Therapeutic Fc‐fusion proteins: Current analytical strategies

Abstract

Fc-Fusion proteins represent a successful class of biopharmaceutical products, with already 13 drugs approved in the European Union and United States as well as three biosimilar versions of etanercept. Fc-Fusion products combine tailored pharmacological properties of biological ligands, together with multiple functions of the fragment crystallizable domain of immunoglobulins. There is a great diversity in terms of possible biological ligands, including the extracellular domains of natural receptors, functionally active peptides, recombinant enzymes, and genetically engineered binding constructs acting as cytokine traps. Due to their highly diverse structures, the analytical characterization of Fc-Fusion proteins is far more complex than that of monoclonal antibodies and requires the use and development of additional product-specific methods over conventional generic/platform methods. This can be explained, for example, by the presence of numerous sialic acids, leading to high diversity in terms of iso-

A review of existing strategies for designing long-acting parenteral formulations: Focus on underlying mechanisms, and future perspectives

“…The strategy of Fc fusion provides the therapeutic protein a longer half-life by combining them with the Fc domain of an immunoglobulin G (IgG) 56 , 57 . Enbrel® (etanercept), was the first Fc fusion protein approved by FDA in 1998; it comprises of the extracellular region of tumor necrosis factor receptor 2 (TNFR2) and has been used in the treatment of rheumatoid arthritis 58 , 59 .…”

Expression of an immunocomplex consisting of Fc fragment fused with a consensus dengue envelope domain III in Saccharomyces cerevisiae

“…New classes of biopharmaceuticals include therapeutic Fc-fusion proteins, which improve half-life and efficiency. To date, thirteen such biopharmaceuticals have been approved in Europe and the United States (Duivelshof et al 2020 ). Interest in the Fc fragment has also been focused on the potential use of the immunocomplex to vaccinate against various infectious diseases and contribute the fight against cancers (Hioe et al 2009 ; Kumar et al 2013 ; DiLillo et al 2015 ; Kim et al 2015 ; Kim et al 2017 ).…”

Purification of Modified Therapeutic Proteins Available on the Market: An Analysis of Chromatography-Based Strategies

“…The general scheme of the purification process for mammalian system-expressed Fc-fusion proteins ( Figure 3 ) involves: i) a capture step, which aims to eliminate most HCP, DNA, and other cell culture-related impurities; ii) an intermediate step for viral inactivation; and iii) a polishing step, oriented to remove any remaining impurities and HMWA ( Shukla and Thömmes, 2010 ). It is important to highlight that the structure of Fc-fusion proteins may vary ( Duivelshof et al, 2021 ). The downstream processing for Fc-fusion proteins typically includes three chromatographic steps, often an affinity chromatography for capture, and two different chromatographic techniques (IEX, HIC or SEC) for polishing.…”

“…Numerous previous reviews describe the history, types of reactions, and purification strategies used for modified therapeutic proteins ( Pfister and Morbidelli, 2014 ; Jafari et al, 2017 ; Jiang et al, 2018 ; Ramos-de-la-Peña and Aguilar, 2020 ; Duivelshof et al, 2021 ). However, they focus mainly on modified therapeutic proteins that have not reached the market yet and are still under development.…”