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Delivery system resisting hepatitis B virus and biological preparation

A technology of polyethylene glycol and dioleoyl phosphatidyl ethanolamine, which is applied in the field of anti-hepatitis B virus and medicine, can solve the problems of adenovirus safety problems, toxic and side effects, etc., and can avoid safety problems and toxic and side effects. Effect

Inactive Publication Date: 2017-05-31
谭旭
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The existing CRISPR-Cas9 delivery in vivo is through the use of adenovirus expression system to express Cas9 and guideRNA, but the safety of adenovirus is still problematic
Moreover, the long-term stable expression of Cas9, a DNA enzyme, in cells by adenoviruses may also cause long-term toxic side effects

Method used

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  • Delivery system resisting hepatitis B virus and biological preparation
  • Delivery system resisting hepatitis B virus and biological preparation
  • Delivery system resisting hepatitis B virus and biological preparation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0117] Example 1: Cas9mRNA and sgRNA-B6 delivered by TT3-O3 formulation lipid nanoparticles in HepAD38 cell line Anti-HBV effect in

[0118] The TT3-O3 formulation delivers encapsulated RNA components in cultured cell lines in vitro without the need for exogenous addition of other substances. We therefore first evaluated the antiviral effects of Cas9 mRNA and sgRNA B6 delivered by TT3-O3 lipid nanoparticles in the HepAD38 cell line stably expressing the HBV genome.

[0119] Figure 7 The anti-HBV effect of Cas9mRNA and sgRNA-B6 delivered by TT3-O3 formulation lipid nanoparticles in HepAD38 cell line is shown. Figure 7 A shows a schematic diagram of the experimental process; Figure 7 B-7C shows the levels of the cell culture supernatant and intracellular hepatitis B virus e antigen respectively, the P value is calculated by t test, and the significance is indicated by *; Figure 7 D shows the real-time fluorescent quantitative PCR detection 3.5kb pre-genomic RNA expres...

Embodiment 2

[0124] Example 2: Cas9mRNA and sgRNA-B5 or sgRNA-B10 delivered by TT3-O3 formulation lipid nanoparticles in small Anti-HBV effect in mouse hepatitis B model

[0125] Its anti-hepatitis B virus effect in the mouse hepatitis B model was evaluated in a method similar to that in Example 1.

[0126] Figure 9 The anti-hepatitis B virus effect of Cas9mRNA and sgRNA-B5 or sgRNA-B10 delivered by TT3-O3 formula lipid nanoparticles is shown in the mouse hepatitis B model. Figure 9 A shows a schematic diagram of the mouse experiment process; Figure 9 B shows ELISA detection serum hepatitis B virus surface antigen level; Figure 9 C shows ELISA detection serum hepatitis B virus e antigen level; Figure 9 D shows that ELISA detects the surface antigen level of hepatitis B virus in the liver; Figure 9 E shows that ELISA detects the hepatitis B virus e antigen level in the liver; Figure 9 F shows the relative expression level of hepatitis B virus 3.5kb pre-genome RNA in the live...

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Abstract

The invention discloses lipid nanoparticles composed of fat polypeptide molecules, dioleoyl phosphatidyl ethanolamine, cholesterol and polyethylene glycol, which coat the sgRNA molecules of Cas9mRNA and target HBV gene conserved domain. The invention also discloses a preparation method and application of the lipid nanoparticles and a pharmaceutical composition and preparation comprising the lipid nanoparticles. The lipid nanoparticles and biological preparation disclosed by the invention can serve as a novel delivery system resisting hepatitis B virus.

Description

technical field [0001] The present invention relates to the field of medicine, in particular to the field of anti-hepatitis B virus, and more specifically to a novel delivery system and biological preparation for anti-hepatitis B virus. Background technique [0002] There are about 350 million people infected with hepatitis B virus in the world, and about 100 million of them are in China. These people cannot be cured because of hepatitis B virus latent. The main reason for the latency of HBV is that its genome exists stably for a long time in the patient's liver cell nucleus in the form of a covalently closed circular DNA (cccDNA). The existing anti-hepatitis B drugs are mainly interferon and nucleoside (acid) analogs, but neither of these two types of drugs can remove cccDNA, so they cannot cure hepatitis B. [0003] The emerging gene editing technology CRISPR-Cas9 system has been widely used in the field of scientific research due to its efficient and specific DNA cuttin...

Claims

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Application Information

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IPC IPC(8): A61K48/00A61K47/69A61K47/60A61K47/26C12N15/113C12N15/88
CPCA61K47/26A61K48/0008A61K48/005C12N15/1131C12N15/88C12N2310/10
Inventor 谭旭董一洲蒋超梅淼李斌
Owner 谭旭
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