Genome-Edited CHO Cells for Non-Fucosylated Antibody Production

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Solution Overview

Problem

Existing antibody drugs face challenges due to inconsistent and changing glycotype components and contents, limiting their production stability and pharmaceutical properties, particularly in terms of ADCC activity and glycosylation heterogeneity.

Innovation Solution

Genome-edited CHO host cells are used to produce antibodies with a unique glycan profile by knocking out the FUT8 gene, resulting in non-fucosylated N-linked oligosaccharides and improved ADCC activity, achieved through the use of TALEN technology to edit the FUT8 gene and create stable, genome-edited CHO-BAT-KF cells.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If conventional CHO cells are used to produce antibodies, then the cells can maintain normal growth and protein expression, but the glycotype components and contents are inconsistent and changing, leading to high glycosylation heterogeneity

Engineering Contradiction:

Improveproduction stabilityVSAvoidglycotype consistency

Core Design Contradiction:

ReliabilityVSStability of the object's composition

Solution Approach 1:

The patent extracts and eliminates the source of glycosylation heterogeneity by knocking out the FUT8 gene in CHO cells, which is responsible for fucosylation. This removes the problematic enzymatic activity that causes inconsistent glycotype composition, thereby stabilizing the glycan profile and improving production reliability.

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

The patent changes the genetic parameter of the CHO cells by introducing genome edits (knockout of FUT8 gene) to alter the glycosylation profile. This parameter change transforms the cell line from producing heterogeneous glycotypes to producing uniform, non-fucosylated glycans, achieving both improved stability and consistent composition.

Inventive Principle:

Principle #35Parameter changes

2Reliability

If the FUT8 gene is knocked out in CHO cells to reduce fucose content, then ADCC activity is enhanced, but the cells require genome editing technology which increases process complexity

Engineering Contradiction:

ImproveADCC activityVSAvoidgenome editing process

Core Design Contradiction:

ReliabilityVSDevice complexity

Solution Approach 1:

The patent replaces traditional mechanical or chemical methods of glycosylation modification with genome editing technology (CRISPR/Cas9 or TALEN). This substitution allows for precise, heritable changes in the FUT8 gene that permanently alter the cell's glycosylation capability, achieving enhanced ADCC activity through a more efficient and controllable approach.

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

3Manufacturing precision

If exogenous sequences are introduced into CHO cells to modify glycosylation, then specific glycan profiles can be achieved, but the cells contain foreign DNA which may affect production stability

Engineering Contradiction:

Improveglycan profile controlVSAvoidproduction stability

Core Design Contradiction:

Manufacturing precisionVSReliability

Solution Approach 1:

The patent takes out the need for exogenous sequences by using knockout strategies that eliminate the FUT8 gene entirely. This approach achieves precise glycan profile control (non-fucosylated glycans) without introducing foreign DNA, thereby maintaining production stability and avoiding potential issues with foreign gene expression or cell line drift.

Inventive Principle:

Principle #2Taking out (Extraction)

Applied Scientific Principles

This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.

Function Achieved in This Case

The approach leads to antibodies with low N-glycosylation heterogeneity, enhanced ADCC effect, and improved pharmaceutical properties, including increased affinity to the FcγRIIIA receptor, thereby stabilizing antibody production and enhancing their therapeutic efficacy.

Implementation Method 1

Methodology Applied

Scientific EffectGene editing:

Implementation Method 2

The glycosylation forms of antibody drugs are mainly N-glycosylation, involving such monosaccharides as glucose, galactose, mannose, N-acetylglucosamine, N-acetylgalactosamine, fucose and sialic acid (NANA, NGNA). Based on the amount of terminal galactose, the two-branched or multi-branched double-antenna complex oligosaccharides connected to Fc fragment Asn297 of the antibody molecule can be divided into G0, G1 (1, 3), G1 (1, 6) and G2

Methodology Applied

Scientific EffectGlycosylation:

Data Source

PatentUS20230203169A1Recombinant antibody having unique glycan profile produced by CHO host cell with edited genome and preparation method thereof

Publication Date: 2023.06.29 BIO THERA SOLUTIONS LTD

AI summary

The present invention, in the field of bioengineering and biotechnology, relates to a method for preparing a recombinant antibody with a unique glycan profile produced by a genome-edited CHO host cell. Specifically, according to a method of the present invention, the TALEN technology is used to edit the FUT8 gene in CHO cells that have been adapted for serum-free suspension growth. The edited CHO host cells can produce recombinant antibodies with a unique glycan profile. The unique glycan profile can be characterized by non-fucosylated N-linked oligosaccharide chains of the antibodies, extremely low N-glycosylation heterogeneity and uniform carbohydrate chains. The antibody prepared by the method of the invention exhibit significantly increased ADCC and greater stability.