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Nucleic acid medicine loading system for targeted therapy and preparation method of nucleic acid medicine loading system

A drug-carrying system and targeted therapy technology, which is applied in the direction of pharmaceutical formulations, medical preparations with non-active ingredients, medical preparations containing active ingredients, etc., can solve problems such as lack of drugs, unstable complexes, complex organic reactions, etc. , to achieve the effect of simple and efficient preparation process, inhibition of cancer cell growth, and high drug loading

Active Publication Date: 2012-10-10
HUNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these drug carrier systems have a series of defects: complex synthesis, purification, and further functionalization, which reduce the feasibility of clinical application of the product; many nanomaterials containing metal or silicon components have poor biocompatibility; many loaded drugs are It is loaded into the carrier through unstable non-covalent binding forces, resulting in limited stability of the drug-carrier complex; in addition, traditional targeted transport can only be achieved through passive targeting, that is, the EPR (Enhanced Permeation and Retention) effect, Carry out targeted delivery of drugs, and the low efficiency of this targeting also makes the effect of targeted therapy very limited; finally, a large number of drug-loading systems can only transport drugs to the cell surface, lacking further active delivery of drugs into cells function of
However, these techniques have a series of disadvantages, such as complex organic reactions, low yield, and high cost of chemical modification.
In some other reports, the non-covalent interaction between the nucleic acid of a specific sequence and the drug was used to form a complex of the drug and the nucleic acid aptamer, but these weak interactions determine the instability of the complex, thus hindering their clinical application. application on

Method used

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  • Nucleic acid medicine loading system for targeted therapy and preparation method of nucleic acid medicine loading system
  • Nucleic acid medicine loading system for targeted therapy and preparation method of nucleic acid medicine loading system
  • Nucleic acid medicine loading system for targeted therapy and preparation method of nucleic acid medicine loading system

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Experimental program
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Effect test

Embodiment 1

[0048] Using an automatic DNA synthesizer, synthesize 1 μmole of the required nucleic acid aptamer by solid-phase synthesis. The resulting product was deprotected, the DNA was dropped from the glass beads used in the synthesis, and the DNA was precipitated with ethanol (70%, 2.5 times the volume of the DNA solution) and NaCl (3 M, 0.1 times the volume of the DNA solution). The precipitated DNA was dissolved in TEAA (0.1 M).

[0049] The above product was purified by HPLC and dried. The DMT protecting group on the DNA was removed with acetic acid, and the DNA was precipitated again by the ethanol precipitation method mentioned above, and dried. The resulting DNA was dissolved in DNA water and stored at low temperature (20 o C) Spare.

Embodiment 2

[0051] In the buffer used (20 mM sodium phosphate (pH 7.0), 150 mM sodium chloride, 0.5 mM EDTA), the following reaction system was prepared: aptamer 20 μM, drug 400 μM, formaldehyde 0.37%. Put the above reaction system at 10 o C, reacted for 12 hours.

[0052] The obtained product was purified by HPLC or molecular sieve, ethanol precipitation and other methods. Such as image 3 As shown, the sample containing only nucleic acid aptamer only absorbs at 260 nm when analyzed by HPLC; in the obtained nucleic acid aptamer-drug adduct, there is both a strong light absorption at 260 nm (mainly from DNA) , also comes from the characteristic absorption of drugs (such as the absorption of doxorubicin at 490nm); in addition, the inventors have found through a large number of experiments that these methods can not only remove unreacted free drugs in the system, but also remove some Drugs that can form complexes with DNA by intercalation (formaldehyde is not added to the reaction system...

Embodiment 3

[0055] The prepared adducts were characterized by mass spectrometry (ESI-MS), the molecular weights of the adducts and an identical nucleic acid aptamer that did not form adducts were compared, and the number of drug molecules on an aptamer DNA chain was calculated. Such as Figure 4 As shown, compared with the sample containing only the nucleic acid aptamer sgc8, the adduct formed by sgc8 and Dox was analyzed by ESI-MS and showed that the adduct contained multiple products with a molecular weight larger than that of sgc8. By comparing the adducts of these products The molecular weight is higher than that of simple sgc8, and the inventors found that they are just integer multiples of the total molecular weight of one Dox and one methylene. Further calculations show that the number of Dox molecules contained in the corresponding adducts is 1-6, respectively.

[0056] The prepared adducts were characterized by fluorescence photometric analysis, and the fluorescence intensity of...

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Abstract

The invention discloses a nucleic acid medicine loading system for targeted therapy and a preparation method of the nucleic acid medicine loading system. The system is adduct consisting of DNA (deoxyribonucleic acid) aptamers as medicine carriers and targeted probes and anticancer medicine loaded onto the DNA aptamers. The targeted identification specificity, high stability and easy modification of the DNA aptamers are utilized, and the DNA aptamer-anticancer medicine adduct is prepared, so small molecule medicine is modified onto the DNA aptamers. The DNA aptamers are very stable at the solid state or low temperature liquid state (such as the temperature below 10 DEG C) and is suitable for being stored for a long time. The adduct maintains the identification specificity of the original DNA aptamers to targeted cancer cells, medicine can be conveyed to cancer cells in a targeted way and can be gradually released at the physiological temperature, the special killing and injury effect on the targeted cancer cells is generated, the side effect is reduced, and the curative effect is improved. The nucleic acid medicine loading system and the preparation method have the advantages that the preparation process is simple, economy is realized, the efficiency is high, and the preparation process is suitable for mass production and is particularly suitable for being used for preparing the targeted medication anticancer medicine.

Description

technical field [0001] The invention belongs to the technical field of targeted drug delivery systems, and relates to targeted drug delivery for cancer treatment, in particular to a nucleic acid aptamer-based carrier and anticancer drug adduct as well as its preparation and application. Background technique [0002] Chemotherapy is currently the most widely used cancer therapy in clinical practice. However, the nonspecificity of traditional chemotherapeutic drugs causes them to induce cytotoxicity and inhibit cell expansion not only to rapidly growing cancer cells but also to rapidly growing normal, healthy cells, including bone marrow hematopoietic cells, hair follicles, oral , digestive tract, reproductive system cells, etc. This toxicity to healthy cells leads to obvious side effects of chemotherapy, and thus only a very limited maximum tolerated drug dose can be used clinically. In addition, the bioavailability of traditional small molecule chemotherapy drugs is very l...

Claims

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Application Information

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IPC IPC(8): A61K47/48A61K31/704A61P31/00A61P35/00A61K47/26
Inventor 谭蔚泓朱贵志
Owner HUNAN UNIV
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