The Hidden Complexity in the Development of Your mRNA Process

A significant number of challenges hide behind the misleading simplicity of mRNA. As the field is expanding to new therapeutic indications, a key question remains. How can we optimize costs and productivity without compromising product quality?

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While not new to the industry, the COVID-19 pandemic thrust mRNA therapeutics into the spotlight. The speed, efficacy, and simplified manufacturing  (i.e., no cell line generation, no virus used) of mRNA offer significant potential for vaccine platforms, protein-replacement therapy, and cancer immunotherapy^{[1, 2]}. However, despite their perceived simplicity, some significant hurdles remain.  

In this blog, we discuss some important challenges associated with mRNA production before exploring the modern solutions required to solve them. These trends are discussed in more detail in our recent webinar – watch now.

mRNA Production

Alongside the varied applications and significant clinical benefits of mRNA therapeutics, such as transient protein expression using the patient’s cells as a “bioreactor ^[1],” they also possess some operational benefits. For instance, mRNA production - especially the in vitro transcription (IVT) reaction - is fast (a few hours) and does not require long seeding and cell expansion procedures for large-scale production. Adding to the processing speed, the molecule design process is relatively “simple”, as the process sequence is known, and large quantities can be produced on a small scale with low CAPEX.

Despite clinical advantages and perceived production simplicity, mRNA processes have various complexities that drug developers must navigate to establish robust manufacturing processes.

mRNA Production Challenges

High Operating Costs
The production of the mRNA by IVT takes only a few hours which, compared to a cell-based process, is very fast and might lead to the idea that the cost of production is low. However, the COGs are relatively high for mRNA production processes. Most COGs are attributed to the high cost of materials, distinct from viral vector production processes.

Evaluating the cost of production is critical; mRNA competes with other modalities, and the choice should also take into consideration the cost per treatment. mRNA-based modalities may not be competitive for all applications (depending on dosage and doses), and cost-reduction strategies are currently in development.

Unique Features of the mRNA Molecule
New types of mRNA are emerging as a solution to challenges such as cost and stability (Figure 1). This diversity may lead to low yield and purity if the complete process from IVT to encapsulation is not optimized.  

Additionally, each new molecule is unique in size (size can vary by a factor of 10) and 3D structure, requiring the entire process to be reworked and optimized.

mRNA Production Processes
While the design process should be relatively simple (since the protein sequence is known), there are multiple choices when producing the mRNA molecule. Depending on the design, the purity and stability of the mRNA vary. Therefore, comprehensive screening and process development activities are essential to ensure that manufacturing is optimized. mRNA production processes are not yet fixed, with some variations in the purification step, capping, or poly-A tail addition, for example. They are also large molecules - up to 10 times the size of a monoclonal antibody (mAb) - requiring different separation or purification technologies than those used in a traditional mAb process.  

Deep process understanding is necessary to design and implement robust processes that yield high-quality products. 

mRNA molecules are also sensitive to degradation by enzymes such as RNase or oxidation. So while the process is enzymatic, caution should be taken to mitigate the risks of RNA degradation

Increasing Doses
Generally, higher and more frequent doses are required for mRNA-based gene therapies (especially protein-replacement therapy) compared to vaccines. Higher doses generally mean increasing the amount of mRNA into one dose while avoiding increasing the concentration of impurities (such as double-stranded and truncated RNA). This challenges purification and IVT processes and creates more regulatory hurdles. 

The number of patients is also likely to be lower for gene therapies compared to vaccines, for example. Therefore, the production scale should be well evaluated. 

How Can Drug Developers Overcome mRNA Production Challenges?

Explore New RNA Formats
As well as typical mRNA molecules, novel constructs with potential therapeutic benefits can also be employed. For example, self-amplifying RNA (saRNA), trans-amplifying RNA (taRNA), and circular RNA (circRNA) each have unique benefits, summarized in Figure 1.

^{Figure 1. Novel RNA types can help solve production challenges}

Quality by Design (QbD)
Applying the principles of QbD can accelerate development timelines and ensure quality is built into the process and product. This approach uses prior knowledge, experimental data, process understanding, and modeling to design a robust process.

In the webinar, Dr Zoltan Kis described how his team built a QbD framework to develop new mRNA products quickly. Using a patient-centric approach, they first define the quality target product profile (QTPP) and then determine the CQAs required to achieve this profile.  

They can then link the CQAs to the critical process parameters (CPPs), a complex relationship that requires modeling to understand. Finally, they define a design space to understand how combinations of process parameters would impact the yield and use this information to define a normal operating range.

High-Performing Purification Solutions
Chromatography-based purification offers selective and scalable purification. Scaled-down chromatography tools (such as the CIM^® Oligo dT18 0.05 mL Monolithic 96-well Plate) facilitate high-throughput screening to develop and optimize a chromatography process across different conditions or for different mRNAs.

Monolithic chromatography operates under low shear conditions and offers different chemistry to adapt to all RNA constructs, meaning they protect the integrity of large, sensitive mRNA molecules and can accommodate all kinds of processes and types of RNA produced.

Powerful Analytical Tools
Improving process productivity requires a deep process and product understanding. mRNA production (IVT) is fast. This creates an analytical bottleneck, which hinders the development and optimization of IVT reactions and purification processes. There is little analytics available specifically for mRNA that can perform quantification, finger printing, or impurity measurement. Automated, at-line analytics provide valuable process insights to accelerate process development at the point of use.

mRNA production processes lack the deep understanding the industry possesses for more mature production processes. However, the field is moving quickly; increased efficiencies and standardization continue to improve mRNA processes, bringing more therapies to clinical phases.

To learn more about overcoming challenges in mRNA production watch the webinar

References

^{1. Van Hoecke, L., & Roose, K. (2019, February 22). How mRNA therapeutics are entering the monoclonal antibody field. Journal of Translational Medicine. BioMed Central Ltd. https://doi.org/10.1186/s12967-019-1804-8}

^{2. Schlake, T., Thran, M., Fiedler, K., Heidenreich, R., Petsch, B., & Fotin-Mleczek, M. (2019, April 10). mRNA: A Novel Avenue to Antibody Therapy? Molecular Therapy. Cell Press. https://doi.org/10.1016/j.ymthe.2019.03.002}