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Serum-free culture medium for protecting disulfide bond integrity of antibody in animal cell culture process

A serum-free culture medium and culture medium technology, applied in cell culture medium, cell culture active agent, animal cells, etc., can solve the problems of increasing the cost of purification stage, depleting antibody drug titer, and not yet protecting antibody disulfide bonds.

Active Publication Date: 2018-10-23
上海倍谙基生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The breakage of the disulfide bond will lead to the change or loss of the spatial structure of the antibody, which will directly affect the effectiveness and safety of the product
In summary, the addition of components and concentrations in serum-free medium will mediate the enzyme-catalyzed cleavage based on thioredoxin system and the non-enzyme-catalyzed cleavage process mediated by redox substances, thereby destroying the advanced structure of antibodies, resulting in antibody drug Questions about safety and efficacy arise
[0005] Most of the currently proposed solutions for protecting the integrity of antibody disulfide bonds are limited to the solid-liquid separation and purification stages after the animal cell culture process. The disadvantages are that on the one hand, the cost of the purification stage is increased, and on the other hand, the titer of antibody drugs is lost.
At present, there is little understanding of the animal cell culture process, especially the design of serum-free medium. At this stage, there is no report on the serum-free medium that protects the integrity of antibody disulfide bonds during animal cell culture.

Method used

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  • Serum-free culture medium for protecting disulfide bond integrity of antibody in animal cell culture process
  • Serum-free culture medium for protecting disulfide bond integrity of antibody in animal cell culture process
  • Serum-free culture medium for protecting disulfide bond integrity of antibody in animal cell culture process

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0182] A serum-free medium that is suitable for CHO cell culture and effectively protects the integrity of antibody disulfide bonds and reduces fragmentation content during CHO cell culture, including reducing substances, neutral substances and oxidizing substances. The contents of each component are as follows , unit mg / L:

[0183] Sodium Selenite 0.1,

[0184] Ferrous sulfate heptahydrate 0.5,

[0185] L-Arginine Hydrochloride 40,

[0186] L-cysteine ​​hydrochloride 60,

[0187] L-Histidine Hydrochloride 30,

[0188] L-Lysine Hydrochloride 90,

[0189] L-serine 30,

[0190] L-threonine 40,

[0191]L-Tryptophan 20,

[0192] L-proline 70,

[0193] L-tyrosine sodium salt 80

[0194] Ascorbic acid 20,

[0195] Riboflavin 0.5,

[0196] Retinol 5.0,

[0197] Tocopherol 5.0,

[0198] Glucose 2500,

[0199] Dithiothreitol 120,

[0200] Alpha Lipoic Acid 2.5,

[0201] Reduced Glutathione 100,

[0202] Albumin 0.3,

[0203] Calcium Chloride Anhydrous 60,

[0204] Po...

Embodiment 2

[0252] A serum-free medium that is suitable for CHO cell culture and effectively protects the integrity of antibody disulfide bonds and reduces fragmentation content during CHO cell culture, including reducing substances, neutral substances and oxidizing substances. The contents of each component are as follows , unit mg / L:

[0253] Sodium Selenite 10,

[0254] Ferrous Sulfate Heptahydrate 10,

[0255] L-Arginine Hydrochloride 180,

[0256] L-cysteine ​​hydrochloride 300,

[0257] L-Histidine Hydrochloride 230,

[0258] L-Lysine Hydrochloride 450,

[0259] L-Serine 300,

[0260] L-Threonine 400,

[0261] L-Tryptophan 350,

[0262] L-Proline 400,

[0263] L-tyrosine sodium salt 450

[0264] Ascorbic Acid 120,

[0265] Riboflavin 8.0,

[0266] Retinol 20,

[0267] Tocopherol 20,

[0268] Glucose 9000,

[0269] Dithiothreitol 300,

[0270] Alpha Lipoic Acid 20,

[0271] Reduced Glutathione 1200,

[0272] Albumin 10,

[0273] Calcium Chloride Anhydrous 450,

[0...

Embodiment 3

[0322] A serum-free medium that is suitable for CHO cell culture and effectively protects the integrity of antibody disulfide bonds and reduces fragmentation content during CHO cell culture, including reducing substances, neutral substances and oxidizing substances. The contents of each component are as follows , unit mg / L:

[0323] Sodium Selenite 5,

[0324] Ferrous Sulfate Heptahydrate 5,

[0325] L-Arginine HCl 100,

[0326] L-cysteine ​​hydrochloride 150,

[0327] L-Histidine Hydrochloride 140,

[0328] L-Lysine Hydrochloride 250,

[0329] L-serine 150,

[0330] L-Threonine 200,

[0331] L-Tryptophan 180,

[0332] L-Proline 250,

[0333] L-Tyrosine Sodium Salt 300,

[0334] Ascorbic acid 70,

[0335] Riboflavin 4.5,

[0336] Retinol 15,

[0337] Tocopherol 16,

[0338] Glucose 5500,

[0339] Dithiothreitol 220,

[0340] lipoic acid 15,

[0341] Reduced Glutathione 600,

[0342] Albumin 5,

[0343] Calcium Chloride Anhydrous 240,

[0344] Potassium chlor...

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Abstract

The present invention relates to a serum-free culture medium for protecting the disulfide bond integrity of an antibody in an animal cell culture process, wherein specifically the culture medium comprises a reducing substance, a neutral substance and an oxidizing substance. According to the present invention, the culture medium can support the rapid growth of cells and the high expression of antibodies in animal cell culture processes; and by designing the type and the concentration of reducing substance in the culture medium, the disulfide bond integrity of antibodies can be well protected, and the content of the finally obtained complete antibody is higher than 95%, such that the effectiveness and the safety of antibody production based on animal cell culture processes can be ensured.

Description

technical field [0001] The invention relates to the field of cell culture, and more specifically, the invention relates to a serum-free medium for protecting the integrity of antibody disulfide bonds during animal cell culture. Background technique [0002] In the global biopharmaceutical market, antibody drugs are the most important category. This type of drug has the characteristics of strong targeting, high specificity and low side effects. It represents the latest development direction in the field of drug therapy, especially in the field of anti-tumor The field of treatment of autoimmune system deficiency has unlimited market prospects. As of 2016, 61 antibody drugs have been launched in the world, with annual sales of 90 billion US dollars, and the average annual compound growth rate in the past 10 years has reached 31.65%. These data reflect the broad market prospects and rapid development momentum of antibody drugs. [0003] Animal cell culture technology is one of...

Claims

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Application Information

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IPC IPC(8): C12N5/07
CPCC12N5/0037C12N2500/12C12N2500/14C12N2500/16C12N2500/24C12N2500/30C12N2500/32C12N2500/34C12N2500/35C12N2500/38C12N2500/84C12N2501/33
Inventor 谭文松范里刘旭平赵亮张理想
Owner 上海倍谙基生物科技有限公司
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