13.01.2022 duration: 3 min

Glycoengineered antibodies: pushing forward biotechnology

The importance of glycosylation patterns (sugar sequences) on proteins and their glycobiology have been intensely investigated in recent years, and the deeper understanding spawned novel applications that push biotechnology forward. As an example, the extensive glycosylation of the SARS-COV-2 spike protein appears to play a role in its immunogenicity and should facilitate the future development of optimized vaccines.

Innovative companies develop techniques to design and control glycosylation patterns of antibodies. A major breakthrough for medicine is the development of glycoengineered antibodies which show high promise to be more potent therapeutics with reduced propensity for adverse side effects.

In this article, we will present you information about glycoengineering, ADCC enhanced antibodies, highlight their potential and advantages, and show evitria’s service offerings in the field of glyco-engineering.

Protein Glycoengineering – advantages of glycoconjugates

Antibodies of the immunoglobulin G class (IgG), and human proteins in general, oftentimes naturally undergo post-translational modifications to be linked to sugars (carbohydrates) and sugar acids (e.g., sialic acid) at specific regions. This enzymatic conjugation process is termed N-glycosylation via glycosyltransferase and sialyltransferase and the resulting proteins are called glycoproteins.

In recent years, immunology research has shown that the type and sequence of sugars (oligosaccharides) in antibody N-glycans play a key role in binding to Fc receptors and hence the recruitment of natural killer (NK), macrophage and neutrophil immune cells. 

Engineering the glycosylation of recombinant antibodies to lack the sugar fucose enhances antibody-dependent cellular cytotoxicity (ADCC). These afucosylated antibodies are up to 10 – 50 times more potent in their effector functions, especially activating the ADCC pathway to fight cancer cells. Therefore, ADCC antibodies are considered very promising candidates for novel therapeutics in the oncology field and the first glycoengineered products are presently being introduced to the market.

Importance of glycoengineered antibodies

In humans, the N-glycosylation of antibodies shows some variability, leading to several glycoforms with a range of immune effector cell recruitment. Analogously, glycan structures of biotechnologically manufactured monoclonal antibodies (mAbs) depend on the production conditions and hence low heterogeneity of N-glycosylation is an important quality criterion of therapeutic antibodies and diagnostics reagents. As researchers of Merck’s biochem division​1​ recently pointed out, the presence of non-human glycosylation patterns from in vitro processes raises the risk of immunogenicity and related side effects.

Thus, glycoengineering and controlling the patterns of N-linked carbohydrate residues and sialylation in biopharmaceuticals is advantageous for biotechnology companies and patients alike, since it simplifies the characterization and comparison of product quality across batches and ensures highly potent treatments with reduced toxicity in vivo. Development and optimization of novel glycoengineering approaches are pushing the biotechnology field forward at an unprecedented speed.

Benefits of glycosylation – optimization of antibodies

The glycosylation pattern of antibodies enhances the stability of the three-dimensional structure, acts as a tag to show the immune system that the antibody belongs to the same individual, and modulates the activation of immune effector cells. These three factors are of major importance for patients in a therapeutic setting, as they determine the safety and activity of therapeutic antibodies.

For the manufacturer of antibodies, glycoengineering holds promise to produce homogeneous glycosylation patterns, which is a major determinant for batch quality. Additionally, homogeneous glycosylation would simplify quality assessment via mass spectrometry and reduce costs of quality control.

Afucosylated antibodies take the potency aspect to the next level, since afucosylation can increase the activation of the ADCC pathway up to 50-fold.

Interested in recombinant antibodies?

    ADCC enhanced antibodies

    The intentional synthesis of antibodies with lacking fucose moieties (afucosylation) allows the manufacture of therapeutic proteins with a significant increase in effector cell recruitment following the ADCC pathway. This is the major mechanism of action of therapeutic antibodies against cancer. Additionally, this pathway can be harnessed to fight bacterial and viral infections, as well as autoimmune diseases.

    Hence, afucosylated antibodies are more potent, specific and versatile therapeutics and can be used in smaller dosages to show the same effect as antibodies that contain fucose, thus lowering the risk of adverse side effects.

    Crucial to this approach is the selective biosynthesis of afucosylated antibodies. One proven implementation is the GlymaxX® platform. 

    Afucosylation with the GlymaxX® platform

    Glycoengineering offered by evitria is based on the GlymaxX ® technology for the modulation of fucosylation in mammalian cells. This unique biosynthetic approach is based on a deflecting enzyme which uses a metabolic fucose precursor as substrate and hence depletes fucose monosaccharides from glycosylation pathways. GlymaxX® allows the fine-tuning of fucose incorporation during protein glycosylation in antibodies, from complete lack to intermediate fucose levels to fully fucosylated glycopeptide variants.

    The GlymaxX® technology is based on a Chinese hamster ovary (CHO cells) expression system. These cell lines exhibit a stable genome and work reliably even in high titer manufacturing processes. A single cell line allows the production of fucose-free and fully fucosylated antibodies, depending on the culture conditions.

    Using the GlymaxX® technology, evitria offers the implementation of afucosylated antibody projects to be completed within 5 weeks.

    1. 1.
      Mastrangeli R, Palinsky W, Bierau H. Glycoengineered antibodies: towards the next-generation of immunotherapeutics. Glycobiology. Published online October 29, 2018:199-210. doi:10.1093/glycob/cwy092
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