Spherical Nucleic Acids as a Divergent Platform for Synthesizing RNA–Nanoparticle Conjugates through Enzymatic Ligation

Herein, we describe a rapid, divergent method for using spherical nucleic acids (SNAs) as a universal platform for attaching RNA to DNA-modified nanoparticles using enzyme-mediated techniques. This approach provides a sequence-specific method for the covalent attachment of one or more <i>in vitro</i> transcribed RNAs to a universal SNA scaffold, regardless of RNA sequence. The RNA–nanoparticle constructs are shown to effectively knock down two different gene targets using a single, dual-ligated nanoparticle construct

Multiplexed Nanoflares: mRNA Detection in Live Cells

We report the development of the multiplexed nanoflare, a nanoparticle agent that is capable of simultaneously detecting two distinct mRNA targets inside a living cell. These probes are spherical nucleic acid (SNA) gold nanoparticle (Au NP) conjugates consisting of densely packed and highly oriented oligonucleotide sequences, many of which are hybridized to a reporter with a distinct fluorophore label and each complementary to its corresponding mRNA target. When multiplexed nanoflares are exposed to their targets, they provide a sequence specific signal in both extra- and intracellular environments. Importantly, one of the targets can be used as an internal control, improving detection by accounting for cell-to-cell variations in nanoparticle uptake and background. Compared to single-component nanoflares, these structures allow one to determine more precisely relative mRNA levels in individual cells, improving cell sorting and quantification

Ribozyme–Spherical Nucleic Acids

Ribozymes are highly structured RNA sequences that can be tailored to recognize and cleave specific stretches of mRNA. Their current therapeutic efficacy remains low due to their large size and structural instability compared to shorter therapeutically relevant RNA such as small interfering RNA (siRNA) and microRNA (miRNA). Herein, a synthetic strategy that makes use of the spherical nucleic acid (SNA) architecture to stabilize ribozymes and transfect them into live cells is reported. The properties of this novel ribozyme−SNA are characterized in the context of the targeted knockdown of O⁶-methylguanine-DNA methyltransferase (MGMT), a DNA repair protein involved in chemotherapeutic resistance of solid tumors, foremost glioblastoma multiforme (GBM). Data showing the direct cleavage of full-length <i>MGMT</i> mRNA, knockdown of MGMT protein, and increased sensitization of GBM cells to therapy-mediated apoptosis, independent of transfection agents, provide compelling evidence for the promising properties of this new chemical architecture