The clinical efficacy and safety of therapeutic monoclonal antibodies (mAbs) are greatly affected by their Fc-glycosylation profile. High mannose N-glycan (HM) affects efficacy (in terms of antibody-dependent cellular cytotoxicity), pharmacokinetics and stability. Although the HM level in endogenous IgG is very low, it is significantly higher in commercially available therapeutic mAbs. Current trends in biopharmaceuticals, such as process intensification and the rise of biosimilars, emphasize the need for a comprehensive understanding of cellular processes and biotechnological process aspects that control the production of high-mannose N-glycans in order to establish a sound manufacturing process.
In order to meet the demands for late-phase clinical trial and market supply, Creative Biolabs has developed an advanced GlycoDegree™ platform that allows controlling HM levels of recombinantly expressed therapeutic antibodies. Our glycoengineering experts have a thorough understanding of the cellular processes as well as the biotechnical aspects that govern the production of HM glycans. By using gene editing technology, our mannose-modified antibodies can be produced in mammalian cell lines with specific functional gene knockouts.
Recombinant mAbs are currently the largest and fastest growing class of biopharmaceuticals, usually expressed in mammalian cells. Currently, there are several trends driving the development and production of mAbs: process enhancements, the trend from humanized mAbs to fully human mAbs, the reformulation of subcutaneously delivered drugs, the rise of biosimilars, and the emergence of antibody-drug conjugates ( ADCs). In each of these areas, high mannose (HM) content is a key aspect.
Most mAbs are N-glycosylated at the heavy chain position Asn297 (EU numbering) in their Fc region. The two oligosaccharide chains (one for each heavy chain) in the Fc structure of an antibody are essential for mAb effector function, stability and pharmacokinetics (PK). When expressed in mammalian cells, glycosylation in the mAb Fc domain is heterogeneous in nature because it consists of a mixture of complex, hybrid, and HM-type N-glycans, which all contribute to the overall therapeutic profile of a given mAb.. In addition to modulating the classical interactions with type I and type II Fc receptors, Fc glycans can also interact with a variety of lectins. Therefore, glycan components are a key quality attribute (CQA), and effective control strategies are needed to ensure consistency between batches and processes to reduce treatment variability and meet regulatory requirements.
Fig.1 Types of N-glycans.
Different studies exploring high mannose Fc glycans have shown that mannose content affects antibody effector functions. HM glycans do not contain fucose, which is known to reduce the mAb interaction with Fcγ receptor III-A (FcγRIIIa), which can induce antibody-dependent cellular cytotoxicity (ADCC). Compared with fucosylated glycans, HM-containing mAbs have enhanced ADCC activity, although they do not reach the level of fucosylated and galactosylated complex glycans. HM glycans showed weak binding to C1q, which is responsible for initiating complement-dependent cytotoxicity. However, when exposed to multivalent surfaces (eg, antigen-binding IgG clusters), they may bind to serum mannose-binding lectin (MBL) 2, thereby activating the lectin complement pathway on mAb-targeted cells. In short, high-mannose Fc glycans have a positive effect on ADCC activity, but have a negative effect on the CDC activity of IgG molecules.
In order to support our mission to achieve the frontier advancement in life sciences, Creative Biolabs has developed many cutting-edge technologies in the field of glycan engineering based on our GlycoDegree™ platform. Through modern recombinant technology, a group of specific glycosyltransferase and/or glycosidase genes can be specifically knocked out or adjusted to produce glycoengineered antibodies with more humanization and homogenous glycosylation, better pharmacokinetics and enhanced pharmacological efficacy. Our scientists are fully confident to provide effective and economically feasible antibody glycoengineering solutions.
Please feel free to send us your project description and requirements, and you will get customized all-in-one services to meet your R&D or cGMP production needs.
Creative Biolabs provides luciferase-based ADCC assay. This Jurkat cell based assay is pioneered by Creative Biolabs, and the methodology is very well accepted by the field. See attached ADCC Reporter Assay Protocol for further details.
All products and services are for Research Use Only. Do Not use in humans.
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Creative Biolabs has established a team of customer support scientists ready to discuss ADCC/CDC optimization strategies, antibody production, bioinformatics analysis and other molecular biology/biotechnology issues.