Chemistry Manufacturing and Controls Development, Industry Reflections on Manufacture, and Supply of Pandemic Therapies and Vaccines.

This publication provides some industry reflections on experiences from the Chemistry, Manufacturing, and Controls (CMC) development and manufacture and supply of vaccines and therapies in response to the COVID-19 pandemic. It integrates these experiences with the outcomes from the collaborative work between industry and regulators in recent years on innovative science- and risk-based CMC strategies to the development of new, high-quality products for unmet medical needs. The challenges for rapid development are discussed and various approaches to facilitate accelerated development and global supply are collated for consideration.

Mapping the molecular basis for growth related phenotypes in industrial producer CHO cell lines using differential proteomic analysis.

Background: The ability to achieve high peak viable cell density earlier in CHO cell culture and maintain an extended cell viability throughout the production process is highly desirable to increase recombinant protein yields, reduce host cell impurities for downstream processing and reduce the cost of goods. In this study we implemented label-free LC-MS/MS proteomic profiling of IgG4 producing CHO cell lines throughout the duration of the cell culture to identify differentially expressed (DE) proteins and intracellular pathways associated with the high peak viable cell density (VCD) and extended culture VCD phenotypes.

Results: We identified key pathways in DNA replication, mitotic cell cycle and evasion of p53 mediated apoptosis in high peak VCD clonally derived cell lines (CDCLs).

Differential expression of miRNAs and functional role of mir-200a in high and low productivity CHO cells expressing an Fc fusion protein.

Objectives: We used miRNA and proteomic profiling to understand intracellular pathways that contribute to high and low specific productivity (Qp) phenotypes in CHO clonally derived cell lines (CDCLs) from the same cell line generation project.

Results: Differentially expressed (DE) miRNAs were identified which are predicted to target several proteins associated with protein folding. MiR-200a was found to have a number of predicted targets associated with the unfolded protein response (UPR) which were shown to have decreased expression in high Qp CDCLs and have no detected change at the mRNA level.

Transcriptomic analysis of IgG4 Fc-fusion protein degradation in a panel of clonally-derived CHO cell lines using RNASeq.

In this study, we report an investigation of a panel of clonally-derived Chinese hamster ovary (CHO) cell lines exhibiting variability in the proportion of full-length IgG4 Fc-fusion protein produced. The recombinant protein was found to be degraded during cell culture into four shorter "clipped" species (three of the four cleavage sites occurred at arginine residues) and preliminary analyses suggested that a host cell enzyme was responsible for proteolysis. To identify the specific enzyme responsible, RNA sequencing was used to identify gene expression differences between the cell lines with a "high" and "low" clipping phenotype.

Clonal variation in productivity and proteolytic clipping of an Fc-fusion protein in CHO cells: Proteomic analysis suggests a role for defective protein folding and the UPR.

Product degradation, such as clipping, is a common quality issue in the production of Fc-fusion proteins from Chinese hamster ovary (CHO) cells. Degradation of proteins is mainly due to the action of either intracellular or extracellular host cell proteases. This study was carried out to understand more fundamentally the intracellular events that may play a role in determining why cell lines from the same cell line development project can vary with regards to the extent of Fc-fusion protein clipping.