stirring condition, wherein the molar weight is IR 820-COOH: triethylamine: ethyl chloroformate: cytarabine ═ 1:3:3:3, the reaction returned to room temperature and continued for 72h under nitrogen protection, and the progress of the reaction was monitored using thin layer plates. After the reaction is finished, the anhydrous DMF is removed by reduced pressure rotary evaporation, and the crude product is obtained by vacuum drying overnight. The crude product was dissolved in methanol, stirred over a small portion of column chromatography silica gel, purified by column chromatography on silica gel, and eluted with a gradient of ethyl acetate and methanol (4:1-2.5:1) to give Ara-IR820 as a pure blue solid in 61.2% yield.
Example 2 nuclear magnetic resonance hydrogen spectrum (1H-NMR) identification of IR820-COOH and Ara-IR820 prodrug chemistryStructure of the product
Approximately 5mg of the IR820-COOH and Ara-IR820 prodrug are respectively weighed, deuterated dimethyl sulfoxide (DMSO-d6) is dissolved and placed in a nuclear magnetic tube, a nuclear magnetic resonance hydrogen spectrum is measured by adopting a 400MHz nuclear magnetic resonance hydrogen spectrum, tetramethylsilane is used as an internal standard, and the chemical shift value (ppm) of the compound is recorded. As shown in FIG. 1, the nuclear magnetic results confirmed that the molar ratio of cytarabine to IR820 in the newly synthesized molecule was close to 1:1, and that the synthesis of the amide bond was confirmed by the appearance of the characteristic peak of H on the amide bond. By passing1The H-NMR spectrum confirmed the successful synthesis of the Ara-IR820 prodrug.
Example 3 preparation of a Metal-organic framework drug delivery System with active targeting function based on Cytarabine and New indocyanine Green Small molecule prodrug (Ara-IR820) as intravenous formulation
Precisely weighing about 50mg of Ara-IR820 prodrug molecule, dissolving in 2mL of water, and adding 0.5mL of Zn (NO) dropwise under stirring3)2·6H2And (2) adding the suspension into an O aqueous solution, dropwise adding the suspension into 5mL of a 2-methylimidazole aqueous solution under the condition of stirring, carrying the MOF nanoparticles to form spontaneously, centrifuging the suspension, washing with water, removing a supernatant, and drying in vacuum to obtain a blue solid Ara-IR820@ ZIF-8.
Uniformly dispersing Ara-IR820@ ZIF-8 in a 3mg/mL Hyaluronic Acid (HA) water solution, and stirring at room temperature for 48 hours; and centrifuging, washing and drying the formed suspension to obtain a blue solid, and uniformly dispersing the blue solid in water by ultrasonic waves to obtain the target intravenous injection preparation.
Experiments show that the concentration of the formed stable suspension of the nano aggregate intravenous injection preparation can reach more than 10 mg/mL.
The sulfonic acid group in the near-infrared dye IR820 can coordinate with metal in a metal organic framework so as to realize the characteristic of high drug loading, the IR820 and an anticancer drug cytarabine are covalently combined through an amido bond so as to drag the cytarabine into an MOF cavity, the two drugs are loaded together in an equal molar ratio, and the construction of a metal organic framework drug delivery system with a targeting function of a small-molecule prodrug is realized.
Example 4HA/Ara-IR820@ ZIF-8 Nanoaggregate morphology Observation
A small amount of dried nanoaggregate was taken and suspended in a 1.5mL Ep tube with 200. mu.L of distilled water. And (3) sucking a drop of suspension liquid on a carbon film copper net, sucking redundant liquid by using filter paper, drying at room temperature, and then placing under a transmission electron microscope to observe the form of the HA/Ara-IR820@ ZIF-8 nano-aggregate. The electron micrograph is shown in figure 2, and the result shows that HA/Ara-IR820@ ZIF-8 can be dispersed in water into a relatively round nanoparticle structure with the diameter of about 100-200nm, the shape is uniform, the dispersibility is good, the nanoparticle size is suitable for intravenous injection administration, and the requirement of Enhanced Permeation and Retention (EPR) effect on the particle size is met. The preparation process of the nanoparticles does not need to add organic solvent and surfactant, does not need to heat, and has simpler and quicker reaction.
Example 5HA/Ara-IR820@ ZIF-8 in vivo pharmacodynamic study of intravenous injection treatment of solid tumors
1. Construction of Breast cancer mouse model
25 healthy BALB/C mice of 6-8 weeks old were taken, placed in a suitable environment, and raised for 1 week. After the mice acclimatized, 0.1mL of 2X 10 density vaccine was inoculated into the right anterior axilla7A/mL suspension of mouse breast cancer (4T1) cells was inoculated and then continued to be fed. Mice were examined daily for growth of tumors in the axilla of the right forelimb. When the tumor volume (V is 1/2 × length × width)2) Up to about 150mm3Thereafter, tumor suppression experiments were started.
