Introduction to principles and technical barriers of monoclonal antibodies

Posted by beauty33 on January 12th, 2020

Monoclonal antibody (Mab) has established leadership in therapeutic biologics. The establishment of a sound production platform is the key to seamlessly transforming the discovery of antibody drugs into clinical and market uses. The advent of biosimilars has made it possible to reduce drug costs and global production of biological products. Currently, monoclonal antibodies can easily achieve high expression in mammalian cell culture. These drivers have led to significant improvements in platform processes. In addition, in order to fulfill these needs, there are also some new trends in biopharmaceutical production processes, for example, the evaluation of different expression systems.

Principle of monoclonal antibody

Antibodies are mainly synthesized by B lymphocytes. Each B lymphocyte has a genetic gene that synthesizes an antibody. There are millions of different B-lymphocyte lines in animal spleens, and B-lymphocytes with different genetic genes synthesize different antibodies. When the body is stimulated by an antigen, many determinants on the antigen molecule activate each B cell with a different gene. Activated B cells divide and proliferate to form the descendants of the cells, that is, clones form multiple clones from the division and proliferation of many activated B cells and synthesize a variety of antibodies. If one cell can be selected to produce a specific antibody and cultured, a single cell can be obtained through division and proliferation to form a cell population, that is, a monoclonal. Monoclonal cells will synthesize a determinant antibody called a monoclonal antibody. B lymphocytes can produce antibodies, but they cannot divide indefinitely in vitro; while tumor cells can be passaged indefinitely in vitro, they cannot produce antibodies. The hybridoma cells obtained by fusing these two cells have the characteristics of two parental cells.

The technical barrier of producing monoclonal antibody

1. Transgenic mouse platform Vs library display

Monoclonal antibodies are classified into murine monoclonal antibodies, chimeric monoclonal antibodies, humanized monoclonal antibodies, and fully humanized monoclonal antibodies according to their species origin. With the continuous development of humanization of monoclonal antibodies, the full-human screening technology represented by phage display and transgenic mice, which both are important means of current full-human antibody screening. Of the two, about 70% of the market's mainstream human antibodies are obtained through screening of transgenic mice. At present, there are three major transgenic mouse platforms.

A new-generation method for obtaining monoclonal antibodies, which directly clones human B cells to obtain fully human antibodies. Thanks to the development of next-generation sequencing and other technologies, human B-cell single-cell cloning technology can achieve high-throughput screening and sequencing to obtain fully human antibodies. Despite various problems, the field of technology is changing rapidly.

2. The construction of cell lines and optimization of medium

At present, mammalian cell expression has become the most important technology for the production of biological drugs, especially antibody drugs. Compared with other eukaryotic cell lines such as yeast, mammalian cells have the advantage of expressing foreign proteins in that they can express target proteins containing complex disulfide bonds or post-translational modifications, and the proteins can be secreted into the culture medium. The degree of basalization is high and uniform, which can effectively improve the efficacy and reduce the side effects of the drug.

Chinese hamster ovary cells (CHO) are currently the most representative engineered cells in the field of recombinant protein production. They have the characteristics of efficient expansion and expression, and can reach 1-10g / L in fed culture. According to statistics, more than 60% of antibody drugs developed using the CHO cell line as an expression vector. CHO cells were isolated for the first time in 1957, and a variety of commercial cell lines have been domesticated, including CHO-K1, CHO-S, and so on.

3. Production of biopharmaceuticals

As well known, large-scale production is the key, while the process quality control is tough. The development process of monoclonal antibody drugs generally needs to go through the steps of construction of an engineered cell bank, development of a shake flask process, development of a pilot process, pilot scale-up, production and purification quality control, and preparations. At present, the expression levels of cell lines are quite different. The expression level of engineered cell lines for antibody production in the world can reach 20-70 PCD (pg / cell / day).