Albumin 3′untranslated region facilitates increased recombinant protein production from Chinese hamster ovary cells

Abstract

Chinese hamster ovary (CHO) cells are used for recombinant protein production in the pharmaceutical industry but there is a need to improve expression levels. In the present work experiments were carried out to test the effectiveness of different 3'untranslated regions (3'UTRs) in promoting production of a naturally secreted luciferase. Seamless cloning was used to produce expression vectors in which Gaussia princeps luciferase coding sequences were linked to the human albumin, immunoglobulin or chymotrypsinogen 3'UTR. Stably transfected CHO cells expressing these constructs were selected. Luciferase activity in the culture medium was increased 2-3-fold by replacing the endogenous 3'UTR with the albumin 3'UTR and decreased by replacement with immunoglobulin or chymotrypsinogen 3'UTR. Replacement of the native 3'UTR with the albumin 3'UTR led to a 10-fold increase in luciferase mRNA levels. Deletion analysis of the albumin 3'UTR showed that loss of nucleotides 1-50, which removed an AU-rich complex stem loop region, caused significant reductions in both luciferase protein expression and luciferase mRNA levels. The results suggest that recombinant protein expression and yield could be improved by the careful selection of appropriate 3'UTR sequences.

3.1 Cell Line Evolution and Engineering

“…This includes strong promoters, transcriptionenhancing elements, various methods for gene amplification, and procedures that allow selection for integration into highly transcribed regions [34,35]. Especially for large glycosylated molecules, such as antibodies, it was hypothesized that these successes created another bottleneck transcriptionally or even posttranslationally.…”

Application of the iPLUS non-coding sequence in improving biopharmaceuticals production

“…Certainly, cis -regulatory sequences present in the UTRs can be used to increase mRNA expression, as well as protein production, either in large scale as for recombinant protein production, or in the cells in the context of an immunization with an mRNA vaccine ( Gómez-Aguado et al, 2020 ). For example, the human albumin 3′UTR increases the production of secreted luciferase by 2-3 fold in CHO cells ( Pearson et al, 2012 ). The structure of the mRNAs in the vaccines mimics endogenous mRNAs ( Pardi et al, 2018 ) and their design has been optimized to elicit a strong and specific immunological response.…”