Protein Corona, with a Twist

The high surface area, unique size-dependent properties, and precise tunability of nanoparticles make them ideal drug delivery and medical diagnostic platforms. However, nanoparticles undergo significant surface compositional changes when introduced into biological systems.¹ First, proteins weakly adsorb to the nanoparticle via noncovalent interactions to form a “soft corona.” Over time, this dynamic arrangement evolves, and weakly bound proteins are displaced by more tightly binding proteins to form a “hard corona.”² Because molecular recognition is key to how biological components interact with and respond to stimuli, the newly adsorbed proteins fundamentally alter the biological activity and fate of the nanoparticle. The protein corona can mask targeting functionalities on the nanoparticle and prevent it from reaching its intended destination; furthermore, if opsonin proteins such as immunoglobulin G (IgG) bind, they can mark the particle for clearance by the mononuclear phagocytic system. In the December 2019 issue of ACS Central Science, Zhang et al. report a strategy to overcome these limitations by proactively defining the nanoparticle protein corona composition to more efficiently enable cell-specific nanoparticle interactions.³

Previous strategies have attempted to minimize protein adsorption by functionalizing the nanomaterial surface with “stealth” polymers, such as polyethylene glycol, polyvinylpyrrolidone, or dextran.⁴ While this strategy successfully increases the circulating half-life of nanoparticles,⁵ modification with hydrophilic stealth polymers can still interfere with targeting functionality. Thus, instead of trying to completely avoid the formation of the protein corona, recent strategies have taken a new approach: recognize that protein corona formation is unavoidable and intentionally define the hard corona protein composition prior to introducing it into a biological system.⁶

This is precisely the strategy employed by Zhang and co-workers. In their present work, they used spherical nucleic acids (SNAs)—gold nanoparticles coated in a dense DNA ligand shell (oligodeoxynucleotides, ODN)—as the nanoparticle of choice. Because of their high DNA density, ability to bind complementary nucleic acid sequences, and unusual capability to enter into a broad range of cell types, SNAs have potential applications in gene silencing and drug delivery therapeutics.⁷ However, as is typical of nanoparticle systems, SNA surfaces are rapidly bound by endogenous proteins in biological systems. To avoid this, the authors used simple electrostatic interactions to deliberately form a hard corona of desired proteins, including antihuman epidermal growth factor receptor 2 (anti-HER2), IgG, and human serum albumin (HSA) (Figure 1). Dynamic light scattering (DLS), ζ-potential measurements, and electrophoretic mobility assays provided corroborating evidence that all three proteins form a hard corona around the SNAs. A series of in vitro characterization studies confirmed that the predefined coronae resisted exchange with serum proteins. Other important features of SNAs, including their ability to hybridize with complementary nucleic acid oligomers, were largely unaffected by the inclusion of a hard protein corona. When introduced to a mixed cell population of HER2+ and HER2– cells, SNAs preadsorbed with anti-HER2 preferentially targeted the former. In a final showcase, the authors preformed IgG and HSA coronae around SNAs and examined macrophage uptake of each particle type. IgG is an opsonin and marks nanoparticles for macrophage-mediated clearance, while HSA is a dysopsonin, which was expected to shield nanoparticles from macrophage uptake.⁸ In contrast to expectations, macrophages were less likely to clear SNAs with either IgG or HSA than bare SNA particles alone. Subsequent inhibitor studies suggested that IgG-modified SNAs are taken up via a different uptake pathway than HSA-modified or unmodified SNAs.

Figure 1.

Spherical nucleic acids (SNAs) predefined with a hard protein corona are able to target specific cell types, avoid macrophage clearance, and/or engage with different uptake machinery, depending on the protein adsorbed.

The results from the present report dovetail nicely with a recent study in which mesoporous silica particles were also shown to benefit from preformed protein coronae.⁹ Taken together, these works hint that predefining the protein corona may be a strategy that is generally applicable to the broad diversity of nanoparticles used in biomedical applications. A major benefit of the present work is its ease of preparation. Proteins are simply preadsorbed on the nanoparticle surface, which yields improvements in targeting efficiency and alters the process by which macrophages engage with the particles. Indeed, it may be the noncovalency of the interaction that enables the adsorbed proteins to reach a thermodynamically stable orientation on the particle surface during preparation. Because SNAs with defined coronae can be easily prepared, the workflow established by Zhang and co-workers is likely to enable rapid screening of antibody-modified SNAs directed to other clinically important targets. However, these targeted efforts should be buttressed with complementary lines of inquiry to bridge the gap between cell studies and in vivo studies. For instance, interrogating protein corona stability and exchange dynamics under shear flow will provide a valuable trove of information to help expedite the translation of nanoparticle-based therapies to clinical applications.

The authors declare no competing financial interest.