Mapping out the pathways for spherical nucleic acid biodistribution

Zhang, Wuliang

doi:https://doi.org/10.21985/n2-3y6b-j173

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Mapping out the pathways for spherical nucleic acid biodistribution

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Oligonucleotides can be used to modulate the regulation of pathological genes that are associated with various diseases. However, due to biological barriers, efficient delivery of oligonucleotides, especially to extrahepatic tissues, remains a challenge. To overcome these barriers, multiple delivery strategies have been developed, ranging from medicinal chemistry to nanotechnology. Nanoparticle-based delivery emerges as a promising approach, marked by its tunable physicochemical and biological properties. Unlike typical nanoparticles that encapsulate oligonucleotides, spherical nucleic acids (SNAs) consist of nanoparticle cores and, onto their surfaces, densely functionalized with radially oriented oligonucleotides. Both the core and the surface ligands can be modulated to enable therapeutic or diagnostic functionalities. In addition, the dense surface oligonucleotides lead to high cellular internalization, compared to linear oligonucleotides, pointing towards their application in biomedicine. Indeed, SNAs have been clinically developed for gene regulation and immunomodulation purposes targeting different forms of cancer, diseases of the skin, and neurological disorders. However, the blood circulation and biodistribution profiles of SNAs need to be improved, especially for uses involving systemic administration. This dissertation seeks to uncover the secrets to prolonging the blood circulation time and elevating SNA accumulation in extrahepatic solid tumors (and the cells that comprise them) by chemically or physically modifying the surface of SNAs. In chapter 2, changing the protein coronae by physically adsorbing active proteins (e.g., antibodies, proteins with low affinity for macrophages) onto SNAs is explored. Significantly, most of the adsorbed active proteins are found to remain on SNA surfaces even in the presence of human serum, and the protein coronae do not hinder the surface oligonucleotides from binding to their complementary strands. This physical adsorption approach for modifying SNAs with a protein corona is easier to implement than the conventional chemical coupling methods and minimizes loss of protein function following bioconjugation. Chapter 3 assesses how modulating multiple polyethylene glycol (PEG) parameters affects the biological fate of SNAs. Different from the previous study on PEGylated SNAs with gold nanoparticle cores, this study uses a liposomal core, allowing noncovalent functionalization with PEG. Unlike covalently functionalized PEGylated SNAs that do not exhibit prolonged blood circulation and high cellular uptake simultaneously, the sheddable PEG shell provides SNAs with both characteristics in a single construct. In addition, the stability of the noncovalently functionalized PEG shell, marked by de-PEGylation kinetics, is a key determinant of the biodistribution and cellular uptake of SNAs. This chapter shows that if one’s aim is to improve the abundance of SNAs inside target cells, simply PEGylating the materials may not be sufficient. Indeed, a set of PEG-relevant factors need to be carefully chosen to achieve this goal. Instead of relying on passive shedding of the protective shell (i.e., PEG in chapter 3), in chapter 4, we incorporate a tumor-associated enzyme-cleavable protective shell into SNAs; this shell can be cleaved at the tumor site selectively. In this work, PEG is replaced with a zwitterionic peptide with a nearly neutral net charge as a less immunogenic alternative. By being protected by an enzyme-responsive zwitterionic peptide shell, such SNAs display prolonged circulation in the bloodstream, enhanced accumulation in solid tumors, and improved internalization into cells in the tumors, as compared with native SNAs. Significantly, this study demonstrates that by having a stimulus-responsive protective shell, SNAs can be made to exhibit more desirable biodistribution; most notably, by replacing PEG with a low immunogenicity ligand, concerns over the PEG-related immunogenicity can be addressed.

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