Targeting early apoptotic genes in batch and fed-batch CHO cell cultures

• Published in: Biotechnology and bioengineering 2006-10, Vol.95 (3), p.350-361
• Main Authors: Wong, Danny Chee Furng, Wong, Kathy Tin Kam, Nissom, Peter Morin, Heng, Chew Kiat, Yap, Miranda Gek Sim
• Format: Article
• Language: English
• Subjects: Adaptor Proteins, Signal Transducing - genetics; ALG-2; Animals; Apoptosis; Apoptosis - genetics; Apoptosis - physiology; Apoptosis Regulatory Proteins - genetics; Apoptosis Regulatory Proteins - metabolism; Biological and medical sciences; Bioreactors; Biotechnology; Calcium-Binding Proteins - genetics; Carrier Proteins - genetics; Cell culture; Cell Cycle Proteins - genetics; CHO; CHO Cells; Cricetinae; Cricetulus; DNA-Binding Proteins - genetics; Down-Regulation - genetics; Endosomal Sorting Complexes Required for Transport; FADD; FAIM; Fas-Associated Death Domain Protein; fed-batch; Fundamental and applied biological sciences. Psychology; Gene Expression Profiling - methods; Genes; Interferon-gamma - metabolism; Oligonucleotide Array Sequence Analysis - methods; Recombinant Proteins - metabolism; REQUIEM
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.20871

Based on the transcriptional profiling of CHO cell culture using microarray, four key early apoptosis signaling genes, Fadd, Faim, Alg‐2, and Requiem, were identified and CHO GT (Gene Targeted) cell lines were developed by targeting these four genes. Two were CHO GTO cell lines overexpressing anti‐apoptotic genes, Faim and Fadd DN and two were CHO GTKD cell lines involving knockdown of Alg‐2 and Requiem which are pro‐apoptotic genes using small interfering RNA (siRNA) technology. Comparisons of these CHO GT cell lines with the parental cell line in batch culture (BC) and fed‐batch culture (FBC) were performed. Compared to parental cells, the CHO GT cell lines showed apoptosis resistance as they significantly delayed and/or suppressed initiator caspase‐8 and ‐9 and executioner caspase‐3 activities during culture. FBC of CHO GT cell lines reached significantly higher maximum viable cell densities (up to 9 × 106 cells/mL) compared with the parental cell line (5 × 106 cells/mL). The recombinant interferon gamma (IFN‐γ) yields were increased by up to 2.5‐fold. Furthermore, it was observed that the IFN‐γ was more highly sialylated. © 2006 Wiley Periodicals, Inc.