Protein Corona Formation on Hard and Polymeric Nanoparticles – Towards Understanding Biocompatibility, Biodistribution, and Efficacy

Abstract

Unpredictable protein adsorption on both hard and soft nanoparticles remains a considerable challenge towards effectively applying nanotechnologies in biological environments. Hard nanoparticles form the basis of many chemical nanosensors. Conversely, soft nanoparticles such as lipid nanoparticles (LNPs) are vital for the successful delivery of mRNA-based vaccines, and offer promising applications in neonatal gene therapy, immunotherapy, and protein replacement therapy. Understanding the biological interactions that both hard and soft nanoparticles undergo upon introduction into biological systems is central to optimize the outcomes of nanoparticle-based delivery biotechnologies in clinical settings.Herein, we present a multimodal study of protein corona composition and dynamics, first on ‘hard’ nanoparticles: spherical polystyrene nanoparticles (a previously studied model nanoparticle) and high aspect ratio single-walled carbon nanotubes (SWCNTs, an understudied nanoparticle). These nanoparticles are exposed to two biofluids of interest: blood plasma (relevant for intravenous applications) and cerebrospinal fluid (relevant for brain imaging and sensing applications). To study these protein coronas, we develop a methodology based on quantitative proteomic mass spectrometry [1] and chemical thermodynamic analysis of real-time protein binding to identify protein corona ‘fingerprints’, enabling quantification of protein abundance and enrichment/depletion relative to the native biofluid, transient kinetics [2], and end-state topology. Interestingly, we find that the heavily studied polystyrene nanoparticles are relatively agnostic in the formation of their protein coronas, demonstrating little preference for particular protein classes or physicochemical properties. Conversely, we find that SWCNTs show strong preference for certain protein classes. Our additional work in machine learing-based analysis shows that corona compositions, and more broadly nanoparticle biofouling, can be drastically different for each nanoparticle type [3].Lastly, we study nano-bio interactions encountered by ‘soft’ nanoparticles: LNPs commonly used for the therapeutic delivery of mRNA. We investigate how modifying (i) the mRNA packaged inside the LNPs and (ii) the ionizable lipid within the LNPs modulate the subsequently formed protein corona in (iii) various biological environments of relevance for delivery applications. Importantly, this workflow is readily translatable across soft polymer-based nanotechnologies of interest, which are understudied due to the experimental complexity of separating nanoparticle-corona complexes from free proteins. This fundamental understanding of protein-LNP interactions could enable more seamless design and clinical application of next-generation LNP carriers to bolster the safe and effective delivery of mRNA and other therapeutics to patients. References Pinals, R.L., et al., Quantitative Protein Corona Composition and Dynamics on Carbon Nanotubes in Biological Environments. Angewandte Chemie (2020).Pinals, R. L., Yang, D., Lui, A., Cao, W. & Landry, M. P. Corona Exchange Dynamics on Carbon Nanotubes by Multiplexed Fluorescence Monitoring. JACS (2020).Ouassil, N.*, Pinals, R.L.*, O’Donnell, J.T.D., Wang, J., Landry, M.P.‡ Supervised Learning Model to Predict Protein Adsorption to Nanoparticles. Science Advances (2022).