Translation of in vitro-transcribed RNA therapeutics

Abstract

In vitro transcribed, modified messenger RNAs (IVTmRNAs) have been used to vaccinate billions of individuals against the SARS-CoV-2 virus, and are currently being developed for many additional therapeutic applications. IVTmRNAs must be translated into proteins with therapeutic activity by the same cellular machinery that also translates native endogenous transcripts. However, different genesis pathways and routes of entry into target cells as well as the presence of modified nucleotides mean that the way in which IVTmRNAs engage with the translational machinery, and the efficiency with which they are being translated, differs from native mRNAs. This review summarises our current knowledge of commonalities and differences in translation between IVTmRNAs and cellular mRNAs, which is key for the development of future design strategies that can generate IVTmRNAs with improved activity in therapeutic applications.

A flexible, high-throughput system for studying mRNA translation kineticsin vitroandin cellulowith HiBit technology

“…An essential aspect of the development of mRNA-based therapeutics is the incorporation of modified nucleotides, which are chosen based on their effect on the translation efficiency of the mRNA and their ability to reduce immunogenic responses(74–76). Importantly, our data demonstrate that the choice of modification will depend on the encoded gene and therefore modified nucleotides should be tested on the therapeutic mRNA, not a reporter protein.…”

Emerging roles of RNA binding proteins in intervertebral disc degeneration and osteoarthritis

“…75 TTP is a zinc finger protein that contains tandem zinc-binding motifs, triggering ARE-containing mRNA destabilization. 76,77 Like HuR, TTP is a ubiquitously expressed protein that has been extensively investigated in cancer, 78 neurodegenerative disorders, 78 and diabetes. 79 Recent studies have shown that TTP is implicated in the pathogenesis of IDD and OA.…”

“…Although members of the ZFP36 family share a similar structure, they might mediate distinct cellular functions according to the cell or tissue type and the nature of the extracellular stimulus 75 . TTP is a zinc finger protein that contains tandem zinc‐binding motifs, triggering ARE‐containing mRNA destabilization 76,77 . Like HuR, TTP is a ubiquitously expressed protein that has been extensively investigated in cancer, 78 neurodegenerative disorders, 78 and diabetes 79 .…”

Synthetic Genes For Dynamic Regulation Of DNA‐Based Receptors

“…A ligand‐responsive cell‐free transcription device (termed ROSALIND) has been also developed for the detection of water contaminants [16] and more recently we have reported the use of transcriptional switches for the detection of specific antibodies [17–19] . Alternatively, cell‐free transcription/translation systems have been also used to develop synthetic genetically engineered circuits for different applications, [20–22] including the production of proteins and the development of nucleic acid constructs for gene therapy. In this context, Kim and Winfree have demonstrated genetic regulatory networks in which in vitro transcriptional circuits regulated only by T7 RNA polymerase and RNase H can be used to generate and degrade RNA signals [23] .…”