mRNA therapies: current landscape and future possibilities

mRNA Therapies and Vaccines

The urgency to tackle the SARS-CoV-2 pandemic brought to the forefront a novel platform of vaccines- messenger RNAs (mRNA). While it may appear that the development of mRNA-based therapies is recent, for more than two decades researchers have been actively working to optimize the stability and delivery of mRNAs for treating various diseases.

The steppingstone in the history of mRNA therapy research was a seminal experiment by Robert Malone from the Salk Institute, in 1989 who showed that synthetic mRNA strands mixed with lipid particles could be used to transfect human cells to express a protein of interest, and serve as a potential ‘eukaryotic gene delivery’ [1]. One of the greatest challenges however was the stability of the synthetic mRNA as well as the cost associated to produce it. Fast forward to the late 2000s and several pharmaceutical companies entered the mRNA field with a view to harness the potential of this exciting platform. The very first clinical trial of an mRNA-based vaccine for cancer immunotherapy was in 2008 that investigated its safety and tolerability [2].  However, the first mRNA vaccine that went through all the clinical stages and was FDA approved was against the virus SARS-CoV-2 and didn’t arrive until 2021 [3].

The demonstration of the safety and efficacy of mRNA-based therapeutics in humans, through the vaccines against SARS-CoV-2, has since laid the groundwork for the development of not only vaccines against other infectious pathogens but also mRNA-based drugs that target several other types of diseases. As of February 2023, more than 90 clinical trials were ongoing globally in the disease areas of cancer and infectious diseases [Fig 1], and several more are in the pipeline at various stages of development from preclinical to phase III [Fig 2].

The Case for mRNA Therapies

In the case of mRNA therapy for cancer, the mechanism of action is the same as for vaccines against pathogens- the mRNA transcripts are taken up by specialized immune cells such as the dendritic that subsequently translate and present the antigen to activate T and B cells against the specific proteins. The difference is that for cancer therapies, the mRNAs are designed against tumor antigens that can be potentially personalized based on the unique mutational signature.  This helps stimulate a highly specific adaptive immune response against the tumor. In addition, multiple, uniquely-designed mRNAs can be delivered simultaneously to broaden the antigenicity. For rare genetic metabolic diseases, mRNA therapy can be viewed as an alternative to protein replacement therapy, where the mRNAs lead to the production of a fully active protein instead of a non-functional one. Additionally, enabling the cells to synthesize the protein from mRNAs allows the proteins to be targeted to the desired cellular compartment such as the mitochondria or the cellular membrane.

But why is mRNA-based therapy being considered over the existing platforms such as DNA and proteins? First, mRNAs are transiently expressed and do not integrate into the host genome. This feature makes them safer overall [4]. Compared to direct protein delivery, mRNAs provide longer lasting expression of the therapeutic protein and therefore longer clinical benefit. In addition, commercial production of COVID-19 mRNA vaccines has led to the optimization and streamlining of the manufacturing process which is relatively simple, fast and inexpensive. The mRNA is produced in a cell-free system, can be designed quickly and scaled to need [5]. These advantages, together with its demonstrated efficacy in inducing a therapeutic response, are opening avenues for many challenging diseases to be addressed through mRNA therapy.

How are mRNA-based therapeutics manufactured?

The development of mRNA therapies is based upon rigorous analysis and quality control to enable safe and effective treatment.

The manufacturing process begins with the construction of a DNA vector into which the template from which the mRNA will be transcribed in vitro, is inserted. Genetic analysis tools such as Sanger sequencing by Capillary Electrophoresis (CE) and qPCR are an integral part of the vector construction process to confirm and validate that the correct sequence of the gene is inserted into the vector backbone. After sequence confirmation, the plasmid is propagated, purified and linearized, following which it is in vitro transcribed (IVT) to synthesize the mRNA transcripts. The mRNA transcripts are then purified from the IVT reaction.  At this stage, qPCR can be used to check for IVT efficacy, residual DNA contamination and adventitious agents such as microbes. The identity of the transcript is confirmed by Sanger sequencing.  In addition, the purity of the final product can be evaluated by using ultrasensitive digital PCR, to detect any residual non-mRNA products, and 16S sequencing, to identify bacterial contamination.

The structure of the mRNA is of key importance as it influences the translation efficiency and stability of the transcript in the cell. For mRNA-based therapeutics, the synthetic mRNA can be chemically modified by manipulating the termini, using altered bases, as well as modifying the sugar backbone to achieve maximum efficacy.[i] These modifications can also ensure the mRNA forms the correct structure and can influence biologic stability.

The delivery mechanism of mRNA for therapeutic purposes can vary. From a lipid nanoparticle-based formulation to the use of polymers such as polyethyleneimine (PEI), several delivery systems and routes of administration continue to be evaluated in an effort to achieve maximum efficacy and safety.

Conclusions & Future Perspectives

The new era of mRNA medicines is just beginning to unfold and the modality is becoming a key asset in the development pipelines of several biotech and pharma companies. While its introduction to the world came through SARS-CoV-2 vaccines, it aptly demonstrated the efficacy and safety of this first-in-class mRNA as a therapeutic. Since then, over 90 mRNA vaccines and drugs are in various stages of clinical testing for diseases that range from infectious to cardiovascular diseases. Several rare metabolic diseases such as Methylmalonic acidemia are also being targeted for mRNA therapy [6] and may bring renewed hope to patients and families that currently face limited treatment options.

Genetic analysis technologies such as dPCR, qPCR and Capillary Electrophoresis are a key part of the development and manufacturing workflow of mRNA therapies and are routinely used to evaluate their efficacy, safety and quality. The simplicity of the mRNA platform together with its low cost and ease of manufacture also opens the door to several economically challenged countries to adopt the technology for manufacturing mRNA therapies, and bringing medicine to millions across the world.

References

1. Malone, R. W., et al. (1989). “Cationic liposome-mediated RNA transfection.” Proc Natl Acad Sci USA 86(16): 6077-6081.
2. Weide, B., et al. (2008). “Results of the first phase I/II clinical vaccination trial with direct injection of mRNA.” J Immunother 31(2): 180-188.
3. FDA News Release https://www.fda.gov/news-events/press-announcements/fda-approves-first-covid-19-vaccine
4. Pardi, N., et al. (2018). “mRNA vaccines – a new era in vaccinology.” Nat Rev Drug Discov 17(4): 261-279.
5. Rosa, S. S., et al. (2021). “mRNA vaccines manufacturing: Challenges and bottlenecks.” Vaccine 39(16): 2190-2200.
6. https://investors.modernatx.com/news/news-details/2021/Moderna-Announces-First-Patient-Dosed-in-Phase-12-Study-of-mRNA-3705-for-Methylmalonic-Acidemia/default.aspx