The effector mechanism of siRNA spherical nucleic acids

Abstract

Spherical nucleic acids (SNAs) are nanostructures formed by chemically conjugating short linear strands of oligonucleotides to a nanoparticle template. When made with modified small interfering RNA (siRNA) duplexes, SNAs act as single-entity transfection and gene silencing agents and have been used as lead therapeutic constructs in several disease models. However, the manner in which modified siRNA duplex strands that comprise the SNA lead to gene silencing is not understood. Herein, a systematic analysis of siRNA biochemistry involving SNAs shows that Dicer cleaves the modified siRNA duplex from the surface of the nanoparticle, and the liberated siRNA subsequently functions in a way that is dependent on the canonical RNA interference mechanism. By leveraging this understanding, a class of SNAs was chemically designed which increases the siRNA content by an order of magnitude through covalent attachment of each strand of the duplex. As a consequence of increased nucleic acid content, this nanostructure architecture exhibits less cell cytotoxicity than conventional SNAs without a decrease in siRNA activity.

Controlled Monofunctionalization of Molecular Spherical Nucleic Acids on a Buckminster Fullerene Core

“…This observation is consistent with the previous findings in which a retarded loading of siRNAs onto SNAs has been observed. 12 , 37 − 39 However, it is notable that the fully loaded SNA ( S4 + 12 equiv of RNA) was virtually stable below the physiological temperature and the observed “’premature’” partial denaturation occurred at a higher temperature. Titration of S4 with the same complementary RNA verified the correct stoichiometry of the melting profiles ( Figure 3 B; note, the concentration of S4 was determined according to the Alexa content).…”

“…Dicer is able to cleave siRNAs from SNAs and release them for the canonical RNA interference pathway. 12 Most of the reported SNAs are polydisperse structures. The polydispersity may be a shortcoming as the data shows a similar behavior, but still a population of particles lacking thorough characterization of the molecules exists, something that is readily accessible for covalent ON conjugates.…”

NAD(P)H:quinone oxidoreductase 1 determines radiosensitivity of triple negative breast cancer cells and is controlled by long non-coding RNA NEAT1

“…SNA gold nanoparticles composed of siRNA targeting the Bcl-2-like protein 12 sequences have been developed and used for early phase 1 clinical trial in patients with recurrent glioblastoma [49]. Using SNAs to carry siRNAs can overcome the issues related to cellular or even tissue delivery in the human body [50]. Furthermore, SNAs composed of NEAT1-targeting sequences can be used to examine their radiosensitizer potential or therapeutic effect on TNBCs.…”

Gene Regulation Using Spherical Nucleic Acids to Treat Skin Disorders

“…induced silencing complex (RISC) which guides it to the target mRNA [66,67]. siRNA-induced RNAi leads to cleavage of the mRNA by Argonaute of the RISC complex, whereas miRNA-induced RNAi either prevents translation or initiates mRNA deadenylation thus destabilizing it [66,68,69] (Figure 2b). Though exocytosis of SNAs from cells is known to occur [70], the exact mechanisms are poorly understood.…”

“…In the antisense pathway, DNA binds to its mRNA target and either prevents protein translation or attracts RNase H which recognizes DNA bound to mRNA and selectively degrades the mRNA leaving the antisense DNA intact. In the RNAi pathway, siRNA or miRNA from SNAs are cleaved from the surface of the SNA core by Dicer, and the antisense strand is loaded onto the RNA-induced silencing complex (RISC) which guides it to the target mRNA [66,67]. siRNA-induced RNAi leads to cleavage of the mRNA by Argonaute of the RISC complex, whereas miRNA-induced RNAi either prevents translation or initiates mRNA deadenylation thus destabilizing it [66,68,69] (Figure 2b).…”