Freeze-dried preparation of recombinant bacillus calmette guerin vaccine for treatment and preparation method and application of freeze-dried preparation

A technology for reconstituting BCG vaccine and freeze-dried preparations, which can be used in freeze-dried delivery, medical preparations of inactive ingredients, drug combinations, etc., can solve problems such as concentration increase, and achieve the effects of preventing damage, high quantity, and good clinical application value.

Pending Publication Date: 2022-07-15
成都安永鼎业生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, because the stability of BCG has an important impact on its effectiveness in clinical use, on the one hand, compared with the BCG preparations used for the prevention of tuberculosis, the concentration of BCG in the therapeutic BCG increases. Stability has brought higher challenges; on the other hand, although the BCG freeze-dried preparation for treatment has been reported, for example, the patent with the notification number CN111588859B discloses a drug containing dextran, sucrose, sodium glutamate , potassium chloride and trehalose as lyoprotectants and BCG as BCG D 2 BCG freeze-dried preparation prepared by PB302 strain
However, because the factors affecting the stability of BCG involve many aspects such as the strain itself, freeze-drying protective agent, and culture medium, and because there are differences in strains between recombinant BCG and BCG, there are differences in their physiological activities and physical and chemical properties. Therefore, the preparation of BCG frozen It is difficult to directly apply the freeze-drying protection agent or freeze-drying process of dry preparations to recombinant BCG

Method used

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  • Freeze-dried preparation of recombinant bacillus calmette guerin vaccine for treatment and preparation method and application of freeze-dried preparation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1. Preparation of the freeze-dried preparation of recombinant BCG for therapeutic use of the present invention

[0036] (1), the preparation of stabilizer: add the following mass fraction components to the water for injection: dextran 3.0%, sucrose 4.0%, sodium glutamate 1.0%, potassium chloride 1.0%, trehalose 4.0%, Tween 0.01%, then adjust the pH to 7.3-7.5 with 10% sodium hydroxide solution, and sterilize for 50 minutes at 116°C and 0.16MPa.

[0037] (2) Passaging of production strains

[0038] 1. Recovery of working seeds: The working seeds of recombinant BCG were inoculated with Roche medium and cultured at 38°C for 2-3 weeks.

[0039] 2. Passaging Sutong medium: Roche strains are inoculated in Sutong medium, and cultured at a suitable temperature (38° C.) and a suitable time (2-3 weeks).

[0040] (3) Preparation of stock solution

[0041] 1. Production of biofilm culture: use the production strain to pass the Sutong medium twice with an appropriate inoc...

Embodiment 2

[0052] Embodiment 2, the preparation of recombinant BCG freeze-dried preparation

[0053] Preparation of stabilizer: add the following components in water for injection: dextran 3.0%, sucrose 4.0%, sodium glutamate 2.0%, potassium chloride 1.0%, trehalose 4.0%, tween 0.01%, and then Adjust the pH to 7.3-7.5 with 10% sodium hydroxide solution, and sterilize for 50 minutes at 116° C. and 0.16 MPa.

[0054] The rest of the method steps are the same as in Example 1.

experiment example 1

[0064] Experimental example 1. Quality evaluation of the preparation of the present invention

[0065] 1. Experimental method

[0066] Performances such as the number of viable bacteria, physical appearance, moisture, and thermal stability before and after freeze-drying of the preparations of the Examples and Comparative Examples were evaluated and compared. The results are shown in Table 2.

[0067]

[0068] As can be seen from the results in the table, the reconstituted BCG freeze-dried preparations (Examples 1 and 2) with the stabilizer of the specific composition of the present invention are added. at 30×10 6 CFU / mg; the product has good thermal stability, and the viable cell rate is more than 50% compared with the storage condition at 4°C (that is, the viable cell count after freeze-drying); and it is conducive to the dispersion of recombinant BCG. The consistency of the number of viable bacteria in the latter sample is good, reflecting that the preparation of the pr...

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Abstract

The invention provides a recombinant bacillus calmette-guerin freeze-dried preparation for treatment. The recombinant bacillus calmette-guerin freeze-dried preparation is prepared from the following components in parts by weight: 12 parts of recombinant bacillus calmette-guerin, 1 to 3 parts of dextran 40, 3.5 to 4.5 parts of cane sugar, 1 to 3 parts of sodium glutamate, 0.8 to 1.3 parts of potassium chloride, 3.3 to 4.5 parts of trehalose and 0.005 to 0.015 part of tween-80. The freeze-dried preparation of the recombinant bacillus calmette-guerin vaccine for treatment provided by the invention is high in viable count, stable in property and easy to preserve, and has good application value as a bladder cancer treatment preparation.

Description

technical field [0001] The present invention relates to the field of biological products, in particular to a freeze-dried preparation of recombinant BCG for therapeutic use, and a preparation method and application thereof. Background technique [0002] BCG was obtained by French scientists Calmette and Guerin by culturing and isolating a strain of Mycobacterium bovis from cattle suffering from tuberculosis on a medium containing glycerol, potato and bovine bile, after 13 years and a total of 231 passages. Attenuated strain of Mycobacterium bovis. Bacille Calmette-Guérin (BCG) vaccine, which is a vaccine prepared from the BCG bacteria cultured by the BCG, can prevent tuberculosis. [0003] In addition to preventing tuberculosis, the researchers also discovered the therapeutic effect of BCG on tumors. BCG can be used as an immunomodulator, which can stimulate the body to enhance immune function and promote the release of immune factors to achieve anti-tumor effects, especia...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/04A61K9/19A61K47/02A61K47/18A61K47/26A61P35/00
CPCA61K39/04A61K9/19A61K47/26A61K47/183A61K47/02A61P35/00A61K2039/522A61K2039/57
Inventor 罗勇张艳杜方川王应春蔡祥苏爽徐志勇
Owner 成都安永鼎业生物技术有限公司
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