The trailblazing success of the COVID-19 vaccine has accelerated the development of mRNA vaccines and therapies. Due to the wide applicability of this technology, there is now demand for rapid, high quality mRNA products within the scientific community. However, current approaches to manufacturing mRNA vaccines from plasmid DNA templates are laborious, error prone and can result in bacterial contaminants. We have developed a streamlined and reliable end-to-end method of manufacturing mRNA and LNP products.
Instead of using plasmid DNA as the initial raw material for mRNA production, our process employs a synthetic “gBlock” DNA template. The template is amplified using PCR instead of bacterial fermentation, significantly decreasing template production time, achieving purified product within a day rather than a week. Our synthetic DNA approach also effectively eliminates endotoxin contamination and avoids truncation of the polyA tail, a common error that arises in plasmid amplification. Furthermore, mRNA synthesised from synthetic templates has consistently higher cellular expression than plasmid-derived mRNA, likely due to the increased integrity and purity.
This mRNA production is easily scalable from 100ug for small in vitro studies to 10mg for preclinical studies. We have optimised encapsulation of mRNA in several different LNP formulations for delivery to in vivo systems. mRNA products undergo a series of analyses including automated gel electrophoresis, high performance liquid chromatography (HPLC), and endotoxin analysis to ensure specificity and purity of the final product. To improve the resolution and sensitivity of mRNA vaccine quality control, we developed a long-read (Oxford Nanopore Technologies (ONT)) sequencing test (VAX-seq) to measure key quality features of our DNA templates Our sequencing workflow allows complete coverage of the length of the mRNA sequence, including the polyA tail, allowing for a quantitative and sensitive measure of mRNA identity and integrity.
The stability and efficacy of our mRNA and LNP products have been established through extensive transfection and expression analysis in various mammalian cell lines. We have shown that our mRNA retains its high integrity following repeated freeze thaw cycling and long-term storage for up to a year.
Overall, our streamlined end-to-end manufacturing pipeline and state-of-the-art analytics allows us to provide high quality mRNA and LNP products quickly and reliably for preclinical evaluation. Through manufacturing hundreds of diverse mRNA vaccines and therapies we developed innovative manufacturing solutions, while enabling the unprecedented growth of the Australian mRNA sector.