The future of mRNA therapies depends on making production faster, safer, and more scalable, and synthetic DNA is emerging as a critical backbone for that transformation. mRNA is a powerful, programmable modality for vaccines, cancer immunotherapies, and gene therapies treatments, but its inherent instability means DNA templates are usually the starting point for large‑scale manufacturing. We interviewed Scientific Advisor Eric Crampon on the considerations that go into using synthetic DNA in producing such therapies. This article will touch on the difference between cell-free approaches to DNA synthesis and cell- or plasmid-based approaches.
Introduction: What are mRNA therapeutics?
Messenger RNA (mRNA) is the genetic blueprint that cells use to produce proteins. By harnessing this natural process, researchers can design mRNA molecules that instruct cells to make therapeutic medicines and enzymes, enabling a wide range of treatments and vaccines. The mRNA encodes viral vectors and other proteins that become the basis for downstream therapies or serve as the active ingredient. It has always been the scientists’ desire to improve the stability, immunogenicity, translation efficiency, and delivery systems to achieve efficient and safe delivery of mRNA.
Because mRNA is inherently unstable and prone to rapid degradation, DNA is the starting template for manufacturing mRNA at scale. Advances in synthetic DNA technologies and cell‑free production systems are making it faster and easier to generate high‑quality DNA templates, which supports rapid design–build–test cycles and initiatives such as the Coalition for Epidemic Preparedness’ (CEPI) 100‑day mission to deliver new vaccines in response to emerging pandemics. These same innovations also facilitate more complex applications, including organ‑targeted delivery and personalized treatments in oncology and cardiovascular disease.
Therapeutic modalities that rely on mRNA include:
- Cell and gene therapies1
- Personalized cancer vaccines encoding patient‑specific neoantigens2
- Prophylactic and therapeutic mRNA vaccines against infectious diseases3
Q: Could you start by describing where you see mRNA therapies and vaccines heading in the next few years?
A: From my perspective, nucleic acids—particularly DNA—have two main application areas: vaccines and advanced therapies. For vaccines, we have been working on using DNA as a starting material instead of viruses as seed for vaccine production, so all sites begin from the same DNA and avoid strain adaptation and discrepancies between manufacturing sites.
This approach aligns with CEPI’s push to provide infectious clones as DNA, which is safer to ship and improves biosafety. In industry, DNA is also critical for gene therapy and cell therapy, and having linear DNA is attractive because it simplifies the upstream process: you avoid plasmid linearization steps and make downstream processing easier.
Looking ahead, DNA is a very promising template for gene correction. RNA is transient and unstable, so expression typically lasts only hours, whereas DNA enables longer expression of the transgene. Linear DNA is also an excellent template for homologous recombination, the cell’s natural DNA repair mechanism, which is ideal for gene replacement. This could be a superior alternative to AAV-based delivery, which faces challenges with pre‑existing immunity and high production costs. If we can replace viral vectors with synthetic DNA, it would be a major advance for gene therapy.
Q: Can you tell me more about quality control? How would synthetic DNA help establish a reference standard, and how does that work in practice?
A: Quality control is very important for next‑generation sequencing (NGS). NGS platforms such as Oxford Nanopore and Illumina are widely used, but there is no widely adopted, defined reference material run with each batch to verify performance. Synthetic DNA standards could fill this gap, providing reference material for both sequence accuracy and potentially semi‑quantitative assessment.
Synthetic DNA can serve as a reference material by being spiked into every sample you run. Because you know its exact sequence, you can confirm that your NGS system reads it correctly and is performing as expected. If you also spike in a known quantity of this nucleic acid, you can tap into established spike‑in–based methods to obtain semi‑quantitative or quantitative readouts, using the known input to approximate the amount of virus or other analytes in your samples, in line with how other DNA and RNA spike‑in controls are already used on sequencing platforms.
Key points regarding reference standards for the industry
Standards in synthetic DNA are crucial because they let researchers turn next‑generation sequencing from a mostly qualitative tool into a reliable, semi‑quantitative measurement system. By adding synthetic DNA “spike‑in” controls of known sequence and concentration to every sample, labs can check that their sequencers and analysis pipelines are working properly, detect technical errors and bias, and normalize variability between runs. These internal standards also define performance metrics such as sensitivity, limit of detection, and linearity, which are essential for validating assays in regulated settings and for applications like low‑frequency variant detection, pathogen load estimation, and clinical decision‑making
Q: During COVID, mRNA vaccines became a major focus. What is the current status of mRNA vaccines, and how do you see that field developing now?
A: “The mRNA vaccine field is somewhat unsettled at the moment due to changing policies, but mRNA remains an ideal tool for pandemics and is central to pandemic preparedness. I expect essentially every major vaccine company to maintain RNA vaccine programs going forward.
Beyond infectious disease, mRNA vaccines are highly promising for cancer vaccines, which is a different application space where there is still clear value. mRNA vaccines will not replace all other modalities, but they will be part of most companies’ portfolios. For example, Pfizer still has strong mRNA vaccine programs for both flu and COVID, and for seasonal markets mRNA is excellent because it can be updated quickly and the manufacturing process is far easier to adjust than virus‑based production. ”
The Role of Synthetic DNA in mRNA Vaccine Manufacturing
Modern mRNA vaccines are typically manufactured using an in vitro transcription (IVT) process, where a DNA template encoding the antigen is transcribed into mRNA in a cell‑free reaction, then chemically capped, polyadenylated, purified, and finally formulated into lipid nanoparticles for delivery. The DNA template can be either a linear fragment or a plasmid that has been linearized. This choice directly affects the following:
- Upstream complexity
- Impurity profiles, e.g., antibiotic resistant sequences
- CMC burden: the amount of analytical work and regulatory documentation required to characterize, control, and justify the template and its impurities3
The above issues are abated by cell‑free linear DNA. Because mRNA platforms are now central to pandemic preparedness efforts (for example CEPI’s 100‑Days Mission), optimizing this template and IVT step has become a major focus for vaccine developers and manufacturing partners.
