How to make the challenge of mRNA delivery easy


Many researchers are facing the challenges of mRNA delivery, through the body into the cytoplasm of cells. In this post, we invite you to discover some useful and efficient solutions to improve and control all the steps of your mRNA delivery project.

Messenger RNA is a new biological entity for drug discovery with promising potential to treat many diseases, especially during the current COVID-19 pandemic. In 2014, the first cancer clinical trial using mRNA encapsulated in lipid nanoparticles (LNP) took place. Then, as we know, 2020 saw the efficiency of the BioNtech mRNA vaccine against COVID-19. A less well known fact, is that it also saw the first mRNA-based gene editing clinical trial. These two treatments both use LNP for mRNA delivery, as you’ll see below.



The favourite mRNA delivery vehicle – Lipid nanoparticles (LNP)

There are already many clinical trials using mRNA with LNP to fight infectious diseases such as Chikungunya and the flu, different type of cancers, and genetic disorders, a summarized in this very interesting review: Hou, X., Zaks, T., Langer, R. et al. (2021).

mRNA delivery with lipid nanoparticles (LNP)

To make it simple, LNP are a mix of lipids of which we can distinguish four main kinds :

  • Cationic lipid: with a positive charge that allows good encapsulation of mRNA that is negatively charged
  • Ionisable lipid: which facilitates the administration through blood and endosomal escape
  • PEG lipid: which reduces non-specific uptake
  • Helper lipid: which plays a role in stabilising the LNP

To improve delivery performance and stability of the nanoparticles, other lipids can be used such as phospholipids and cholesterol.

Lipids of the nanoparticles themselves can be toxic. To avoid such drawback and immune response, the solution would be biodegradable lipids such as used by Moderna (SM102) and BioNtech (ALC-0315) for their respective mRNA vaccines against COVID-19. Both solutions are ionisable lipids that are better eliminated in the body.

Based on our broad offer of lipids solutions, you can quickly access the 4 BNT126b2 vaccine lipids :

But LNP can trigger pro-inflammatory cytokine production. As you can read in this post on our blog, we offer you a complete solution to quickly and accurately quantify cytokines all together such IFN-gamma, IL-1alpha, IL-6, IL-10 and many more if needed.

No time for the quantification? Let our lab experts do the job for you – discover our Biomarker service.

Emerging alternatives to lipid nanoparticles

Lipid nanoparticles are not the unique solution. We are also observing platforms of mRNA delivery with micelles, such as polylactide-based or PEG-based micelles.

This summer, the first gene editing in mouse brain was successful thanks to Trilink’s CAS9 mRNA delivery using micelles. Take a look at the details in this post: CleanCap® IVT Cas9 mRNA in Micelles Used for Genome Editing in Mouse Brain for the First Time

Recently, direct PEGylation of mRNA without cationic lipids was shown to be efficient (Yoshinaga et al 2021). Here the PEGs have been conjugated to oligonucleotides complementary to the mRNA. Are you facing any questions about PEGs or Conjugation? We’ll be pleased to offer the right products or services to answer your needs.

When looking for non-synthetic solutions, exosomes are a good lead as natural vehicles from the blood. Some platforms develop encapsulation and mRNA delivery with patient cell-derived exosomes. We are getting deeper into personalised treatment.

Monitor mRNA delivery with high quality control CleanCap mRNA

How do you monitor and thus optimise mRNA delivery into cells, taking in account that mRNA is fragile and the cells could response against foreign RNA as if it were a viral attack?

CleanCap reagent for mRNA delivery
CleanCap technology

The capping matter:

It must be with a CAP1 structure, because it’s the natural shield in mammalians that reinforces the mRNA. Trilink Biotechnology, the worldwide leader in mRNA technology, has developed ideal reagents for co-transcriptional CAP1 capping close to 100% of efficiency in one pot:

The latter one is used in the BNT162b2 of BioNtech’s vaccine against COVID-19.

CleanCap® capping also helps the cells better accept the exogenous mRNA. However, U-depletion in the coding sequence greatly reduces the cellular response.  The remaining Uracile (around 15% ideally) can be masked with substitution to N1mePseudo-U, Pseudo-U or 5-mo-U (5-methoxy-UTP). This robust design is required and has been used by Trilink to develop reliable control mRNA.

Start with a reliable and flexible mRNA transfection reagent

A possible strategy is to start testing the proof of concept with mRNA coding for the protein of interest, before investing in extensive delivery optimisation studies. For this purpose, a robust transfection reagent which is efficient both in cell culture and injection for mouse is ideal. RJH Biosciences have developed an efficient transfection reagent mRNA-Fect which, versus classical transfection solutions (e.g. Lipofectamine in the picture below), have shown better and higher performance with adherent cells, suspension cells and in vivo (specific protocol upon request).

mRNA-Fect compare to Lipofectamine
mRNA-Fect with adherent cellsSuspension dependent cells transfected with mRNA-Fect

Now, how do you handle the challenges of mRNA delivery?

The success of research with mRNA is definitely linked to mRNA delivery performance. Many solutions are in development with continuous efforts to improve it. For now, there is not yet an optimal and universal vehicle. mRNA delivery studies are really part of projects and require reliable control. That explains, as illustrated by their popularity, why CleanCap® control mRNA are becoming the gold standard.

To help you to answer these questions, at tebu-bio, you’ll benefit from our strong expertise in the mRNA world, thanks to a broad range of high quality solutions and services (e.g. our new IVT Laboratory Services), and dedicated technical experts to find the best solutions and to accompany you throughout your project.

What’s your next step? Any Questions?

Contact Dimitri, our mRNA technical expert through the comments form below, he’ll be happy to advise!


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