The impact of expression vector position on transgene transcription allows for rational expression vector design in a targeted integration system.

Site-specific integration (SSI) technology has emerged as an effective approach by the pharmaceutical industry for the development of recombinant Chinese hamster ovary (CHO) cell lines. While SSI systems have been demonstrated to be effective for the development of CHO cell lines, they can be limiting in terms of both transgene expression and in the case of multi-specifics, the ability to generate the correct product of interest. To maximize the performance of Pfizer's dual SSI expression system for expressing monoclonal and multi-specific antibodies, we used a novel approach to investigate the positional effect of transgenes within expression vectors by engineering nucleotide polymorphisms (NP)s to use as biomarkers to track the level of transcript output from each expression vector position.