How AstraZeneca can improve lipid nanoparticle formulations for gene delivery using neutron scattering


In recent year, new therapeutic modalities involving delivery of biological molecules such as RNA, peptides and oligonucleotides have shown promising results to treat diseases that are currently hard to tackle with standard small molecules approaches. As an example, last year Alnylam announced the first therapy using small interference RNA (siRNA) approved by the Federal Drug Administration (FDA) for the treatment of the polyneuropathy of hereditary transthyretin-mediated amyloidosis. This sounds very exciting, but it also shows that there might be a long road for the approval of RNA-based therapies given the small number of approvals, especially messenger RNA (mRNA) given the small number of clinical trials that are still in early phases. Therapeutics containing siRNA aim to knockdown protein production while mRNA intends to produce proteins needed in the body. Additionally, mRNA is a very large molecule compared to siRNA, which makes it difficult to be transported  through the body to reach specific cells in different organs and to cross the cell membrane without the help of a delivery vehicle.

Lipid nanoparticles (LNPs) formed by cationic ionizable lipids (CILs) are potential delivery vectors since they do not elicit strong immune responses compared to viral vectors and are more efficacious compared to other non-viral alternatives. To improve the performance of LNPs containing mRNA, it is necessary to understand how their structure is related with cellular uptake and endosomal escape, an important step for the success of this type of therapies. LNPs also contain helper lipids such as cholesterol, phospholipids and poly(ethylene glycol) (PEG) lipids and have, therefore, a very complex structure to determine.


Small angle neutron scattering (SANS) in combination with isotopic contrast variation gave us important structural information that it was not accessible previously by other methods. Since hydrogen and deuterium have different scattering profiles, selective deuteration of the lipid molecules in the LNPs and the solvent allowed us to highlight the structure of the LNPs and to pinpoint the location of the different components. The figure below shows some of the important results of the published work.[1] In this work, it was found that the phospholipid DSPC and the PEG-lipid were mainly located at the surface of the LNPs. This was the first time that one technique explicitly showed this structure since previous molecular dynamic simulations predicted that the phospholipid was evenly distributed across the LNPs. We used this information to vary the size and surface composition of the LNPs, which translated to an increase of protein production and therefore more effective LNPs. We found that the surface of the LNPs most likely play a major role in endosomal scape and, thus, its design can help to improve the performance of the formulations.

Figure A) Left: Cryo-TEM image of LNPs, Right: SANS data of LNPs in buffer with different D2O content and lipid distribution according to the model. Figure B) Left: Surface area per DSPC with constant (pink) and variable (cyan) surface composition, Right: in vitro expression of hEPO protein in human adipocytes for LNPs with constant (pink) and variable (cyan) DSPC surface area.


The determination of the structure of the LNPs has been coordinated by the research scientists Marianna Yanez Arteta, Aleksandra Dabkowska and Lennart Lindfors in Pharmaceutical Sciences, IMED Biotech Unit, AstraZeneca, Gothenburg, Sweden. This study had collaborators from University of Gothenburg and Jülich Centre for Neutron Science. The experiments were performed at the Research Neutron Source Heinz Maier-Leibnitz (FRM II).

What is next?

We still need to learn more about how changes in the composition of the LNPs will affect their performance. To optimize the targeting of these nanoparticles to specific cells, we need to understand better how the variations in the structure of the LNPs affect their interaction with membrane proteins. Dr. Federica Sebastiani, in the group of Professor Marité Cardenas at Malmö University together with AstraZeneca and Attana, will start to tackle part of this challenge soon. In her research, she will focus on understanding how the structure of LNPs affect their interactions with lipoproteins and how can this information be used to optimize in vitro and in vivo studies. This work is of high importance as it aims to improve pre-clinical work since AstraZeneca is committed to the ethical use of animal models in research.

[1] M. Yanez-Arteta et al, Successful reprogramming of cellular protein production through mRNA delivered by functionalized lipid nanoparticles, PNAS, 115, E3351 (2018) (


Marianna Yanez Arteta, PhD
Senior Scientist Formulation/Physical Chemistry
Pharmaceutical Science, Advanced Drug Delivery
L240, Pepparedsleden 1, Mölndal, 431 83, Sweden
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