Author: Junaid Kashir, Associate Professor, Department of Biology, Khalifa University
The recent COVID-19 pandemic led to drug regulatory authorities clearing the use of mRNA-based vaccines against the outbreak. However, introducing RNA into cells to produce resultant molecules has been common practice in developmental biology/reproductive sciences to inhibit, induce, and identify several factors as potential therapeutics and diagnostics in humans.
1. What is mRNA technology?
mRNA-based therapies involve translating exogenous mRNA into functional proteins, either within individual cells, either in a laboratory setting or within organisms. The first such clinical application was using mRNA as a novel rabies vaccine in 2013 (NCT02241135). The most recent example of this approach was of course in 2020, with the advent of mRNA-based vaccines against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), with the Pfizer-BioNTech (BNT162b2), Moderna (mRNA-1273), and CureVac101–103 vaccines being the fastest to be developed in medical history.
The concept is that the host cell(s) converts the mRNA sequence to express the antigen of interest, which is degraded into small peptides that will be presented as endogenous antigens, and thus ‘training’ the immune systems in preparation for a viral infection which will present with similar antigens, triggering humoral and cellular responses.
2. How does mRNA technology relate to embryology?
RNA is very closely linked within animal reproduction and embryos, especially within the female cell; the oocyte/egg. Indeed, for conversion of eggs/oocytes into embryos, the proper storage of mRNA is essential. These mRNAs play a significant role in supporting embryogenesis. While no new mRNA is made until later stages of embryogenesis, the female gamete contains all the required machinery to convert mRNA in to protein or peptides. Indeed, researchers have been using such properties in their research, by injecting RNA into oocytes and observing the effects of the resultant proteins upon fertilisation and embryogenesis.
3. How can mRNA technology be applied for fertility treatments?
The injection of mRNA into oocytes and embryos has long been thought to represent a potentially powerful therapeutic approach within various areas of embryology and fertility treatment. An example is oocyte activation, which involves the release of calcium within the oocyte at fertilisation, a pivotal signal in every animal species studied to date. These rises in calcium are triggered by a soluble factor, a sperm-specific phospholipase C zeta (PLCζ) in mammals, introduced into the oocyte by the fertilising sperm. Sperm from human patients that are defective in this process are unable to release the required calcium and are infertile. However, when this sperm is co-injected into oocytes alongside mRNA coding for PLCζ, calcium is released, and fertilisation able to proceed, the idea of course being that the oocytes active transcriptional machinery converts the RNA to protein, leading to activity.
Collectively, such studies suggest that this approach represents a potentially immensely important therapeutic avenue to treat cases of infertility. Indeed, such an approach could also be extended to other markers of embryogenic health, which could represent targets in the fertility clinic to improve successful embryogenic rates to regulate keys events during embryogenesis to perhaps enhance chances of successful pregnancy within the clinic.
4. What other areas could mRNA technology benefit in the fertility clinic?
mRNA technology also could be applied in a diagnostic aspect, especially within the context of calcium signals at fertilisation which are extremely indicative of embryogenic competency. However, current methods of imaging calcium involves using calcium-specific dye (dyes that emit light in the presence of calcium) hold a number of concerns – namely that these methods require constant light exposure which lethally damages the developing embryo, and thus cannot be used in clinical human fertility treatments. Furthermore, the delivery method of some dyes involves either cell permeabilisation or bulk injections that may damage the cell. A proposed alternative is to use mRNA that encodes for genetically encoded calcium indicators (GECIs), which operates based upon fluorescence resonance energy transfer (FRET), which is much more efficient and safer than current methods.
5. Are there any other ways to deliver mRNA to eggs/oocytes apart from injection?
Quite astoundingly, nature already has concoted a way of delivery endogenous RNA to eggs/oocytes – via sperm! While most sperm-based mRNAs that are transferred to the oocyte are degraded, these transcripts play a huge role crucial to the early embryonic development. Thus, research has begun considering sperm as a ‘delivery vehicle’ to transport therapeutic/diagnostic RNA, with some recent advancements using spherical mesoporous silica nanoparticles (MSNPs), liposome complexes/nanoparticles, lipid and other materials-based nanoparticles, polymer-based strategies, and most intriguingly, the use of hydrogels which can be easily engineered to deliver RNA in a specific and controlled spatiotemporal manner.
6. Why have such methods not been expanded into the fertility clinic yet?
As with any emerging technologies, there is always initial (and justified) concern surrounding its use. In the case of mRNA technology, issues used to consist of aspects such as mRNA efficiency, safety, stability, immunogenicity, and efficacy of delivery systems. However, the recent explosive interest in the therapeutic use of mRNA has significantly reduced these areas of concerns by the introduction of chemical modifications and enhanced delivery systems such as exosomes and lipid nanoparticles, with further modifications designed to enhance mRNA stability.
However, there do remain some areas that require elucidation before application within the context of fertility and embryogenesis can proceed. For instance, exposure to heavy metals could alter mRNA expression pattern (increasing or decreasing levels). Similarly, some drugs can affect mRNA synthesis, altering expected production. The area of most concern, specific to embryos, is that mammalian embryos possess a retrotransposon-encoded reverse transcriptase activity, which generate a wave of ‘reverse transcription’ that could result in DNA being generated from the introduced mRNA that could possibly be integrated into the genome – resulting in ‘transgenic’ babies. However, it is possible that given recent advances in chemical mRNA modifications that could perhaps prevent this integration.
7. How soon can such methods be applied?
mRNA therapeutics represent a potentially powerful tool to enhance quality and success of fertility treatments. Indeed, RNA has been traditionally a widely used technique in an extensive repertoire of methods used to study reproductive biology. While, such applications have been traditionally brimming with limitations and safety considerations, preventing clinical application, the recent advances and enhanced interest in mRNA technology should cause us to reconsider our traditional reluctance. Given how often such technology has already been used in the field, with significant advances already been made, such technology could be applied following appropriate safety studies in cellular and animal systems.