The Formidable Challenge of Controlling High Mannose-Type N-Glycans in Therapeutic mAbs

mAbs are currently the prime focus in biopharmaceutical drug development. Glycosylation is a critical quality attribute for mAbs because their clinical efficacy and safety are significantly affected by their glycosylation profile, which is generally heterogeneous, profoundly dependent on the manufacturing process, and thus prone to variations depending on cell culture conditions. As opposed to endogenous IgGs, marketed therapeutic mAbs contain higher levels of high mannose glycans, which can affect efficacy, pharmacokinetics, and stability. Current trends in biopharmaceutical manufacturing, such as process intensification and the rise of biosimilars, emphasize the need for a thorough understanding of the cellular processes, as well as the biotechnical process aspects that govern the production of high mannose-type N-glycans, in order to establish robust manufacturing processes. The clinical efficacy and safety of therapeutic monoclonal antibodies (mAbs) are significantly affected by their Fc-glycosylation profile. High mannose-type N-glycans (HM) affect efficacy (in terms of antibody-dependent cell cytotoxicity), pharmacokinetics, and stability. While in endogenous IgGs the HM levels are very low, they are significantly higher in marketed therapeutic mAbs. In order to meet the demands for late-phase clinical trial and market supply, process intensification is required. Since glycosylation profiles are sensitive to process variations and changes, controlling HM levels in robust manufacturing processes presents a formidable challenge and requires a thorough understanding of the cellular processes as well as the biotechnical aspects that govern the production of HM glycans. The clinical efficacy and safety of therapeutic monoclonal antibodies (mAbs) are significantly affected by their Fc-glycosylation profile. High mannose-type N-glycans (HM) affect efficacy (in terms of antibody-dependent cell cytotoxicity), pharmacokinetics, and stability. While in endogenous IgGs the HM levels are very low, they are significantly higher in marketed therapeutic mAbs. In order to meet the demands for late-phase clinical trial and market supply, process intensification is required. Since glycosylation profiles are sensitive to process variations and changes, controlling HM levels in robust manufacturing processes presents a formidable challenge and requires a thorough understanding of the cellular processes as well as the biotechnical aspects that govern the production of HM glycans. a mechanism of antibody-mediated target cell destruction where an immune cell lyses a target cell. a class of biopharmaceutical drugs composed of an antibody linked to a cytotoxic payload. a biopharmaceutical that is highly similar, but not identical, to a licensed medicinal product (‘originator’). a family of endocytic receptors that bind through carbohydrate-recognition domains (CRDs). Type I CLRs are calcium-dependent and have multiple CRDs [e.g., macrophage mannose receptor (MR)]. Type II CLRs contain a single CRD and can be either calcium-dependent, like Dectin 2 and DC-SIGN, or calcium-independent. a mechanism by which antibody-coated target cells, after binding to C1q, are destroyed through activation of the classical complement pathway. a continuous membrane system involved in the synthesis, folding, modification, quality control, and transport of proteins. also called CD16, binds to the Fc of IgG antibodies and activates ADCC. the fragment crystallizable region consists of a paired set of antibody heavy chain domains that bear the highly conserved N-glycosylation sites. It interacts with cell surface receptors responsible for the effector functions. a cell culture process with continuous or intermittent addition of nutrients to control the metabolic activity. The product remains in the bioreactor until the end of the run. Fc-fusion proteins are composed of the Fc domain of IgG genetically linked to another protein of interest. antibodies that consist entirely of human sequences. the attachment of an oligosaccharide structure, also referred to as glycan, to the asparagine (Asn) residue within the Asn–X–Ser/Thr consensus sequence. The N-glycosylation profile refers to the qualitative and quantitative composition of a mixture of structurally different glycans (complex-, hybrid-, and high mannose-type N-glycans). enzymes that catalyze the transfer of glycosyl groups from a nucleotide donor sugar to a substrate acceptor. intermediate glycoforms of the glycosylation pathway. The general term refers to glycans comprising between five and nine mannose residues. antibodies with sequences engineered to increase the resemblance with human antibodies, which, however, retain a low percentage of murine sequence regions. a modified fed-batch procedure in which a perfusion culture is utilized to inoculate the production bioreactor at a very high cell density in order to increase productivity. high mannose-type N-glycans containing five, six, seven, eight, or nine mannose residues, respectively. serum protein that, upon binding to bacterial carbohydrate, activates lectin-complement pathway. a series of unit operations required to produce the final product. For biotherapeutics, this includes the cell culturing, purification, formulation, as well as fill-&-finish processes. a group of molecules comprising nucleotides linked to sugars, which act as glycosyl donors in glycosylation reactions [e.g., uridine diphosphate–N-acetylglucosamine (UDP–GlcNAc) as the donor of N-acetyl glucosamine]. the concentration of osmotically active particles in a solution. It is an important parameter for intracellular ion homeostasis in cell culture. a perfusion cell culture process involves the constant feeding of fresh media and removal of spent media and product.