Characterization of a cathepsin D protease from CHO cell-free medium and mitigation of its impact on the stability of a recombinant therapeutic protein.

During purification process development of a recombinant therapeutic protein, an endoproteolytic activity endogenous to the Chinese hamster ovary (CHO) cells and leading to degradation at particular hydrophobic amino acid residues (e.g., Phe and Trp) was observed when processing at acidic pH.

Alkaline cation-exchange chromatography for the reduction of aggregate and a mis-formed disulfide variant in a bispecific antibody purification process.

During the purification development of a bispecific antibody, cation-exchange chromatography was screened for its ability to separate a prominently expressed (>12%) mis-formed disulfide bond variant, termed MAb-diabody, and aggregate from the product of interest. The influence of pH, product load (g of product per liter of resin) and linear velocity on the separations were evaluated for the strong cation-exchange resins SP Sepharose HP and POROS(®) HS50. Cation-exchange chromatography is commonly operated distant to the isoelectric point of a molecule, generally leading to acidic conditions for antibody purification.

Evaluation of microfluidics reactor technology on the kinetics of virus inactivation.

Mammalian cell lines constitute an important part in the manufacture of therapeutic proteins. However, their susceptibility to virus contamination is a potential risk to patient safety and productivity, and has led to the development of a repertoire of virus inactivation techniques. From a process development viewpoint, the challenge is to demonstrate the required log reduction in virus content without a significant loss in product titer or quality.

Evaluation of viral removal by nanofiltration using real-time quantitative polymerase chain reaction.

Nanofiltration is commonly introduced into purification processes of biologics produced in mammalian cells to serve as a designated step for removal of potential exogenous viral contaminants and endogenous retrovirus-like particles. The LRV (log reduction value) achieved by nanofiltration is often determined by cell-based infectivity assay, which is time-consuming and labour-intensive. We have explored the possibility of employing QPCR (quantitative PCR) to evaluate LRV achieved by nanofiltration in scaled-down studies using two model viruses, namely xenotropic murine leukemia virus and murine minute virus.

Selective enhancement of multipotential hematopoietic progenitors in vitro and in vivo by IL-20.

In a search for novel growth factors, we discovered that human interleukin-20 (IL-20) enhanced colony formation by CD34+ multipotential progenitors. IL-20 had no effect on erythroid, granulocyte-macrophage, or megakaryocyte progenitors. IL-20 transgenic mice increased the numbers and cell cycling of multipotential but not other progenitors.