The speed at which a recombinant protein product progresses into clinical trials is of vital importance for both small biotechnology
companies as well as the biopharma groups of large pharmaceutical companies. For mammalian cell lines, two major impacts on
the project timeline are the ability to quickly identify a product candidate and subsequently produce a high-expressing cell
line for that product. The advent of various computer-based protein design methodologies and antibody discovery technologies
for developing protein therapeutics has resulted in large numbers of protein or antibody variants that must be screened to
identify the best clinical candidate.
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For these variant candidates, proper screening requires milligram levels of protein production in mammalian cells for testing
candidate material. This testing is initially performed in vitro, but in many instances animal studies are required for identification
of the best potential clinical candidate. Animal studies can require multi-gram levels of the protein variants. Production
and screening of these variants and selection of a final candidate is a time-consuming process in most research groups, and
generally conflicts with what are now common aggressive timelines for moving products forward into the clinic. Typically,
the initial variants are produced from a transient expression system, and then after clinical candidate selection, the cell
line development process is restarted, a stable cell line is produced, and the final clinical candidate is master cell banked
for manufacturing. The GPEx (Gene Product Expression) technology allows protein variants to be screened as part of the manufacturing
cell line generation process. Milligram to multi-gram levels of protein are produced from an initial stable pool of clonal
cell lines for variant characterization and selection. Final clonal cell line selection from this pooled line either proceeds
in parallel with variant in vitro/in vivo screening or after clinical candidate selection has been completed. This method
of variant selection eliminates the need for a total restart in cell line development after the clinical candidate has been
identified.
The GPEx technology utilizes retrovectors to stably insert single copies of genes into dividing mammalian cells.1 Retrovectors deliver genes coded as RNA that, upon entry into the cell, are reverse-transcribed to DNA and integrated stably
into the host cell genome. Two enzymes, reverse transcriptase and integrase, provided in the vector particle, allow conversion
to DNA and gene insertion. These integrated genetic inserts are maintained through subsequent cell divisions as if they were
endogenous cellular genes.2
Characteristics of GPEx Technology
Figure 1. Recombinant Protein Cell Line Development Method.
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- The gene inserts target "active" regions of the cell genome allowing for high levels of protein production from the cell pools
and lines.3, 4, 5
- Antibiotic selection and gene amplification are not required, resulting in shorter clonal cell line development timelines.
- Stable cell pools producing milligram to gram quantities of variant proteins prior to selection of a high-expressing clone
allows for screening of protein variants and optimal selection of final clinical candidates.
- GPEx technology inserts each copy of the transgene at a different genomic location, producing very stable gene insertions
and stable expression from both pooled and clonal selected cell lines.
CHO Cell Line Development
