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Application of hydrophobic modified polypeptide in preparation of microRNA related nucleic acid delivery system

A nucleic acid delivery and hydrophobization technology, applied in the field of biomedicine, can solve the problems of unclear function of microRNAs, restriction of microRNAs function, unsatisfactory delivery efficiency, etc., to achieve inhibition of tumor growth and migration, low cytotoxicity, and short production cycle Effect

Pending Publication Date: 2022-04-22
WEST CHINA HOSPITAL SICHUAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It is worth noting that the function of the newly discovered microRNAs may not be clear, and the inefficiency of microRNAs delivery in cells limits the research on the function of microRNAs
At present, cationic liposomes and polymers have been used for the delivery of microRNA, but due to its potential cytotoxicity, safer and more effective carriers still need to be developed, and although the delivery efficiency in vitro is high, its delivery efficiency in vivo but lower
[0003] MicroRNA-related nucleic acids, such as microRNA analogs and microRNA inhibitors, have also been extensively studied in recent years. However, the delivery efficiency of these microRNA-related nucleic acids in vivo is also unsatisfactory.

Method used

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  • Application of hydrophobic modified polypeptide in preparation of microRNA related nucleic acid delivery system
  • Application of hydrophobic modified polypeptide in preparation of microRNA related nucleic acid delivery system
  • Application of hydrophobic modified polypeptide in preparation of microRNA related nucleic acid delivery system

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0072]For example, the preparation of the delivery system will be handled, formulated, incubated and used in solution, which may therefore contain pharmaceutically acceptable concentrations of salts, buffers, preservatives and various media.

[0073] In order to prepare a suitable dosage form, various types of pharmaceutically acceptable stabilizers, excipients, lyoprotectants and other auxiliary components can be added. The main dosage form of the vaccine prepared by the delivery system for delivering microRNA-associated nucleic acid of the present invention is an injection, which can be made into an injection or a freeze-dried preparation. The route of use can be direct intratumoral injection.

[0074] If desired, the vaccine may further include an adjuvant to assist in the induction or restimulation of an immune response, including prolonging or boosting the immune response. The adjuvant can be selected from various adjuvants known in the art according to actual usage cond...

Embodiment 1

[0086] Example 1 Preparation and Characterization of DP7-C / microRNA Complex and DP7-C / inhibitor Complex

[0087] 1. The particle size and potential of DP7-C complexed with microRNA and inhibitor respectively

[0088] For the synthesis of DP7-C and the preparation of DP7-C micelles, please refer to the Chinese patent application with application number 201710527268.4 (document number CN 107446019 A). Dilute the 10mg / ml DP7-C stock solution stored at 4°C to 20μg / ml, add 4μg microRNA or 5μg microRNA inhibitor to 1ml, mix well and incubate at room temperature for 10min. Put the prepared solution into the particle size potential cup, and use the Malvern particle size potential meter (Malvern ZetaSizer, UK) to measure the particle size distribution and zeta potential of DP7-C micelles and their complexes respectively compounded with microRNA and inhibitor Size Characterization Determination. All experiments were repeated three times to obtain the median value.

[0089] The result...

Embodiment 2

[0096] Example 2 Efficiency and serum stability of DP7-C transfection microRNA and inhibitor

[0097] 1. The efficiency of DP7-C transfection of microRNA and inhibitor to CT26 cells and 4T1 cells

[0098] CT26 cells and 4T1 cells were cultured in 24-well plates with 1640 dual medium (RPMI 1640 + 10% FBS + 1% PS), and CT26 cells were plated in 1 × 10 per well. 5 , 4T1 cells were plated 2×10 per well 5 . After 24 hours of plating, replace the culture medium with 200 μl of 1640 double-free medium, add 30 μl of carrier / FAM-microRNA or carrier / FAM-inhibitor complexes incubated in double-free 1640 for 10 minutes, and the carrier materials are DP7-C and PEI25K respectively and Lipo2000. Grouped as follows:

[0099] DP7-C(1.2μg) / FAM-microRNA(0.24μg), DP7-C(0.96μg) / FAM-inhibitor(0.24μg), Lipo2000(0.48μg) / FAM-microRNA(0.24μg), Lipo2000(0.48μg ) / FAM-inhibitor(0.24μg), PEI25K(0.48μg) / FAM-microRNA(0.24μg), PEI25K(0.48μg) / FAM-inhibitor(0.24μg). After 4 hours, the culture solution was ...

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Abstract

The invention belongs to the field of biological medicine, and particularly relates to application of hydrophobic modified polypeptide in preparation of a microRNA related nucleic acid delivery system. The invention aims to solve the technical problems of improving the intracellular delivery effect of microRNA related nucleic acid and reducing the cytotoxicity of a carrier. According to the scheme for solving the technical problem, the application of the hydrophobic modified polypeptide in preparation of the microRNA related nucleic acid delivery system is provided, the sequence of the cationic polypeptide is VQWRIRVAVIRK, and hydrophobic modification is that a hydrophobic fragment is coupled to the nitrogen terminal of the cationic polypeptide. According to the present invention, the hydrophobic modified VQWRIRVAVIRK polypeptide can efficiently deliver microRNA related nucleic acid to cells so as to well provide the vaccine effect, and further has advantages of low toxicity, safety, short preparation period, and good application prospect.

Description

technical field [0001] The invention belongs to the field of biomedicine, and in particular relates to the use of hydrophobically modified polypeptides in the preparation of microRNA-related nucleic acid delivery systems. Background technique [0002] RNA interference has a highly efficient inhibitory effect on protein expression at the post-transcriptional level, so it has broad application prospects in disease treatment. This approach can even target previously untreatable targets, resulting in a range of potential therapeutic effects. MicroRNA is a small non-coding RNA molecule, 20-24 nucleotides in length, that regulates mRNA degradation or translational repression of target genes through incomplete base pairing with the 3'-UTR (untranslated region), and Can target multiple mRNA transcripts, thereby regulating the expression of target genes. The expression changes of microRNA regulate the important biological processes of tumorigenesis, growth, invasion and metastasis....

Claims

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Application Information

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IPC IPC(8): A61K47/42A61K9/107A61K39/00A61P35/00A61P35/04C07K7/08C12N15/87
CPCA61K47/42A61K9/1075A61K39/00C07K7/08C12N15/87C12N15/111A61P35/00A61P35/04C12N2310/141C12N2310/113C12N2320/32A61K2039/53Y02A50/30
Inventor 杨莉张瑞
Owner WEST CHINA HOSPITAL SICHUAN UNIV
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