Beyond viability: Advancing CHO cell culture process strategies to modulate host cell protein levels.

Chinese Hamster Ovary (CHO) cells are widely utilized in bioprocessing industry for monoclonal antibody (mAb) production. In many instances, challenges persist in achieving sufficient clearance of Host Cell Proteins (HCPs) in the final drug substance. While purification strategies usually offer substantial HCP clearance, certain "problematic" HCPs, particularly lipases, continue to pose significant challenges.

Enhancing protein productivities in CHO cells through adenosine uptake modulation - Novel insights into cellular growth and productivity regulation.

Maximizing production potential of recombinant proteins such as monoclonal antibodies (mAbs) in Chinese Hamster Ovary (CHO) cells is a key enabler of reducing cost of goods of biologics. In this study, we explored various strategies to utilize adenosine mediated effects in biologics manufacturing processes. Results show that supplementation of adenosine increases specific productivity by up to two-fold while also arresting cell growth.

diaPASEF Enables High-Throughput Proteomic Analysis of Host Cell Proteins for Biopharmaceutical Process Development.

Monitoring and quantifying host cell proteins (HCPs) in biotherapeutic production processes is crucial to ensure product quality, stability, and safety. Liquid chromatography-mass spectrometry (LC-MS) analysis has emerged as an important tool for identifying and quantifying individual HCPs. However, LC-MS-based approaches face challenges due to the wide dynamic range between HCPs and the therapeutic protein as well as laborious sample preparation and long instrument time.

Contributions of Chinese hamster ovary cell derived extracellular vesicles and other cellular materials to hollow fiber filter fouling during perfusion manufacturing of monoclonal antibodies.

Hollow fiber filter fouling is a common issue plaguing perfusion production process for biologics therapeutics, but the nature of filter foulant has been elusive. Here we studied cell culture materials especially Chinese hamster ovary (CHO) cell-derived extracellular vesicles in perfusion process to determine their role in filter fouling. We found that the decrease of CHO-derived small extracellular vesicles (sEVs) with 50-200 nm in diameter in perfusion permeates always preceded the increase in transmembrane pressure (TMP) and subsequent decrease in product sieving, suggesting that sEVs might have been retained inside filters and contributed to filter fouling.

Comparison of process mass intensity (PMI) of continuous and batch manufacturing processes for biologics.

Biologics encompasses a wide variety of therapeutics including monoclonal antibodies, fusion proteins, and enzymes, among others. The biologics market is growing at a rapid pace and different manufacturing processes, including continuous manufacturing processes, are being increasingly adopted. There is a strong drive to assess the sustainability of such processes.

An innovative strategy to recycle permeate in biologics continuous manufacturing process to improve material efficiency and sustainability.

Intensified perfusion processes are an integral part of continuous manufacturing for biopharmaceuticals which enable agile operations and significant reduction in cost of goods. However, they require large volumes of media to support robust cell growth and maintain high productivity, posing substantial challenges to operations, logistics, and process sustainability. This study explores a novel strategy for reprocessing and reusing permeate from perfusion cultures for mAb production.

Streamlined life cycle assessment of single use technologies in biopharmaceutical manufacture.

The rapid growth of biologics as the preferred modality in several therapeutic areas has led to changes in the environmental profile of pharmaceutical manufacturing for some companies. The increased use of single use technologies (SUT) in biologics manufacturing has been accompanied by a greater public awareness of plastics waste, but the full life cycle environmental impacts of SUT have had limited study. Therefore, a segment of American Chemical Society Green Chemistry Institute Pharmaceutical Roundtable member companies undertook a streamlined cradle-to-gate life cycle assessment on a biological bulk drug substance (BDS) manufacturing process utilizing SUT at the 2000 L scale.

Understanding the effect of increased cell specific productivity on galactosylation of monoclonal antibodies produced using Chinese hamster ovary cells.

Achieving optimal productivity and desired product quality of the therapeutic monoclonal antibody (mAb) is one of the primary goals of process development. Across the various mAb programs at our company, we observed that increasing the specific productivity (q) results in a decrease in the % galactosylation (%Gal) level on the protein. In order to gain further insight into this correlation, cells were cultured under different process conditions such as pH or media osmolality or in the presence of supplements such as sodium butyrate.

Quantitative assessment of environmental impact of biologics manufacturing using process mass intensity analysis.

Process mass intensity (PMI) is a benchmarking metric to evaluate the efficiency of a manufacturing process, which is indicative of the environmental impact of the process. Although this metric is commonly applied for small molecule manufacturing processes, it is less commonly applied to biologics. In this study, an Excel based tool developed by the ACS GCI Pharmaceutical Roundtable was used to calculate PMI of different manufacturing processes for a monoclonal antibody (mAb).

Platelet GpIba binding to von Willebrand Factor under fluid shear:contributions of the D′D3-domain, A1-domain flanking peptide and O-linked glycans.

Background: Von Willebrand Factor (VWF) A1-domain binding to platelet receptor GpIbα is an important fluid-shear dependent interaction that regulates both soluble VWF binding to platelets, and platelet tethering onto immobilized VWF. We evaluated the roles of different structural elements at the N-terminus of the A1-domain in regulating shear dependent platelet binding. Specifically, the focus was on the VWF D'D3-domain, A1-domain N-terminal flanking peptide (NFP), and O-glycans on this peptide.

von Willebrand factor (VWF) propeptide binding to VWF D'D3 domain attenuates platelet activation and adhesion.

Noncovalent association between the von Willebrand factor (VWF) propeptide (VWFpp) and mature VWF aids N-terminal multimerization and protein compartmentalization in storage granules. This association is currently thought to dissipate after secretion into blood. In the present study, we examined this proposition by quantifying the affinity and kinetics of VWFpp binding to mature VWF using surface plasmon resonance and by developing novel anti-VWF D'D3 mAbs.

Epigenetic profile of the euchromatic region of human Y chromosome.

The genome of a multi-cellular organism acquires various functional capabilities in different cell types by means of distinct chromatin modifications and packaging states. Acquired during early development, the cell type-specific epigenotype is maintained by cellular memory mechanisms that involve epigenetic modifications. Here we present the epigenetic status of the euchromatic region of the human Y chromosome that has mostly been ignored in earlier whole genome epigenetic mapping studies.

Disulfide trapping of protein complexes on the yeast surface.

Protein complexes are common in nature and play important roles in biology, but studying the quaternary structure formation in vitro is challenging since it involves lengthy and expensive biochemical steps. There are frequent technical difficulties as well with the sensitivity and resolution of the assays. In this regard, a technique that can analyze protein-protein interactions in high throughput would be a useful experimental tool.