(185d) Secretory Pathway Analysis of High Productivity in Mammalian Cell Culture through Genomics and Proteomics

High productivity in mammalian cell culture is achieved via a high integral viable cell density (i.e., more cells), high specific productivity (more antibody secreted per cell), or as a result of a balance between the two. The molecular portrait of what goes on inside cells with a high productivity phenotype is not completely clear. It is for this reason that we analyzed the gene and protein expression profiles of 2 different Chinese Hamster Ovary (CHO) cell lines secreting 2 different clinical stage monoclonal antibodies with particular emphasis placed on those genes/proteins involved in the secretory pathway. Each cell line was cultured under conditions in laboratory-scale bioreactors that facilitated at least a 2 fold difference in final titer levels. One cell line achieved high titers due to a high specific productivity, and the other achieved a high titer due to a high integral viable cell density and specific productivity. Through use of a CHO oligonucleotide microarray we were able to probe our samples against CHO sequences. Numerous genes with a direct or indirect role in the secretory pathway were identified. Samples from identical bioreactor cultures were also fractionated to purify ER and Golgi organelles to enrich those proteins most likely associated with the secretory pathway. Verification of organelle extraction was done via TEM, protein separation was achieved through 2-dimensional gel electrophoresis, and protein identification was achieved through nano-flow ESI Q-TOF MS/MS analysis. The information gathered through these efforts will support our ultimate goal of better understanding a high producer phenotype.