2. Tumor inhibition experiment in vivo
In the experiment, a female BALB/C mouse with 4T1 mouse breast cancer cells is taken as an animal model, and the in vivo anti-tumor effect of HA/Ara-IR820@ ZIF-8 is researched. Mice were randomly divided into 5 groups of 5 mice each. Respectively injecting a normal saline solution, a cytarabine aqueous solution, an IR820 aqueous solution, an Ara-IR820@ ZIF-8 nanoparticle aqueous solution and an HA/Ara-IR820@ ZIF-8 nanoparticle aqueous solution into a tail vein. Wherein the dosage of the IR820 in the group of IR820 aqueous solution, Ara-IR820@ ZIF-8 nanoparticle aqueous solution and HA/Ara-IR820@ ZIF-8 nanoparticle aqueous solution is 2mg/kg, the dosage of the group of cytarabine aqueous solution is consistent with that of the preparation group, the drug is taken once every two days, and after the group of IR820 aqueous solution, Ara-IR820@ ZIF-8 nanoparticle aqueous solution and HA/Ara-IR820@ ZIF-8 nanoparticle aqueous solution is taken, a mouse receives near-infrared laser irradiation5 minutes, power 1W/cm2The administration time points were recorded as days 1, 4, 7, 10, 13, and 16, respectively. Body weight and tumor volume of mice were measured every two days. Mice can eat and drink water freely during administration.
In order to evaluate the antitumor efficacy of HA/Ara-IR820@ ZIF-8 in vivo, tumor-bearing mice were used as animal models to compare the changes in tumor volume and body weight of mice in the normal saline group (as a control), cytarabine aqueous solution group, IR820 aqueous solution group, Ara-IR820@ ZIF-8 nanoparticle aqueous solution group and HA/Ara-IR820@ ZIF-8 nanoparticle aqueous solution group. Tumor volume and relative body weight were calculated by measuring tumor length, tumor length and mouse body weight (real-time measurement of body weight W/initial body weight W)0) The results are shown in FIG. 3.
The tumor volume of the raw material medicine and the preparation group is obviously smaller than that of the normal saline group, which shows that the raw material medicine and the preparation both have obvious inhibition effect on the growth of tumors; moreover, the results show that the tumor inhibition rate of the HA/Ara-IR820@ ZIF-8 nanoparticle aqueous solution group on the mammary tumor of a mouse after tail vein injection is obviously higher than that of a non-targeted Ara-IR820@ ZIF-8 nanoparticle aqueous solution group, a cytarabine bulk drug group, an IR820 bulk drug group and a normal saline group, and the anti-tumor effect is optimal.
The body weight change of the mice can be used as an index to measure the systemic toxicity of the drug to the animals. As can be seen from fig. 3, the relative body weights of the saline group and the formulation group showed a slow and continuous increase from the day of administration. The relative body weights of the two raw material medicines are always in a remarkable reduction state. The weight of the mice in the raw material medicine solution group is obviously reduced, which shows that the mice can generate serious systemic toxicity; the body weight change of the preparation group is not much different from that of the normal saline group, which shows that the systemic toxicity of the medicine can be effectively reduced after the medicine is prepared into a prodrug, is entrapped and delivered to tumor tissues in a targeted manner.
The invention synthesizes cytarabine and new indocyanine green micromolecule prodrug Ara-IR820 for the first time, the prodrug molecule not only overcomes the defects that Ara is easy to be metabolized and inactivated in vivo and the like, but also improves the toxicity of the drug to solid tumors, and expands the anti-tumor application range of Ara. Ara-IR820 is loaded into the pore cavity of a metal organic framework ZIF-8 to prepare nanoparticles by a simple and rapid method, active targeted HA modification is realized, the method is economical and environment-friendly, and can be used for mass production, and the solid is easy to store and transport, so that the possibility is provided for industrial production. The HA/Ara-IR820@ ZIF-8 nanoparticle intravenous injection preparation HAs low toxicity, good safety and high solid tumor inhibition rate, and provides possibility for realizing cytarabine intravenous injection administration for treating solid tumors. In addition, the prodrug is grafted with the compound to generate the prodrug, so that the cytarabine is entrapped in the pore cavity of the metal-organic framework and is prepared into the metal-organic framework drug delivery system with the active targeting function, and a foundation is laid for the later research of the metal-organic framework drug delivery system based on the Ara small-molecule prodrug.
