Biomedical Engineering Reference
In-Depth Information
10.
Biotechnol Prog
. 2010 Sep;
26(5):1431-7. Rapid protein
production using CHO stable
transfection pools. Ye J, Alvin K,
Latif H, Hsu A, Parikh V,
Whitmer T, Tellers M, de la Cruz
Edmonds MC, Ly J, Salmon P,
Markusen JF. Merck & Co., Inc.,
Bioprocess Research and
Development, Rahway, NJ 07065.
During early preclinical development of therapeutic
proteins, representative materials are often required
for process development, such as for
pharmacokinetic/pharmacodynamic studies in
animals, formulation design, and analytical assay
development. To rapidly generate large amounts of
representative materials, transient transfection is
commonly used. Because of the typical low yields
with transient transfection, especially in CHO cells,
here we describe an alternative strategy using stable
transfection pool technology. Using stable
transfection pools, gram quantities of monoclonal
antibody (mAb) can be generated within 2 months
posttransfection. Expression levels for monoclonal
antibodies can be achieved ranging from 100 mg/L
to over 1000 mg/L. This methodology was
successfully scaled up to a 200 L scale using
disposable bioreactor technology for ease of rapid
implementation. When fluorescence-activated cell
sorting was implemented to enrich the transfection
pools for high producers, the productivity could be
improved by about threefold. We also found that an
optimal production time window exists to achieve
the highest yield because the transfection pools were
not stable and productivity generally decreased over
length in culture. The introduction of Universal
chromatin-opening elements into the expression
vectors led to significant productivity improvement.
The glycan distribution of the mAb product
generated from the stable transfection pools was
comparable to that from the clonal stable cell lines.
