Bicistronic Expression Vector for Stable Antibody Production
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Solution Overview
Problem
Current methods for producing antibody therapeutics face challenges in achieving high-yield, stable gene expression in mammalian cells, particularly in co-expressing light and heavy chains along with amplification genes, leading to inefficient production of blockbuster antibodies.
Innovation Solution
A bicistronic expression vector is designed, incorporating an untranslated region (UTR) and an intron, which includes a first expression cassette for the antibody light chain gene and a second cassette with an internal ribosome entry site (IRES) for the heavy chain gene and amplification gene, enabling simultaneous expression as if they were a single gene.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional methods are used to co-express light chain, heavy chain, and amplification genes, then gene expression stability is improved, but production yield remains insufficient
Solution Approach 1:
The invention divides the expression system into two separate monocistronic expression vectors: one vector expresses the light chain gene combined with amplification gene (under CMV promoter), and another vector expresses the heavy chain gene (under EF1α promoter). This segmentation allows independent optimization of each expression cassette, achieving both stable gene expression through amplification and high production yield through efficient promoter-driven expression.
2Reliability
If multiple separate vectors are used for co-transfection, then expression stability is improved, but device complexity increases
Solution Approach 1:
The invention merges the light chain gene and amplification gene into a single monocistronic expression cassette under one promoter (CMV), eliminating the need for bicistronic constructs. This combining approach simplifies the vector design while maintaining stable co-expression through selective amplification, reducing the complexity of the overall expression system.
3Device complexity
If bicistronic vector is used to express multiple genes, then device complexity is reduced, but manufacturing precision deteriorates
Solution Approach 1:
The invention segments the expression system into separate monocistronic vectors with distinct promoters (CMV for light chain+amplification, EF1α for heavy chain). This segmentation provides precise control over each gene's expression level and timing, avoiding the regulatory complications of bicistronic designs while maintaining manageable vector complexity.
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
This approach allows for high-efficiency, stable expression of antibodies by co-expressing light and heavy chains along with amplification genes, significantly improving the production yield of therapeutic antibodies like bevacizumab, tocilizumab, and others.
Implementation Method 1
a second cassette with an internal ribosome entry site (IRES) for the heavy chain gene and amplification gene, enabling simultaneous expression as if they were a single gene
Implementation Method 2
A promoter is a site on DNA, in which RNA polymerase can bind and initiate mRNA synthesis due to the presence of binding sites for various transcription factors and TATA box
Implementation Method 3
a first expression cassette including 'promoter-untranslated region (UTR)-intron-antibody light chain gene-polyA'
Data Source
AI summary
The present invention relates to a bicistronic expression vector for antibody expression, an animal cell transfected with the expression vector, and a method for producing an antibody including culturing the animal cell, in which the expression vector includes a first expression cassette including ‘promoter-UTR-intron-antibody light chain gene-polyA’ and a second expression cassette including ‘promoter-UTR-intron-antibody heavy chain gene-internal ribosome entry site (IRES)-amplification gene-polyA’. An expression vector capable of expressing a desired antibody with high efficiency can be constructed using the bicistronic expression vector including an intron for antibody expression according to the present invention, and the expression vector can produce the antibody by culturing the transfected animal cell with stability and high efficiency.