Emerging Technologies Capable of Unlocking the Promise of Biologically Targeted mRNA Therapies

Emerging Technologies Capable of Unlocking the Promise of Biologically Targeted mRNA Therapies

In a recent article published in Nature BiotechnologyResearchers reviewed technological advances that will unlock the promise of biologically targeted messenger ribonucleic acid (mRNA) therapies beyond vaccines.

Study: unlocking the promise of mRNA therapies.  Image Credit: nobeastsofierce/Shutterstock
To study: Unlocking the promise of mRNA therapies. Image Credit: nobeastsofierce/Shutterstock

Background

The first part of the review focused on the design and purification of mRNA payloads, including novel forms such as circular mRNA (circRNA) and self-amplifying mRNA (saRNA). The second part discussed improved mRNA packaging systems, including ionizable lipid nanoparticles (LNPs), to improve cargo delivery.

In part three, the researchers reviewed the engineering of packaging systems that will facilitate the targeting of mRNA therapies to specific tissues. The fourth and fifth parts discussed strategies that enable the treatment of chronic diseases through mRNA therapies and a synopsis of current clinical trends in mRNA therapies. Finally, the researchers emphasized the short-term and long-term scope of new mRNA therapies.

Background

The unprecedented success of coronavirus disease 2019 (COVID-19) vaccines based on the mRNA technology platform has renewed interest in this therapeutic area. However, several challenges still prevent establishing mRNA technology as a general therapeutic modality with broad applicability against various clinical conditions.

Advances in the area of ​​protein expression, packaging systems, tissue selection, and chronic dosing

Immunization requires minimal levels of protein expression, whereas mRNA therapy requires a 1000-fold higher protein level to reach a therapeutic threshold. Efficient delivery to solid organs remains a challenge. Even the tissue bioavailability, circulatory half-life, and efficiency of the LNP-based carrier could be rate-limiting when delivered to the target tissue. Even with optimized mRNA chemical modifications and advanced LNPs, chronic dosing eventually activates innate immunity, in parallel attenuating the expression of therapeutic proteins.

An individual mRNA has a cap, 5′ and 3′ untranslated regions (UTRs), an open reading frame (ORF), and a polyadenylated (poly(A)) tail. There have been advances in the design of each of these components. The most notable of these are:

i) Improved 5′ cap analogs which improve translational ability, but more importantly, cap efficiency from 70% to 95%.

ii) optimization of the length of the poly(A) tail has been shown to be essential in balancing the synthetic capacity of an mRNA cargo.

iii) Optimization of the UTR sequence could improve protein expression of an mRNA cargo by a few fold, allowing its customization for the target biological area and disease-driven microenvironment.

iv) Studies have documented over 130 natural chemical modifications to mRNA so far. Chemically modified nucleosides, particularly uridine moieties, such as methylpseudouridine, can reduce recognition by toll-like receptors of innate immunity by up to 100-fold, which, in turn, markedly increases protein expression after live transfection of mRNA cargoes. In the future, clinically effective unmodified therapeutic mRNAs that will hide from the immune system and have higher translational efficiency may become available. livesimilar to chemically modified mRNA vaccines.

Likewise, saRNAs could be favorable for enzyme replacement therapies. They require ~10-fold less RNA for protein expression of similar magnitude compared to linearly modified mRNA and are under live testing, a scalable process for vaccine production. Another alternative to linear mRNA is circRNA, which has been shown to double the life of mRNA. invitro. circRNA circumvents the need for costly 5′ capping, the tedious 3′ poly(A) tail, and increases total protein yield without increasing protein expression levels compared to linearly modified mRNA.

Overcoming challenges related to the breadth of protein expression, in parallel with mRNA structural optimizations, could alleviate the need for repeated dosing, a major requirement that makes it difficult to treat chronic diseases using mRNA therapies. Conventional treatment involves systemic injection of recombinant coagulation proteins (factor VIII/IX) three to seven times per week due to their relatively short half-life of ~12 hours. In contrast, preclinical studies in mice have shown that a single weekly systemic injection of 0.2 to 0.5 mg kg−1 modified linear mRNA could treat hemophilia A and B while maintaining protein levels above a clinically relevant threshold.

There are four types of mRNA packaging systems: biomimetic, lipid- and cell-based packaging, and extracellular vesicle-based packaging. LNPs were first reported six decades ago and have since undergone several advances, leading to their first clinical use as a small RNA interference (siRNA) delivery vehicle. The other three packaging systems are still in the preclinical evaluation stage.

Cationic lipids induce cytotoxicity and exhibit low transfection efficiencies due to rapid splenic and hepatic clearance. In contrast, ionizable cationic lipids are neutral, preventing them from cellular or molecular recognition. Therefore, upon cellular uptake, they fuse with endosomes, releasing the mRNA cargo into the cell cytoplasm for translation. LNPs composed of MC3, which first received regulatory approval in 2018, show ~20-fold lower median effective dose (ED50) in animal models and are also currently used in COVID-19 mRNA vaccines.

The scope of mRNA therapy

Compared to mRNA vaccines that have successfully completed phase III clinical trials, most mRNA therapies are in early phase I clinical trials, primarily focused on safety. mRNA therapies could deliver any protein locally or systemically, including enzymatic, secreted, mitochondrial membrane, intracellular, receptor, and gene-editing proteins. However, only two clinical studies have produced encouraging results on its efficacy and safety.

The secreted proteins offer “nearest neighbor” effects beyond the few cells that are transfected. Like paracrine vascular endothelial growth factor (VEGF), they could have clinical applications through tissue-specific delivery. Recent studies on VEGF, with live delivery systems, are expanding the potential role of mRNA therapies in wound healing, peripheral vascular physiology, and bone repair.

Conclusions

The future of mRNA drugs may depend on the rapid evolution of mRNA cargo, intracellular transporters, and live delivery systems along with deep biological and clinical knowledge. However, the versatility of mRNA could trigger therapeutic opportunities and thus its other innovative applications are expected to come in the near future. For example, recent research has shown the utility of mRNA technology to live expression of intracellular antibodies for the treatment of heart failure and as in vitro disease modeling tool.

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