Finally, although the present invention has been described in detail with reference to the foregoing embodiments, it will be apparent to those skilled in the art that modifications and equivalents can be made in the embodiments described above, or equivalents may be substituted for elements thereof. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention. Although the embodiments of the present invention have been described with reference to the accompanying drawings, it is not intended to limit the scope of the present invention, and it should be understood by those skilled in the art that various modifications and variations can be made without inventive efforts by those skilled in the art based on the technical solution of the present invention.
Claims (7)
1. A preparation method of a metal organic framework drug delivery system with an active targeting function of cytarabine and a new indocyanine green small molecule prodrug Ara-IR820 is characterized in that the prodrug Ara-IR820 is dissolved in water, the prodrug Ara-IR820 is added into a zinc nitrate hexahydrate aqueous solution in a dropwise manner under the stirring condition, and a mixed solution is added into a 2-methylimidazole aqueous solution in a dropwise manner under the stirring condition; centrifuging, washing and drying the formed suspension to obtain a blue solid, namely a metal organic framework drug delivery system Ara-IR820@ ZIF-8 based on cytarabine and the new indocyanine green small molecule prodrug;
uniformly dispersing Ara-IR820@ ZIF-8 in an aqueous solution of hyaluronic acid HA, and stirring at room temperature; centrifuging, washing and drying the formed suspension to obtain a blue solid, namely a metal organic framework drug delivery system HA/Ara-IR820@ ZIF-8 with an active targeting function based on cytarabine and a new indocyanine green small molecule prodrug Ara-IR 820;
the structural formula of the prodrug Ara-IR820 is shown below:
2. the preparation method of claim 1, wherein the prodrug Ara-IR820 is prepared by combining neoindocyanine green with cytarabine through an amido bond to realize chemical modification of cytarabine amino.
3. The preparation method according to claim 2, wherein the reaction process of cytarabine and neoindocyanine green is as follows: reacting the new indocyanine green with 6-aminocaproic acid under the condition of adding triethylamine to generate a first-step product; and (3) separating and purifying the product, and reacting the product with cytarabine under the catalysis of triethylamine and ethyl chloroformate to generate the cytarabine and the new indocyanine green micromolecule prodrug Ara-IR 820.
4. The method of claim 3, comprising the steps of:
(1) dissolving a certain amount of new indocyanine green in anhydrous DMF, adding triethylamine, dissolving a certain amount of 6-aminocaproic acid in anhydrous DMF, adding the obtained solution into the reaction solution, and reacting under the conditions of inert gas protection and 85 ℃ oil bath to obtain IR 820-COOH;
(2) IR820-COOH purification: carrying out reduced pressure rotary evaporation to remove the anhydrous DMF (dimethyl formamide) serving as a reaction solvent, and carrying out vacuum drying to obtain a crude product; dissolving the crude product in methanol, purifying by silica gel column chromatography, and performing gradient elution by ethyl acetate and methanol to obtain an IR820-COOH pure product which is a blue solid; the volume ratio gradient of the ethyl acetate to the methanol is 10:1-3: 1;
(3) dissolving a certain amount of IR820-COOH in anhydrous DMF, adding triethylamine and ethyl chloroformate, and reacting under the conditions of inert gas protection and ice bath;
(4) dissolving a certain amount of cytarabine in anhydrous DMF, and adding the dissolved cytarabine into the reaction solution obtained in the step (3) to prepare the small molecule prodrug Ara-IR820 of cytarabine and neoindocyanine green;
(5) Ara-IR820 purification step: carrying out reduced pressure rotary evaporation to remove the anhydrous DMF (dimethyl formamide) serving as a reaction solvent, and carrying out vacuum drying to obtain a crude product; dissolving the crude product in methanol, purifying by silica gel column chromatography, and performing gradient elution by ethyl acetate and methanol to obtain an Ara-IR820 pure product which is a blue solid; the volume ratio gradient of the ethyl acetate to the methanol is 4:1-2.5: 1;
in the step (1), the ratio of the new indocyanine green: 6-aminocaproic acid: the molar ratio of triethylamine is 1:5: 5;
the IR 820-COOH: cytarabine: triethylamine: the molar ratio of ethyl chloroformate was 1:3:3: 3.
5. A metal organic framework drug delivery system with active targeting function based on cytarabine and a novel indocyanine green small molecule prodrug Ara-IR820 obtained by the preparation method of any one of claims 1 to 4.
6. Use of the system of claim 5 in the preparation of an anti-neoplastic agent for treating or ameliorating cancer in a tissue or organ, including but not limited to leukemia, solid tumors.
7. Use according to claim 6, wherein the system of claim 5 is ultrasonically dispersed homogeneously in water to produce an intravenous formulation.
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