Sodium chloride impacts glycosylation and N- and O-glycan site occupancy of an Fc-fusion protein.

Fc-fusion proteins are highly complex molecules, difficult to manufacture at scale. In this work, undesired proteoforms were detected during the manufacture of a therapeutic fusion protein produced in CHO cells. These species were characterized using gel electrophoresis, size exclusion chromatography and liquid chromatography-mass spectrometry leading to the identification of low molecular weight proteoforms presenting low N- and O-glycan site occupancy, as well as a low sialylation content. Upstream process parameters were investigated, and fusion protein quality was shown to be linked to the sodium chloride content of the medium. A mitigation strategy was developed to avoid formation of unwanted glyco-variants, resulting in an increased yield of highly glycosylated Fc-fusion protein. The effect of sodium chloride was shown to be independent of the osmolality increase and was hypothesized to be linked to a modulation of Golgi acidity, which is required for the correct localization and function of glycosyltransferases. Altogether, this study highlights the importance of the salt balance in cell culture media used to produce highly sialylated and occupied glycoproteins, helping to maximize the yield and increase robustness of processes aiming at producing biopharmaceutical complex therapeutic molecules.

Biological Durability of Wood-Polymer Composites-The Role of Moisture and Aging.

Knowledge about the resistance of wood-polymer composites (WPCs) to biological attack is of high importance for purpose-oriented use in outdoor applications. To gain this knowledge, uniform test methods are essential. EN 15534-1 (2018) provides a general framework, including the recommendation of applying a pre-weathering procedure before the biological laboratory tests. However, the procedure's manner is not specified, and its necessity assumes that a durability test without such pre-weathering will not produce the structural changes that occur during outdoor use. To verify this assumption, this study examined the influence of natural, ground-level pre-weathering on the material properties of different WPC variants, which were tested at intervals of six months in four durability tests under laboratory conditions in accordance with EN 15534-1 (2018). Weathering factors were calculated from determined characteristic values such as mass loss, and loss in moduli of elasticity (MOE) and rupture (MOR). The weathering factors based on mechanical properties tended to decrease with increasing weathering duration. The expected negative influence of pre-weathering on these material properties was thus not confirmed. The weathering factors based on mass loss were subject to high variation. No significant effect of pre-weathering on mass loss due to fungal attack became evident. Overall, the necessity of a pre-weathering step in biological durability tests shall be questioned based on the presented results.