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Non-viral vector formed based on covalent assembly of terpyridyl ruthenium catalytic oligopeptides, and preparation method and application of non-viral vector

A non-viral carrier, terpyridine ruthenium chloride technology, applied to the preparation method of peptides, chemical instruments and methods, medical preparations of non-active ingredients, etc.

Pending Publication Date: 2021-10-26
ANHUI NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Despite great progress, limitations remain

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  • Non-viral vector formed based on covalent assembly of terpyridyl ruthenium catalytic oligopeptides, and preparation method and application of non-viral vector
  • Non-viral vector formed based on covalent assembly of terpyridyl ruthenium catalytic oligopeptides, and preparation method and application of non-viral vector
  • Non-viral vector formed based on covalent assembly of terpyridyl ruthenium catalytic oligopeptides, and preparation method and application of non-viral vector

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Embodiment 1

[0038] A DNA-[Ru(bpy) based 3 ] 2+ A preparation method for catalyzing the covalent assembly of short peptides to form a non-viral vector, the preparation method comprising the following steps:

[0039](1) Heat equal volumes of DNA S1 solution and DNA S2 solution with a concentration of 2 μM to 90°C, take it out after 10 minutes, cool to room temperature, and then store in the upper layer of the refrigerator at a temperature of 4-10°C until the dsDNA structure is formed, Obtain dsDNA solution;

[0040] The gene sequence of the DNA S1 is: 3'-GGA TTG GAG TTC CTC CAG CGT GCG CCA TCC TTC CCA TCCTCC TCC -5'; the underlined part is the ASO sequence that can reduce the expression of the anti-apoptotic protein Bcl-2. If it is replaced with a nucleic acid sequence that can treat other diseases, the targeted treatment of other diseases can also be achieved;

[0041] The gene sequence of the DNA S2 is: 3'-GAG GAA CTC CAA TCC-5';

[0042] (2) 1mL 2mM [Ru(bpy) 3 ] Cl 2 The solut...

Embodiment 2

[0046] Others are with embodiment 1, test [Ru(bpy) when carrying out step (2) 3 ] Cl 2 , DNA-[Ru(bpy) 3 ] 2+ fluorescence spectrum, such as image 3 As shown in a), generate DNA-[Ru(bpy) 3 ] 2+ When the 625nm characteristic fluorescence peak intensity increases, it proves that DNA-[Ru(bpy)3] 2+ The formation of small molecule; Carry out the measurement of Zeta potential when carrying out step (2-4) simultaneously, as image 3 As shown in b), when [Ru(bpy) 3 ] Cl 2 dsDNA solution was added to the solution to prepare DNA-[Ru(bpy) 3 ] 2+ , DNA-[Ru(bpy) 3 ] 2+ The absolute value of the Zeta potential of dsDNA becomes smaller than that of dsDNA, which is due to [Ru(bpy) 3 ] 2+ With a positive charge, it is further proved that [Ru(bpy) 3 ] 2+ Successful insertion in dsDNA yields DNA-[Ru(bpy) 3 ] 2+ ; when DNA-[Ru(bpy) 3 ] 2+ After mixed with YY-12 and irradiated with light to obtain a non-viral vector, the zeta potential was further corrected, which may be due to ...

Embodiment 3

[0048] Others are the same as in Example 1, when carrying out step (4), test the fluorescence spectrum of YY-12 and the non-viral vector obtained by the reaction, such as image 3 As shown in a), the fluorescence peak of the generated peptide-nucleic acid nanospheres is 410nm, which proves the formation of non-viral vectors; after the reactants are mixed evenly, the ultraviolet absorption diagrams of the test system are sampled under different irradiation times, and the test time is cut off at 600s, 6min Afterwards, the peak value tends to be flat and no longer rises, and the UV absorbance value at this time reaches the maximum, which also proves the success of the covalent cross-linking of the short peptide and the successful preparation of a non-viral vector.

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Abstract

The invention discloses a non-viral vector formed based on covalent assembly of terpyridyl ruthenium catalytic oligopeptides, and a preparation method and application of the non-viral vector. The preparation method comprises the following steps of: mixing antisense oligonucleotide and a terpyridyl ruthenium chloride solution, and performing incubation to obtain a DNA-[Ru(bpy)3]<2+> solution; and mixing the DNA-[Ru(bpy)3]<2+> solution, a tyrosine-containing targeting peptide solution and an ammonium persulfate solution, performing white light irradiation, and performing centrifuging to obtain the non-viral vector. The method has the characteristics of simple synthesis, greenness, high efficiency and the like; and the non-viral vector can be applied to gene delivery or used as a targeted drug.

Description

technical field [0001] The invention belongs to the technical field of gene delivery carriers, and in particular relates to a non-viral carrier based on the covalent assembly of short peptides catalyzed by terpyridine ruthenium and its preparation method and application. Background technique [0002] Drug therapy involves a variety of complex mechanisms. Building an intelligent and adaptive system is a major challenge in current cancer research and clinical practice. Cancer cells can develop a variety of defense strategies to evade the effects of therapeutic drugs. These defense mechanisms are often associated with changes in the expression levels of resistance-associated proteins, ultimately leading to the failure of anticancer treatments. To address this challenge, antisense oligonucleotides (ASO) and small interfering RNA (siRNA) are nucleic acid therapy methods that can inhibit the expression of drug resistance-related proteins to restore the drug sensitivity of cancer c...

Claims

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Application Information

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IPC IPC(8): C07K7/08C07K1/107C12N15/87A61K9/00A61K41/00A61K47/64A61K47/69A61K48/00A61P35/00
CPCC07K7/08C07K19/00C12N15/87A61K48/0025A61K41/0057A61K9/0009A61P35/00A61K47/6929A61K47/64Y02B20/00
Inventor 王广凤李娜
Owner ANHUI NORMAL UNIV
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