Polymeric and lipid nanoparticles (NPs) are an effective and safe method for therapeutic delivery. At each of the various steps in nanoparticle development and manufacturing (i.e., research and design, quality assurance), assessing the features of these vehicles is essential. Some features (i.e., packaging characteristics, cargo location, structure assembly, and dynamic behavior during purification) provide important insights that can further the understanding of structure-property function and the optimization of NP design. However, payload distribution and capacity - key characteristics that inform pharmacodynamics, pharmacokinetics, and delivery efficiency – are difficult to characterize at the single NP level with current methods. Therefore, there is a clear need for new methods that improve the assessment of NP payloads, particularly at the single NP level.
Technology Overview
Researchers at Johns Hopkins University have developed a multi-laser cylindrical illumination confocal microscopy (CICS) technique that can assess cargo and carrier content of delivery vehicles at single-NP level. The technique features in-line examination in a flow set-up, 100% mass detection efficiency and single-cargo sensitivity. When multiple fluorescent tags are used, it differentiates distinct populations within a formulation (e.g., free mRNA, mRNA-loaded lipid nanoparticles, and empty lipid nanoparticles), and then quantitatively resolve single-cargo/carrier fluorescence to give a distribution of single-NP payload level. In addition, the method can be coupled with hydrodynamic separation to resolve coupled data of size (diameter) and payload of the same NP, which provides unprecedented level of details for investigation of NP assembly mechanisms.
Stage of Development
Proof of concept work has been completed and demonstrated for mRNA lipid nanoparticles (LNPs, published as a research article in Nature Communications), in which the distribution of mRNA copy per LNP was successfully characterized for multiple formulations, with populations of empty (unloaded) LNPs identified. We are working with more collaborators to apply the technique to analyze different cargos, including proteins, small molecule drugs, and other nucleic acids in diverse NP systems including PLGA NPs, exosomes, and extracellular vehicles (EVs). We are seeking for more collaborators with interest in characterizing single-NP payloads.
Publication
https://www.nature.com/articles/s41467-022-33157-4
https://hub.jhu.edu/2022/09/23/cics-platform-for-measuring-genetic-medicine-in-lipid-nanoparticles/
