Spherical Nucleic Acid Nanoparticles: Therapeutic Potential

Abstract

Spherical nucleic acids (SNAs) are highly oriented, well organized, polyvalent structures of nucleic acids conjugated to hollow or solid core nanoparticles. Because they can transfect many tissue and cell types without toxicity, induce minimum immune response, and penetrate various biological barriers (such as the skin, blood-brain barrier, and blood-tumor barrier), they have become versatile tools for the delivery of nucleic acids, drugs, and proteins for various therapeutic purposes. This article describes the unique structures and properties of SNAs and discusses how these properties enable their application in gene regulation, immunomodulation, and drug and protein delivery. It also summarizes current efforts towards clinical translation of SNAs and provides an expert opinion on remaining challenges to be addressed in the path forward to the clinic.

Fig. 1

Schematic showing the various spherical nucleic acid (SNA) structures that have been developed. As a generalizable technology, SNAs consist of a ‘shell’ of nucleic acids surrounding a solid or hollow core nanoparticle. Reproduced with permission from [102]

Fig. 2

Oligonucleotide dimensionality dictates cellular endocytosis. a Confocal microscopy shows that Cy5-ssDNA cannot readily enter C166 cells. b SNAs prepared with the same DNA as utilized in part a display substantial cellular uptake within 2 h after incubation. This demonstrates that the 3D architecture of SNAs is critical to their cellular interactions. Reprinted with permission from Choi et al. Mechanism for the endocytosis of spherical nucleic acid nanoparticle conjugates. Copyright 2013. Proc Nat Acad Sci USA [39]. Cy5 cyanine 5, Cy5-ssDNA Cy5-labeled single-stranded DNA, C166 cells mouse endothelial cell line, SNA spherical nucleic acid

Fig. 3

Scheme depicting RNA interference therapy. Ordinarily, DNA is transcribed into mRNA, which is then translated into protein. In RNA interference therapy, siRNA (shown here) or miRNA is delivered into cells complexes with the RISC and guides it to complementary mRNA molecules in the cytoplasm, which are subsequently degraded. Reprinted with permission from Kreuzberger et al. Nanoparticle-mediated gene regulation as a novel strategy for cancer therapy. Copyright 2017. Delaware J Public Health [42]. mRNA Messenger RNA, miRNA microRNA, RISC RNA-induced silencing complex, siRNA small interfering RNA

Fig. 4

Topically applied SNAs can penetrate skin to regulate target genes and improve wound healing. a Cy5-labeled SNAs (red) are present throughout the stratum corneum and nucleated epidermis of human skin equivalents (EpiDerm; MatTek) following a single application. Blue, Hoechst 33343-stained nuclei. Scale bar = 50 μm. Reprinted with permission from Zheng et al. Topical delivery of siRNA-based spherical nucleic acid nanoparticle conjugates for gene regulation. Copyright 2012. Proc Nat Acad Sci USA [19]. b Representative images of wounds in diabetic obese mice treated with NS-SNA or GM3S-SNA. Topical application of GM3S-SNAs resulted in more rapid wound healing. Reprinted with permission from Randeria et al. siRNA-based spherical nucleic acids reverse impaired wound healing in diabetic mice by ganglioside GM3 synthase knockdown. Copyright 2015. Proc Nat Acad Sci USA [20]. Cy5 cyanine 5, NS-SNA nonsense SNA, GM3S ganglioside GM3 synthase, GM3S-SNA SNAs targeting GM3S, NS-SNA nonsense SNA, PBS phosphate buffer saline, SNA spherical nucleic acid

Fig. 5

Topically applied SNAs for treatment of psoriasis. a Timedependent uptake of Cy5-labeled liposomal SNAs in NHEK cells. Nuclei are blue (DAPI). Scale bars = 20 µm. b Cy5-L-SNAs diluted to 30 µM in Aquaphor/PBS (1:1) and applied to normal or psoriatic human skin explants in lifted cultures are visible throughout the samples after 24 h incubation. Scale bars = 50 µm. c, d Psoriatic skin samples exposed to no treatment, vehicle, L-SNAs containing scrambled RNA (Scr + L-SNA), or L-SNAs containing TNF-α-targeted RNA (TNF + L-SNA) were assessed clinically by modified PASI score. All data shown in c are with imiquimod treatment, with modified PASI scores for untreated mice all equal to 0. Reprinted with permission from Lewandowski et al. Topically delivered tumor necrosis factor-α-targeted gene regulation for psoriasis. Copyright 2017. J Investigative Dermatol [51]. Cy5 cyanine 5, IMQ imiquimod, L-SNAs liposomal SNAs, NHEK cells normal human epidermal keratinocyte cells, NT no treatment, PASI Psoriasis Area Severity Index, PBS phosphate buffer saline, Scr scrambled RNA, SNA spherical nucleic acid, TNF-α tumor necrosis factor–alpha, Veh vehicle

Fig. 6

IS-SNAs demonstrate increased potency in vitro and reduce tumor growth in mice bearing lymphoma tumors. a Significance of the orientation of CpG ODNs in IS L-SNA function. Comparison of IS L-SNAs made with CpG 7909 TLR9 agonists on their exterior to liposomes made with CpG 7909 on their interior, with or without an exterior coating of d-A₂₀ all-PO ODNs. The external presentation of CpG 7909 ODNs in L-SNAs is critical to their superior performance. b Tumor growth curve of mice bearing subcutaneous E.G7-OVA lymphoma tumors treated with either liposomal IS-SNAs containing tocopherol-modified CpG ODNs and OVA antigens, or with freely delivered OVA alone or in combination with CpGs. The experimental design is shown above the figure. Reprinted with permission from Radovic-Moreno et al. Immunomodulatory spherical nucleic acids. Copyright 2015. Proc Nat Acad Sci USA [68]. CpG ODNs short synthetic single-stranded DNA molecules containing unmethylated CpG dinucleotides, E.G7-OVA mouse lymphoma cell line engineered to overexpressed ovalbumin protein, IS-SNA immunostimulatory SNA, L-SNA liposomal SNA, ODN oligodeoxynucleotides, OVA ovalbumin, PBS phosphate buffer saline, PO phosphodiester, SEAP secreted alkaline phosphatase inducible by NF-κB, SNA spherical nucleic acid

Fig. 7

Scheme of various nanostructures that utilize the nucleic acid ‘shell’ of SNA architectures to facilitate cell uptake and drug delivery. a Drugs such as doxorubicin (DOX) can be loaded within polymer nanoparticles that are then coated with amine-terminated oligonucleotides via EDS-NHS chemistry. Reprinted with permission from Banga et al. Drug-loaded polymeric spherical nucleic acids: enhancing colloidal stability and cellular uptake of polymeric nanoparticles through DNA surface-functionalization. Copyright 2017. American Chemical Society [73]. b Camptothecin (CPT) can be conjugated to DNA via azide bonds and these DNA–drug amphiphiles will selfassemble into DNA–drug nanostructures. Reprinted with permission from Tan et al. Light-triggered, self-immolative nucleic acid-drug nanostructures. Copyright 2015. American Chemical Society [74]. c DNA-PTX micelles can be produced by conjugating PTX to DNA via disulfide linker. Reprinted with permission from Tan et al. blurring the role of oligonucleotides: spherical nucleic acids as a drug delivery vehicle. Copyright 2016. American Chemical Society [35]. CPT camptothecin, DOX doxorubicin, PTX paclitaxel, SNA spherical nucleic acid