Effect of Protein Corona on The Transfection Efficiency of Lipid-Coated Graphene Oxide-Based Cell Transfection Reagents.

Erica Quagliarini,Francesco Cardarelli,Giulio Caracciolo,Gianmarco Ferri,Riccardo Di Santo,Daniela Pozzi,Sara Palchetti

doi:10.3390/pharmaceutics12020113

Abstract

Coating graphene oxide nanoflakes with cationic lipids leads to highly homogeneous nanoparticles (GOCL NPs) with optimised physicochemical properties for gene delivery applications. In view of in vivo applications, here we use dynamic light scattering, micro-electrophoresis and one-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis to explore the bionano interactions between GOCL/DNA complexes (hereafter referred to as ”grapholipoplexes”) and human plasma. When exposed to increasing protein concentrations, grapholipoplexes get covered by a protein corona that evolves with protein concentration, leading to biocoronated complexes with modified physicochemical properties. Here, we show that the formation of a protein corona dramatically changes the interactions of grapholipoplexes with four cancer cell lines: two breast cancer cell lines (MDA-MB and MCF-7 cells), a malignant glioma cell line (U-87 MG) and an epithelial colorectal adenocarcinoma cell line (CACO-2). Luciferase assay clearly indicates a monotonous reduction of the transfection efficiency of biocoronated grapholipoplexes as a function of protein concentration. Finally, we report evidence that a protein corona formed at high protein concentrations (as those present in in vivo studies) promotes a higher capture of biocoronated grapholipoplexes within degradative intracellular compartments (e.g., lysosomes), with respect to their pristine counterparts. On the other hand, coronas formed at low protein concentrations (human plasma = 2.5%) lead to high transfection efficiency with no appreciable cytotoxicity. We conclude with a critical assessment of relevant perspectives for the development of novel biocoronated gene delivery systems.

Highlights

In the last years, the interest in manifold nanomaterials has rapidly been growing
dynamic light scattering (DLS) and microelectrophoresis measurements showed that GO-cationic lipid (GOCL) NPs had optimal physicochemical properties for gene delivery purposes, as they were small in size (212.3 ± 9.1 nm, diameter) and positively charged
Despite the huge amount of preclinical data involving the use of gene delivery systems, their clinical application is far from established

Summary

Introduction

The interest in manifold nanomaterials has rapidly been growing. Among these, graphene oxide (GO) is promising as a potential nano-vector for several biomedical applications because of its peculiar physicochemical surface properties (e.g., high dispersibility in aqueous solvents, high biocompatibility and high specific surface area) [1,2,3,4]. Factors shaping a PC, such as the NPs’ properties (e.g., surface area, surface charge, shape and solubility); protein source (e.g., plasma vs serum) and environmental influences (e.g., incubation temperature, exposure time and shear stress) have all been largely documented [15,16,17,18,19] Among these factors, the amount of proteins in the physiological environment may significantly affect the tissue engineering (TE) of gene delivery systems [14]. We have recently understood that, depending on the route of administration (e.g., systemic vs administration), PC of gene delivery systems can change in response to bodily fluids with completely different protein concentrations (e.g., gastrointestinal fluid [21] vs lung surfactant [22], etc.) For all these reasons, exploring the bionano interactions with the biological milieu is emerging as the missing link between benchtop discoveries and clinical applicability of gene delivery systems [7]

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Discussion

Conclusion