Post-translational modifications in the context of therapeutic proteins

Abstract

The majority of protein-based biopharmaceuticals approved or in clinical trials bear some form of post-translational modification (PTM), which can profoundly affect protein properties relevant to their therapeutic application. Whereas glycosylation represents the most common modification, additional PTMs, including carboxylation, hydroxylation, sulfation and amidation, are characteristic of some products. The relationship between structure and function is understood for many PTMs but remains incomplete for others, particularly in the case of complex PTMs, such as glycosylation. A better understanding of such structural-functional relationships will facilitate the development of second-generation products displaying a PTM profile engineered to optimize therapeutic usefulness.

Electrospray ionization quadrupole ion-mobility time-of-flight mass spectrometry as a tool to distinguish the lot-to-lot heterogeneity in N-glycosylation profile of the therapeutic monoclonal antibody trastuzumab

“…Asparagine at position 297 in the heavy chains is N-glycosylated and heterogeneity in this glycosylation profile is common for IgG antibodies [4,5]. The most common glycan structures for IgG possess zero, one, or two terminal galactose (G) residues with or without a fucose (F) and are defined as G0, G0F, G1F, and G2F [6,7].…”

Using cell engineering and omic tools for the improvement of cell culture processes

“…The functional biological activity of the glycoprotein is directly affected by glycosylation, including its trafficking and folding within the host cell (Scallon et al 2006;Walsh and Jefferis 2006;Willey 1999). Once secreted, solubility, aggregation, stability and immunogenicity of the protein may be affected by its glycosylation pattern (Kobata 1992;Sinclair and Elliott 2005;Willey 1999;Wyss and Wagner 1996).…”

“…N-glycosylation is initiated in the ER, and O-glycosylation can be initiated in either the ER or Golgi apparatus, but due to processing inconstancies glycans can have frequent structural heterogeneities (Patel et al 1992;Seth et al 2006a;Varki 1998). The most common therapeutic monoclonal antibody approved biopharmaceutical is of the immunoglobulin G (IgG) class (Walsh and Jefferis 2006). This glycoprotein has several isoforms which are cell line and clone dependent, plus the structure is influenced by the culture environment (Chee Furng et al 2005;Patel et al 1992;Raju et al 2000).…”

“…A silencing approach has also been used in CHO cells to increase the antibody effector function of ADCC by introducing siRNA targeting a1,6 fucosyltransferase (FUT8) mRNA (Mori et al 2004). Recently, one of the first biopharmaceuticals was approved for the market that included engineered modifications of glycosylation which extent its half life in human patients (Walsh and Jefferis 2006).…”

“…In 2006 it was reported that over 165 new biopharmaceutical entities received FDA approval (Walsh 2006). Mammalian cell lines command an effective monopoly towards production of complex therapeutic proteins that require post-translational modifications (Walsh 2006;Walsh and Jefferis 2006). This unique advantage outweighs the costs associated with mammalian cell culture, which are far greater in terms of development time and manufacturing when compared to bacterial culture.…”