title: Reciprocal Translocation Observed in End-of-Production Cells of a Commercial CHO-Based Process.
abstract: During the validation of an additional working cell bank derived from a validated master cell bank to support the commercial production continuum of a recombinant protein, we observed an unexpected chromosomal location of the gene of interest in some end-of-production cells. This event-identified by fluorescence in situ hybridization and multicolour chromosome painting as a reciprocal translocation involving a chromosome region containing the gene of interest with its integral coding and flanking sequences-was unique, occurred probably during or prior to multicolour chromosome painting establishment, and was transmitted to the descending generations. Cells bearing the translocation had a transient and process-independent selective advantage, which did not affect process performance and product quality. However, this first report of a translocation affecting the gene of interest location in Chinese Hamster Ovary cells used for producing a biotherapeutic indicates the importance of the demonstration of the integrity of the gene of interest in end-of-production cells.
  
  
  
  The expression of recombinant therapeutic proteins in mammalian cells depends on the establishment of a cell line with the gene of interest integrated in the host genome and stably expressed over time. Before being used for commercial production, cell lines are submitted to a qualification program in order to ensure their phenotypic and genotypic characteristics and the efficacy and safety of the product. During the production life cycle of a therapeutic protein, additional cells banks have to be validated after exhaustion of the current qualified cell bank in order to support the commercial production continuum of the recombinant protein. It is during the validation of an additional working cell bank derived from a validated master cell bank that we detected a different chromosome bearing the gene of interest in a portion of cells at the end of the upstream production phase. In our case, this event did not affect the process performance, the product quality, or its safety profile, but it highlights the need to characterize the integrity of the gene of interest in end-of-production cells when producing recombinant proteins for human use.
authors.affiliation: Merck Serono SA, Corsier-sur-Vevey, Biotech Process Sciences, Fenil-sur-Corsier, Switzerland; yolande.rouiller@merckgroup.com.
authors.forename: Yolande
authors.initials: Y
authors.lastname: Rouiller
authors.affiliation: Merck Serono SA, Corsier-sur-Vevey, Biotech Process Sciences, Fenil-sur-Corsier, Switzerland;
authors.forename: Beate
authors.initials: B
authors.lastname: Kleuser
authors.affiliation: Istituto Ricerche Biomediche Antoine Marxer RBM, Merck Group, MS-TDS-B2 Department, Colleretto Giacosa, Italy;
authors.forename: Emiliano
authors.initials: E
authors.lastname: Toso
authors.affiliation: Merck Serono SA, Global Research and Development, Coinsins, Switzerland;
authors.forename: Wolf
authors.initials: W
authors.lastname: Palinksy
authors.affiliation: Merck Serono S.p.A., MS-TDA-P Department, Guidonia Montecelio, Italy; and.
authors.forename: Mara
authors.initials: M
authors.lastname: Rossi
authors.affiliation: Istituto Ricerche Biomediche Antoine Marxer RBM, Merck Group, MS-TDS-B2 Department, Colleretto Giacosa, Italy;
authors.forename: Paola
authors.initials: P
authors.lastname: Rossatto
authors.affiliation: Bioclarma srl, Torino, Italy.
authors.forename: Davide
authors.initials: D
authors.lastname: Barberio
authors.affiliation: Merck Serono SA, Corsier-sur-Vevey, Biotech Process Sciences, Fenil-sur-Corsier, Switzerland;
authors.forename: Hervé
authors.initials: H
authors.lastname: Broly
mesh_terms: D000818:Animals; D001688:Biological Products / Q000378:metabolism*; D001702:Biopharmaceutics / Q000379:methods*; D019149:Bioreactors; D016466:CHO Cells; D020223:Chromosome Painting; D033481:Chromosomes, Mammalian; D003412:Cricetulus; D005786:Gene Expression Regulation; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D051379:Mice; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D019076:Transgenes; D014178:Translocation, Genetic
chemical_list: D001688:Biological Products; D011994:Recombinant Proteins; D013762:Tetrahydrofolate Dehydrogenase
keywords: CHO cells; Cell banks; Commercial process; Genetics; Recombinant protein; Validation