Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Recombinant adeno-associated viral vector with human papillomavirus type 16 multi-point mutant E7<mm> antigen genes, method for constructing recombinant adeno-associated viral vector and application thereof

A technology of human papillomavirus, hpv-16e7mm, which is applied in the field of preparation of anti-HPV-16 infection and therapeutic drugs for related diseases, and can solve problems such as safety risks

Active Publication Date: 2015-12-23
GUANGDONG TOPHEALTH BIOTECH
View PDF3 Cites 2 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, there are certain risks in terms of safety in stimulating the occurrence of immune responses with wild-type HPV-16E7 antigen in vivo and in vitro

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Recombinant adeno-associated viral vector with human papillomavirus type 16 multi-point mutant E7&lt;mm&gt; antigen genes, method for constructing recombinant adeno-associated viral vector and application thereof
  • Recombinant adeno-associated viral vector with human papillomavirus type 16 multi-point mutant E7&lt;mm&gt; antigen genes, method for constructing recombinant adeno-associated viral vector and application thereof
  • Recombinant adeno-associated viral vector with human papillomavirus type 16 multi-point mutant E7&lt;mm&gt; antigen genes, method for constructing recombinant adeno-associated viral vector and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0072] Example 1. Construction of recombinant adeno-associated virus vectors AAV / HPV-16E7 and AAV / HPV-16E7 mm

[0073] 1. Materials and their sources:

[0074] A. Four kinds of adeno-associated virus (AAV) vectors: pAAV / p5 with AAVp5 promoter, pAAV / CMVp with macrophage virus (CMV) promoter, pAAV / SV40p with SV40 virus early promoter and pAAV / β-actinp with human β-actin promoter. Known adeno-associated virus vectors have a p5 promoter. In order to increase the transcription level of the target gene, the p5 promoter in the recombinant adeno-associated virus vector can be replaced with a macrophage virus (cytomegalovirus, CMV) promoter, beta actin The promoter and one of the SV40 virus promoters, the four AAV vectors are only different in the promoter, and the rest of the gene structure is exactly the same, that is, it has the complete repeat terminal fragment (TR) sequence at both ends of the AAV2 type, and at both ends of the TR There are 9 nucleotide fragments inserted at th...

Embodiment 2

[0089] Embodiment 2, preparation of recombinant adeno-associated virus (rAAV) and virus titer determination

[0090] Materials and their sources:

[0091] A. Carrying HPV-16E7 constructed in Example 1 mm and HPV-16E7 antigen gene recombinant adeno-associated virus vector (AAV / HPV-16E7 mm and AAV / HPV-16E7).

[0092] B. Auxiliary plasmid pHelper containing the Rep gene and Cap gene of AAV: successfully constructed by the inventors of this patent application Liu Yong et al. (Liu, Y., SantinAD., ManeM., Chiriva-Internati, M., ParhamGP., RavaggiA ., and Hermonat, P.L. Transduction and Utility of the Granulocyte-Macrophage Colony-Stimulating Factor Gene into Monocytes and Dendritic Cells by Adeno-Associated Virus. Journal of Interferon and Cytokine Research. 20:21–30. 2000).

[0093] C. AAV-HEK293 cells containing adenovirus genes (E1, E2A, E4, VAI and VAII genes) integrated in the cell chromosome and expressed: established by Liu Yong, the inventor of this patent application, et...

Embodiment 3

[0113] Embodiment 3, recombinant adeno-associated virus AAV / HPV-16E7 mm Observation on tumorigenicity of primary cervical epithelial cells infected by virus and AAV / HPV-16E7 virus

[0114] Materials and their sources:

[0115] A. rAAV virus: the recombinant adeno-associated virus (AAV / HPV-16E7 that embodiment 2 obtains mm virus and AAV / HPV-16E7 virus).

[0116] B. Keratinocyte-SCF cell culture medium: purchased from American Life Technology Company.

[0117] C. Primary cervical epithelial cells: obtained by separating from normal cervical epithelial tissue by conventional methods.

[0118] Tumorigenicity Observation Experiment

[0119] Put the primary cervical epithelial cells into a 10.0cm cell culture dish, immediately add 10mL Keratinocyte-SCF cell culture medium, and culture them in a carbon dioxide incubator at 37°C. After the cells are completely adhered to the wall, take out the culture dish, remove 7mL of the culture medium, and then add the recombinant adeno-asso...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

PropertyMeasurementUnit
Titeraaaaaaaaaa
Login to View More

Abstract

The invention discloses a recombinant adeno-associated viral vector with HPV-16 (human papillomavirus-16) E7<mm> antigen genes and a method for constructing the recombinant adeno-associated viral vector. The method includes mutating tumorigenicity HPV-16 E7 antigen genes at multiple points to obtain non-tumorigenicity E7 antigen genes; inserting the mutated genes into an adeno-associated viral vector without structural genes to obtain the recombinant adeno-associated viral vector. The recombinant adeno-associated viral vector and the method have the advantages that the mutant HPV-16 E7<mm> antigen genes carried by recombinant adeno-associated viruses can be delivered into monocyte-dendritic cell lines to be used for stimulating effect cells of immune systems; as proved by experiments, growth of HPV-16 E7 positive cells can be effectively inhibited by CTL (cytotoxic lymphocytes) induced by DC (dendritic cells) infected by the recombinant adeno-associated viruses for patients in in-vitro and in-vivo manners, or the HPV-16 E7 positive cells can be effectively killed by the CTL for the patients in the in-vitro and in-vivo manner, and tumorigenicity can be prevented; the recombinant adeno-associated viral vector or associated products can be used for preparing anti-tumor medicine for treating HPV-16 infection or diseases associated with the HPV-16 infection.

Description

technical field [0001] The invention relates to a carrier in the biological field and its application, in particular to a human papillomavirus type 16 (Human Papillomavirus Type 16, HPV-16) multi-point mutant type E7 mm The recombinant adeno-associated virus vector (rAAV) of the antigen gene and its construction method and its application in the preparation of anti-HPV-16 infection and related diseases (such as tumors caused by HPV-16 infection) therapeutic drugs. , its construction method and its application in the preparation of medicines against HPV-16 infection and related diseases. Background technique [0002] The genetic structure of adeno-associated virus (AAV) has been identified. In 1983, Samulski et al. described the terminal repeats of AAV (upstream 5' end segment, downstream 3' end segment) (SamulskiRJ, SrivastavaA, BernsKI, MuzyczkaN. Rescueofadeno-associated virus from recombinant plasmamids: gene correctionwithin the terminal repeats of AAV. Cell.33:135-143....

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12N15/864C12N7/01C12N5/10A61K48/00A61K39/12A61P31/20A61P35/00
Inventor 刘勇陈巧林曾昭鹏高洪吉龚研浩董文娟张慧
Owner GUANGDONG TOPHEALTH BIOTECH
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com