What Are Personalized mRNA Vaccines?
Personalized mRNA cancer vaccines begin by sequencing an individual patient’s tumor to identify neoantigens, then using computational methods to select neoantigens predicted to be immunogenic. These neoantigens are encoded in a patient‑specific mRNA construct that is manufactured by first generating a matching DNA template and then performing in vitro transcription, making DNA design and synthesis speed a central constraint in the manufacturing workflow. Cell‑free, linear synthetic DNA templates mitigate this bottleneck by eliminating plasmid cloning, reducing contamination and the CMC burden, and enabling shorter design‑to‑dose timelines, which is essential for clinically meaningful deployment of individualized vaccines.
Q: Cancer mRNA vaccines are highly personalized. How do you see synthetic DNA enabling such personalized therapies? What becomes possible that may not be feasible today?
A: Personalized mRNA vaccines require rapid adjustment to each patient’s tumor mutations, and that is where fast DNA synthesis is critical. You do not necessarily need large quantities of DNA; for personalized mRNA vaccines, milligram‑scale DNA can be sufficient. Linear DNA simplifies the process because you can go directly into RNA synthesis without plasmid cloning and preparation.
Some genes are difficult or unsafe to express in Escherichia coli, which can make plasmid production challenging. One advantage of Ribbon’s cell‑free linear DNA technology is that you avoid cloning in E. coli, allowing you to produce long or problematic DNA sequences that might otherwise be difficult to handle in bacterial systems. This is relevant not only for cancer vaccines but also for concepts like oncolytic viruses, where you may want to rapidly generate and adjust viral constructs for personalized use.
Q: What is the current status quo in terms of plasmid‑based bacterial production versus the cell‑free approach that Ribbon Bio enables? How does the cell‑free approach change the manufacturing of mRNA vaccines and other mRNA‑based therapies?
A: The key advantage of cell‑free production is the simplification of the CMC (Chemistry, Manufacturing, and Controls) package. Because you are not using living cells like E. coli, the process keeps endotoxin levels low, and you avoid many of the tests required to show you are below specified endotoxin limits in each batch.
When you use plasmid DNA as a template for in vitro transcription, you must also demonstrate that residual antibiotic resistance genes—such as kanamycin resistance used in plasmid selection—are below regulatory thresholds. Some groups try to work around this by using minicircles or other designs to reduce antibiotic sequences, but these add complexity. In a cell‑free system without plasmids and antibiotics, you eliminate these issues by design, which simplifies CMC requirements and reduces QC costs.
Q: In cell and gene therapy, people often say “the process is the product.” Beyond CMC simplification, what process considerations change if you manufacture gene or cell therapies using a cell‑free approach instead of a bacterial plasmid‑based process?
A: Because you can directly produce linear DNA, you remove the need for plasmid linearization, which is normally the first step before in vitro transcription. With linear DNA templates, you may also be able to replace expensive endonuclease treatments—for example, Benzonase or DNase I, which cut both linear and supercoiled DNA—with exonucleases that specifically digest unwanted DNA ends, since all your DNA is in solution and accessible.
Synthetic DNA and mRNA in Cell and Gene Therapies
In cell and gene therapies, synthetic DNA is increasingly central both as a template for in vitro–transcribed RNA (for example, for engineered cell products) and as a repair template for genome editing, where linear DNA supports homology‑directed repair and other recombination‑based mechanisms. Cell‑free, linear DNA manufacturing reduces reliance on plasmids and bacterial fermentation, thereby avoiding endotoxin and antibiotic‑resistance sequences, simplifying CMC, and shortening development timelines for complex modalities such as CAR‑T, in vivo gene editing, and engineered cell therapies. As manufacturing paradigms shift toward platform processes for mRNA and gene‑modified cells, high‑quality linear synthetic DNA becomes a key enabling technology, helping break the DNA bottleneck and aligning with the regulatory view that in these products, the process effectively defines the product
Conclusion
Taken together, advances in cell‑free, linear synthetic DNA directly reinforce the future of mRNA vaccines, personalized mRNA cancer vaccines, and cell and gene therapies by providing faster, cleaner, and more scalable templates that simplify CMC and support truly responsive, patient‑tailored manufacturing.
References:
1. Schürmann, Paul J., et al. Therapeutic application of mrna for genetic diseases. WIREs Nanomedicine and Nanobiotechnology, vol. 17, no. 3, May 2025, https://doi.org/10.1002/wnan.70019.
2. Sahin, U., Schmidt, M., Derhovanessian, E. et al. Individualized mRNA vaccines evoke durable T cell immunity in adjuvant TNBC. Nature 651, 1088–1096 (2026). https://doi.org/10.1038/s41586-025-10004-2
3. Guideline on the Quality Aspects of mRNA Vaccines | EMA. European Medicines Agency, 27 Mar. 2025, www.ema.europa.eu/en/documents/scientific-guideline/draft-guideline-quality-aspects-mrna-vaccines_en.pdf.
