Analysis on The Challenges and Bottlenecks of mRNA Manufacturing

The development and use of mRNA as a new class of drug modalities has shown great potential in the treatment of a variety of human diseases, including infectious diseases, cancer, and genetic diseases. Modified in vitro (IVT) mRNA has been shown to be feasible and safe for use in vaccinations, protein replacement therapy, antibody therapy, and more. Controlling the manufacturing and supply process is critical to successfully bring mRNA-based medicines.

During the Covid-19 pandemic, mRNA vaccines have come into the spotlight as a leading-edge technology used by many companies for vaccine development. In fact, an mRNA vaccine candidate is the first to enter phase I clinical trials. mRNA technology has several advantages that make it an attractive alternative to traditional vaccines and even DNA vaccines. Unlike attenuated or inactivated vaccines, mRNA is precise in that it only expresses the specific antigen and induces a directed immune response. In addition, it promotes humoral and cellular immune responses and induces the innate immune system.

Compared to DNA-based vaccines, mRNA is more effective because expression does not require nuclear entry, and is safer because there is almost zero chance of random genome integration. Furthermore, the expression of the encoded antigen is transient as the mRNA is rapidly degraded by cellular processes and no traces are found after 2-3 days. The flexibility of the mRNA vaccine platform also benefits manufacturing, as changes in the encoded antigen do not affect the physicochemical properties of the mRNA backbone, thus enabling standardization of production. Furthermore, since production is based on in vitro cell-free transcription reactions, safety concerns with cell-derived impurities and viral contaminants commonly found in other platforms are minimized.

Two forms of mRNA structure are being extensively studied for vaccine applications: regular or non-replicating mRNA and self-amplifying mRNA. In traditional mRNA formats, the antigen of choice is flanked only by the UTR region, 3' poly(A) tail, and 5' cap. This format has several advantages—the molecule is simple and small, and since no other proteins are encoded, it reduces the likelihood of unwanted immune responses. However, this mRNA expression is limited by its transient nature and higher mRNA doses may be required to achieve high expression.

Efforts have been made to overcome this bottleneck through the use of sequence optimization and formulation [34]. Self-amplifying mRNA (saRNA) is based on the addition of a viral replicase gene, enabling the mRNA to replicate itself. Typically, sequences from single-stranded RNA viruses are used, such as alphaviruses, flaviviruses, and picornaviruses. Following cytoplasmic delivery, this type of mRNA produces high levels of the target antigen. Despite the use of viral genes, no viral infectious particles or virus-like particles were observed during expression, reducing safety concerns. Evaluation of a saRNA vaccine for protection against H1N1/PR8 infection in a mouse model showed that a 64-fold lower dose was required to induce an immune response compared to conventional mRNA vaccine counterparts.

One of the most important advantages of mRNA over traditional vaccines is its relative simplicity to manufacture. To produce mRNA products with specific quality attributes, a series of manufacturing steps must be performed. Currently, a complete fabrication platform is still lacking, and various step combinations are possible. These can be divided into upstream processing, including the enzymatic generation of mRNA, and downstream processing, including the unit operations required to purify the mRNA product. These complement the LNP formulation and Fill-to-Finish steps.

Still, the choice of unit operations depends on the purpose. For example, lab-scale production typically involves a one-step synthesis reaction followed by nuclease digestion and precipitation. The exact unit operation used will have an impact on the manufacturing price and cost per dose. Ultimately, cost will be largely influenced by the amount of RNA per dose, the titer produced, and the scale of production used. The purchase price of 5' cap analogs and modified UTP seems to have an impact on cost.

The interest in this new class of vaccines stems from their flexibility, safety and precision compared to traditional methods. The growing number of clinical trials for cancer treatments and infectious diseases demonstrates the industry's growing interest in bringing these types of vaccines to market. mRNA vaccines are precise, safe, and flexible, and can be easily mass-produced for clinical-grade applications. These vaccines could be the answer in terms of manufacturing to quickly respond to epidemic outbreaks.