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A kind of pharmaceutical composition and its application

A composition and drug technology, applied in the field of biomedicine, can solve problems such as inability to activate pDC

Active Publication Date: 2021-02-05
郭文江
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Type B CpG-ODN is characterized by the presence of TCGTCG and GTCGTT, and has strong immunostimulatory activity on B cells, but cannot activate pDC

Method used

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  • A kind of pharmaceutical composition and its application
  • A kind of pharmaceutical composition and its application
  • A kind of pharmaceutical composition and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061]Purification experiment and purity detection of sample subtilisin

[0062]1.1 Preparation of fermentation broth

[0063]The strain is Bacillus subtilis B.subtilis CVCC20076, purchased from the China Industrial Microbial Culture Collection and Management Center; it will be inoculated on a solid medium and cultivated until the lawn is covered with the surface of the medium for the first generation; scrape the lawn and inoculate to In the liquid medium, shake culture until the second generation of the bacterial solution; inoculate it into the seed tank, stir culture until the production seeds are the third generation, inoculate the production seeds into the culture tank, stir culture and harvest 200L of fermentation broth .

[0064]1.2 Purification steps and results

[0065]Centrifugation and filtration: Centrifuge the fermentation broth on a centrifuge at 4°C, centrifuge at 10,000 r / min for 15 minutes to obtain the supernatant, and discard the precipitate.

[0066]Anion exchange chromatography...

Embodiment 2

[0073]In vivo anti-tumor activity experiment of subtilisin and nucleic acid

[0074]2.1 Experimental equipment and experimental animals

[0075]Electronic balance (Shanghai Precision Scientific Instruments, JA1003N), mouse scale (Shanghai Yousheng Weighing Apparatus Co., Ltd., ASC-A), blood cell analyzer (BT2000-PLUS), mouse gastric tube, etc.

[0076]C57BL / 6, Balb / c mouse, all female; grade SPF; age: 5-7 weeks; weight: average body weight at the beginning of the experiment 18.0-25.0g; from Shanghai Slack Laboratory Animal Co., Ltd. (SLAC ).

[0077]2.2 Grouping of experimental animals and model establishment

[0078]First, according to body weight, they were randomly divided into model control group, positive drug group, subtilisin group, nucleic acid group, subtilisin nucleic acid compound formula group, and combined administration group, with 8 animals in each group.

[0079]Model establishment: Dilute various tumor cells with saline 1:3 to prepare cell suspensions, and inoculate the well-growing ...

Embodiment 3

[0102]Application of subtilisin and nucleic acid as vaccine adjuvant in recombinant hepatitis B vaccine

[0103]3.1 Materials

[0104]Recombinant yeast hepatitis B vaccine, Shenzhen Kangtai, batch number c201301003; Mouse hepatitis B virus antibody (HBVAb) ELISA test kit.

[0105]3.2 Experimental animals

[0106]Clean Balb / c mice, female mice, 6-8 weeks old, weighing 18-22g.

[0107]3.3 Main reagents

[0108]Subtilisin, CpG2006, polyinosin (purchased from Hangzhou Meiya Pharmaceutical Co., Ltd.).

[0109]3.4 Animal grouping and immunoassay

[0110]The mice were randomly divided into 6 groups with 10 mice in each group. The grouping situation is as follows: 1. Normal saline control; 2. Recombinant HBsAg protein vaccine; 3. Recombinant HBsAg protein vaccine 60μg+CpG2006(5μg); 4. Recombinant HBsAg protein vaccine 60μg+ Polyinosin (5μg); 5. Recombinant HBsAg protein vaccine 60μg+subtilisin (25μg); 6. Recombinant HBsAg protein vaccine 60μg+CpG2006(5μg)+subtilisin (25μg); 7. Recombinant HBsAg protein vaccine 60μ...

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Abstract

The invention relates to the field of biomedicine, in particular to a pharmaceutical composition and its application. A pharmaceutical composition mainly consists of subtilisin and nucleic acid substances, and the mass content of subtilisin in the composition is 0.01-98%. In the present invention, subtilisin and nucleic acid substances are used in combination to find that subtilisin has the characteristics of enhancing the activity of nucleic acid substances, and the two cooperate with each other, and can be used as a drug for treating cancer and viral infectious diseases, as well as a vaccine adjuvant Related drugs, and / or other drugs, antigens or adjuvants are compounded to form a composition for the treatment of related diseases to achieve better anti-tumor and immune-enhancing effects; and compared with ligands of nucleic acid substances, subtilin The drug has the characteristics of low toxicity and obvious effect, which shows that the pharmaceutical composition provided by the present invention also has the characteristics of low toxicity and obvious effect.

Description

Technical field[0001]The present invention relates to the field of biomedicine, in particular to a pharmaceutical composition and its application.Background technique[0002]In 1980, Nusslein-Volhard et al. discovered the Toll-like receptor gene during the study of the embryonic development of Drosophila [Nusslein-Volhard C, Wieschaus E., Nature, 1980, 287(5785):795-801]. Currently, they are breast-feeding There are 11 members of the human TIRs family that have been found in animals and humans. Dendritic cells, the body's strongest antigen-presenting cells, can express TLRs. With the help of TLR to recognize molecules with PAMP such as LPS, GpG-DNA, peptidoglycan, lipoprotein, and cell wall components of mycobacteria, dendritic cells are activated and mature, providing costimulatory signals for adaptive immunity. Therefore, TLR is a bridge for the activation of dendritic cells caused by microbial components. Toll-like receptors have a regulatory effect on the type of adaptive immune r...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K38/17A61K39/39A61P35/00A61P31/12A61P37/04A23L33/10A61K31/505
CPCA23V2002/00A61K31/505A61K38/1709A61K39/39A61K2039/55516A61K2039/55561A61K2300/00A23V2200/324A23V2200/308
Inventor 郭文江
Owner 郭文江