Despite recent advances in improving titers for therapeutic proteins such as antibodies to the 10 g/L scale, these high yields can only be achieved in select mammalian hosts. Regardless of the host or product, strong promoters are required to obtain high levels of transgene expression. However, the regulatory elements (promoters) employed to drive this expression are rather limited in variety and are usually either viral in nature or screened empirically during vector design. This results in suboptimal synthetic parts that may be subject to silencing and/or limited potential. Thus, we wished to employ a more systematic approach to identify and design new synthetic promoters using endogenous elements. To do so, we established a workflow to design these elements by: (1) analyzing the transcriptomics profile of a specific cell line under a desired, representative cell culture condition, (2) identifying genetic motifs using bioinformatics that can be used to rationally construct synthetic promoters, and (3) evaluating the performance of these synthetic promoters using model proteins. During this design-build-test cycle, we investigated the underlying design rules for spacing and order of transcription factor binding sites in synthetic promoters. Overall, this approach of using an ‘omics-guided workflow for designing synthetic promoters enables the facile construction of high expression vectors for immediate use in current production hosts.

Transcriptomics-guided design of regulatory elements: A workflow to create synthetic promoters for mammalian hosts

251st National Meeting (2016)

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