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RNA interference blood vessel stent coating material with local antiplatelet effect and local anticoagulation blood vessel stent prepared therefrom

A technology of RNA interference and vascular stents, which is applied in anticoagulant treatment, coating, blood diseases, etc., can solve the problems of no combination of vascular stents and no anti-platelet effect

Inactive Publication Date: 2013-10-23
BEIJING ZHONGFU YOUXIN MEDICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because the use of RNAi technology can specifically inhibit the expression of specific genes, this technology has been widely used in the field of gene therapy for exploring gene functions and infectious diseases and malignant tumors, but it has not been used to silence human cyclooxygenase 1. Gene PTGS1 and human P2RY12 type ADP receptor gene play an anti-platelet effect, and there is no report on the combined use with vascular stents

Method used

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  • RNA interference blood vessel stent coating material with local antiplatelet effect and local anticoagulation blood vessel stent prepared therefrom
  • RNA interference blood vessel stent coating material with local antiplatelet effect and local anticoagulation blood vessel stent prepared therefrom
  • RNA interference blood vessel stent coating material with local antiplatelet effect and local anticoagulation blood vessel stent prepared therefrom

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0095] Example 1: Determination of target sequences in cyclooxygenase 1 gene and P2RY12 type ADP receptor gene.

[0096] The gene sequence data of the cyclooxygenase 1 gene (hereinafter referred to as PTGS1) and the P2RY12 type ADP receptor gene (hereinafter referred to as P2RY12) were obtained from the public database published by the National Center for Biological Information (NCBI), and the corresponding transcripts were used here. sequence, the messenger RNA (mRNA) sequence. After obtaining the sequence data, first organize the sequence into FASTA format text, and then use VBA (Visual Basic for Application) computer language programming to carry out computer-aided manual search for the target sequence in the sequence. The search standard is formulated according to our long-term laboratory experience . Specifically include:

[0097] (1) The length reference is 19nt,

[0098] (2) Among the 6 bases at the 5' end and the 3' end, the sum of the numbers of guanine (G) and cyt...

Embodiment 2

[0133] Example 2: Assembly, synthesis, purification and annealing of cloned inserts of expression-type short-chain interfering RNA recombinant plasmids targeting PTGS1 and P2RY12

[0134] Assembly reference for cloned inserts Figure 5 The model diagram is carried out, and the specific assembly results are as follows.

[0135] Insert sequence assembly for PTGS1 gene:

[0136] Seq ID No. 7siR-PTGS1-is1-F:

[0137] 5'-GATCCAAGGGTGGGGAGGGGATGGTTCAAGAGACCATCCCCTCCCCACCCTTAGA-3'

[0138] Seq ID No.8siR-PTGS1-is1-R:

[0139] 5'-AGCTTCTAAGGGTGGGGAGGGGATGGTCTCTTGAACCATCCCCTCCCCACCCTTG-3'

[0140] Seq ID No.9siR-PTGS1-is2-F:

[0141] 5'-GATCCTTCTTGCTGTTCCTGCTCCTTCAAGAGAGGAGCAGGAACAGCAAGAAAGA-3'

[0142] Seq ID No. 10siR-PTGS1-is2-R:

[0143] 5'-AGCTTCTTTCTTGCTGTTCCTGCTCCTCTCTTGAAGGAGCAGGAACAGCAAGAAG-3'

[0144] Seq ID No. 11siR-PTGS1-is3-F:

[0145] 5'-GATCCTTCACCCACTTCCTGCTCATTCAAGAGATGAGCAGGAAGTGGGTGAAAGA-3'

[0146] Seq ID No.12siR-PTGS1-is3-R:

[0147] 5'-AGCTTCTTTCACCCACT...

Embodiment 3

[0173] Example 3: Assembly fragments connected to vector, E.Coli transformation and screening

[0174] Material:

[0175] Double-stranded inserts prepared in Examples 1 and 2.

[0176] The CMV vector plasmid was purchased from Ambion Company, USA.

[0177] Endonuclease, T4 DNA ligase, Escherichia coli (E. Coli), LB medium, agarose, and ampicillin were all purchased commercially.

[0178] method

[0179] The CMV vector plasmid was digested with BamHI and HindIII at 37°C for 1 hour, and the buffer used for digestion was the buffer distributed with the endonuclease by the supplier. After the digestion is completed, it is separated by 2% agarose electrophoresis, recovered by the conventional low-melting point agarose recovery method, and then ligated with the annealed double-stranded insert sequence. The ligation reaction formula is as follows:

[0180] 50mM Tris-HCl (pH 7.5)

[0181] 10mM MgCl 2

[0182] 1mM ATP

[0183] 10mM dithiothreitol

[0184] 90ng of annealed doub...

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Abstract

The invention which provides an RNA interference blood vessel stent coating material with the local antiplatelet effect and a local anticoagulation blood vessel stent prepared therefrom relates to the blood vessel stent coating technology in the field of biomedicines. The coating material is characterized in that: the effective component of the coating material comprises at least one RNA interference recombinant plasmid, and an inserted DNA fragment contained in the RNA interference recombinant plasmid can express short interference RNA which can inhibit the expression of human cyclooxygenase-1 gene PTGS1 or human P2RY12 type ADP receptor gene. According to the coating material of the invention, a gene regulation approach is adopted to realize the antiplatelet mechanism of two antiplatelet medicines which comprise cyclooxygenase inhibitors which are represented by aspirin and ADP receptor inhibitors which are represented by clopidogrel, the blood vessel intervention treatment anticoagulation is localized, and usage taboos and a series of side effects, which are caused by the systematic distribution of the medicines, are avoided.

Description

technical field [0001] The invention relates to a vascular stent coating technology in the field of biomedicine, in particular to an RNA interference vascular stent coating material with a local anti-platelet effect and a stent made of the same. Background technique [0002] Percutaneous coronary intervention (PCI for short) has become one of the important treatments for coronary heart disease. At present, there are more than 1,200 institutions in my country carrying out PCI, with an annual operation volume of 250,000 cases. The essence of PCI is to open the coronary artery and realize the revascularization of the diseased area. The equipment used in PCI has gradually developed from balloon expansion to vascular stents. The development of vascular stents has gone through bare metal stents (BMS for short) and drug-eluting stents (DES for short). While the use of drug-eluting stents has brought huge benefits, there are also obvious problems. Wherein, the most serious proble...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61L33/06A61L31/08A61L31/10A61K48/00A61P7/02
Inventor 周儒伦郭威早张建军
Owner BEIJING ZHONGFU YOUXIN MEDICAL TECH