From a medical point of view, some of the main benefits of developing mAb-based therapies are:

1. Specificity: Monoclonal antibodies are highly specific and can be designed to target a particular antigen or cell type, making them useful in many applications.
2. Low toxicity: Monoclonal antibodies are generally well-tolerated and have a low risk of side effects compared to traditional chemotherapies.
3. Long half-life: Monoclonal antibodies have a longer half-life (from few days to several weeks) in the body than traditional drugs, which can reduce the frequency of and improve patient compliance.

Recombinant antibodies are generated in vitro using an expression host (a cell used as a factory) that can yield high quantities. Mammalian systems, particularly Chinese Hamster Ovary (CHO) or human embryonic kidney (HEK) cell lines, are the preferred hosts for producing antibodies. Monoclonal antibodies can be expressed either transiently or stably by the host cells. Transient production is useful to obtain small amounts of antibody within short timelines. When large amounts are needed or when a therapeutic application is considered, a stable cell line is developed to ensure reproducible and GMP compliant production of the antibody.

Transient and stable expression are two methods for producing recombinant proteins, including monoclonal antibodies, using mammalian cells as expression hosts.

• Transient expression refers to a short-term process where plasmid DNA containing the gene of interest is introduced into the cells, and the protein is expressed for a limited period, typically a few days. Transient expression is suitable for small-scale production or quick screening of various expression constructs.
• Stable expression involves the integration of the gene of interest into the host cell genome, resulting in long-term expression of the protein. A stable cell line can be used for larger-scale production of recombinant proteins, including monoclonal antibodies, for therapeutic or commercial purposes. Developing a stable cell line requires more time and effort for the selection of and hyper-producing clone but is compliant with future scale-up and GMP manufacturing of the monoclonal antibody.

Several criteria should be considered, such as format of the molecule, final production scale, desired antibody properties, and the intended application. Here are the most commonly used hosts for production of antibody modalities:

• Mammalian cells: Chinese Hamster Ovary (CHO) or Human Embryonic Kidney (HEK) cell lines, are widely used and remain the expression system of choice to produce therapeutic mAbs. This expression system allows to perform post-translational modifications and produce correctly folded and functional full-length antibodies. However, this comes at a price. Cell line development is usually a lengthy process compared to other expression systems. Also, biomass production and protein purification are expensive due to the cost of raw materials and additional activities required to guarantee viral safety of the generated drug substance.
• Bacterial expression systems: Escherichia coli (E. coli) is commonly used for producing small fragments of antibodies that do not require post-translational modifications. Bacterial expression systems are cost-effective and easy to use. Bacterial strains can indeed be generated in a small amount of time, the bacteria grow fast (with a doubling time of 30 min compared to 20 for mammalianh for mammalian cells!), and the cultures can reach high cell density.
• Yeast expression systems: Komagataella phaffii (also known as Pichia pastoris) is often considered as an alternative to bacteria to produce therapeutic antibodies and fragments. Yeast expression systems offer advantages such as low cost, high yield, and fast production times. They can effectively secrete proteins into the culture supernatant, like mammalian cells, which greatly facilitates downstream purification. Strain generation is somewhat longer than in bacteria, though, since it requires screening for hyper-producers but the productivity is usually higher. To date, production of full-length antibodies in pastoris remains challenging. One reason is the peculiar glycosylation profile of proteins produced in this yeast which is often incompatible with therapeutic application of the antibodies.