Molecular definition of predictive indicators of stable protein expression in recombinant NS0 myeloma cells.
We have generated a molecular description of the loci at which stability/instability of expression of a monoclonal antibody (MAb) (anti-CD38) occurs within the GS-NS0 expression system. Critically, these data show that, in the absence of changes to copy number for the recombinant gene sequences, all cell lines examined exhibit a progressive loss (instability) in expression of mRNA during prolonged culture. However, not all cell lines express instability at the level of MAb protein production. The molecular distinction between stable and unstable production at the protein level is a reflection of the cellular amount of recombinant mRNA encoding MAb. Our data indicate a threshold level, a putative saturation point for utilisation of mRNA in translational/secretory events, that defines stability or instability of protein production. Above this level of recombinant mRNA expression, cell lines are stable, whereas below this level cell lines will show instability of protein production. Our studies indicate that absolute levels of expression of recombinant mRNA encoding for MAb in the GS-NS0 expression system offer a potential predictive indicator for the selection of stable cell lines for scale-up. These studies identify molecular facets of host cell biology of generic interest for gene regulation and expression and define techniques and approaches for enhancement of recombinant protein expression and process development.
Barnes
Louise M
LM
2.205 School of Biological Sciences, University of Manchester, Stopford Building, Oxford Road, Manchester, M13 9PT, United Kingdom. louise.m.barness@man.ac.uk
Bentley
Catherine M
CM
Dickson
Alan J
AJ
D036541:ADP-ribosyl Cyclase / Q000235:genetics / Q000378:metabolism; D051997:ADP-ribosyl Cyclase 1; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000235:genetics*; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism; D015415:Biomarkers / Q000032:analysis*; D045744:Cell Line, Tumor; D020869:Gene Expression Profiling / Q000379:methods*; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D014644:Genetic Variation / Q000235:genetics; D008562:Membrane Glycoproteins; D051379:Mice; D009194:Myeloma Proteins / Q000235:genetics* / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000096:biosynthesis*
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D015415:Biomarkers; D008562:Membrane Glycoproteins; D009194:Myeloma Proteins; D011994:Recombinant Proteins; D036541:ADP-ribosyl Cyclase; C498155:Cd38 protein, mouse; D051997:ADP-ribosyl Cyclase 1