Although the messenger RNA (mRNA) and its roles in the cell were discovered more than half a century ago, it took more than four decades to consider its uses as biologics for human therapeutics. Nowadays, synthetic mRNA, produced in vitro by various enzymatic and non-enzymatic processes, is broadly used in vaccination, immunotherapeutics and even transient gene compensation.
Nevertheless, artificial mRNA synthesised in vitro and delivered exogenously suffers many limitations that hamper its potential. Firstly, even chemically modified exogenous RNA remains highly immunogenic due to its recognition by the Toll-Like Receptors on the external side of the endosomes and the cell membrane. Such innate immune activation induces an interferon response, which can be poorly tolerated and lead to serious adverse events. Secondly, mRNA is a fragile molecule, which is rapidly degraded in biological medium by abundant nucleases, making its uses in therapeutics difficult. Altogether, these limitations explain the difficulty in handling therapeutic mRNA, especially in cases of systemic administration for gene compensation. Therefore the development of novel alternatives is required.
To overcome these issues, the production of synthetic mRNA in the cell itself remains a desirable goal: it would prevent the stimulation of the innate immune response, as well as the rapid decay of mRNA in the biological medium. However, this goal was unachievable until recently due to the complexity of transcription in eukaryotic cells.
The synthesis of the RNA chain is performed by the nuclear RNA polymerase II and associated transcription factors – a huge protein complex of amazing complexity. Moreover, once produced, the eukaryotic mRNA chain undergoes various modifications involved in multiple functions of the eukaryotic cells. For instance, the capping at the 5’-end of the mRNA chain and polyadenylation at its 3’-end, which both occur at post-transcription, are instrumental for the recognition of the mRNA by the translation machinery and therefore, protein synthesis in the cell.
The C3P3 system is the first artificial system designed to synthesise artificial mRNA in the eukaryotic cells. This enzymatic system, now in its second generation, relies on a single monomeric enzyme that was developed by a complex synthetic biology process and transcribes a specific DNA template. Once expressed in the cytoplasm of the cell, the enzyme not only produces the mRNA chain, but also generates the key modifications required for the translation of mRNA.
The system has multiple applications – e.g. bioproduction, screening, transgenics and others – since it is able to produce any mRNA in virtually any eukaryotic cell, including for instance mammals, plants or yeast. Among these applications, human therapeutics appears particularly attractive due to the potential of mRNA.
At the 4Bio Synthetic Biology and Gene Editing Strand, we will present preliminary results of the uses of the C3P3 system for human therapeutics and more specifically, a novel approach to gene compensation. This therapeutic approach, that we call synthetic gene therapy, makes it possible to express or inhibit genes in a target tissue. The therapeutic potential of the synthetic gene therapy will be illustrated with the results of EUK-LPR, our first synthetic gene therapy that is pro-regenerative and has demonstrated efficacy in animal models.