The ability to downregulate specific genes using systemically delivered short RNA molecules and the cellular mechanism known as RNA interference has been shown previously in mouse and non-human primate models. Davis et al. have now demonstrated for the first time in humans that a short interfering RNA (siRNA) molecule can be systemically delivered using nanoparticles to a solid tumour. The siRNA mediates directed cleavage of its target mRNA, thereby also reducing the protein level. This proof-of-principle study confirms the potential of this technology as a human therapeutic. It has previously been shown in mice and non-human primates that systemically delivered short RNA molecules can inhibit gene expression. Here it is shown that a short interfering RNA (siRNA) can be systemically delivered, using nanoparticles, to a solid tumour in humans. The siRNA mediates cleavage of its target mRNA, thereby also reducing levels of the encoded protein. This proof-of-principle study confirms the potential of this technology for treating human disease. Therapeutics that are designed to engage RNA interference (RNAi) pathways have the potential to provide new, major ways of imparting therapy to patients1,2. Long, double-stranded RNAs were first shown to mediate RNAi in Caenorhabditis elegans3, and the potential use of RNAi for human therapy has been demonstrated by the finding that small interfering RNAs (siRNAs; approximately 21-base-pair double-stranded RNA) can elicit RNAi in mammalian cells without producing an interferon response4. We are at present conducting the first in-human phase I clinical trial involving the systemic administration of siRNA to patients with solid cancers using a targeted, nanoparticle delivery system. Here we provide evidence of inducing an RNAi mechanism of action in a human from the delivered siRNA. Tumour biopsies from melanoma patients obtained after treatment show the presence of intracellularly localized nanoparticles in amounts that correlate with dose levels of the nanoparticles administered (this is, to our knowledge, a first for systemically delivered nanoparticles of any kind). Furthermore, a reduction was found in both the specific messenger RNA (M2 subunit of ribonucleotide reductase (RRM2)) and the protein (RRM2) levels when compared to pre-dosing tissue. Most notably, we detect the presence of an mRNA fragment that demonstrates that siRNA-mediated mRNA cleavage occurs specifically at the site predicted for an RNAi mechanism from a patient who received the highest dose of the nanoparticles. Together, these data demonstrate that siRNA administered systemically to a human can produce a specific gene inhibition (reduction in mRNA and protein) by an RNAi mechanism of action.

PEGylated gold nanoparticles are decorated with various amounts of human transferrin (Tf) to give a series of Tf-targeted particles with near-constant size and electrokinetic potential. The effects of Tf content on nanoparticle tumor targeting were investigated in mice bearing s.c. Neuro2A tumors. Quantitative biodistributions of the nanoparticles 24 h after i.v. tail-vein injections show that the nanoparticle accumulations in the tumors and other organs are independent of Tf. However, the nanoparticle localizations within a particular organ are influenced by the Tf content. In tumor tissue, the content of targeting ligands significantly influences the number of nanoparticles localized within the cancer cells. In liver tissue, high Tf content leads to small amounts of the nanoparticles residing in hepatocytes, whereas most nanoparticles remain in nonparenchymal cells. These results suggest that targeted nanoparticles can provide greater intracellular delivery of therapeutic agents to the cancer cells within solid tumors than their nontargeted analogs.

Intracellular delivery of nucleic acids as gene regulation agents typically requires the use of cationic carriers or viral vectors, yet issues related to cellular toxicity or immune responses hamper their attractiveness as therapeutic candidates. The discovery that spherical nucleic acids (SNAs), polyanionic structures comprised of densely packed, highly oriented oligonucleotides covalently attached to the surface of nanoparticles, can effectively enter more than 50 different cell types presents a potential strategy for overcoming the limitations of conventional transfection agents. Unfortunately, little is known about the mechanism of endocytosis of SNAs, including the pathway of entry and specific proteins involved. Here, we demonstrate that the rapid cellular uptake kinetics and intracellular transport of SNAs stem from the arrangement of oligonucleotides into a 3D architecture, which supports their targeting of class A scavenger receptors and endocytosis via a lipid-raft–dependent, caveolae-mediated pathway. These results reinforce the notion that SNAs can serve as therapeutic payloads and targeting structures to engage biological pathways not readily accessible with linear oligonucleotides.

Nanoparticles are being investigated for numerous medical applications and are showing potential as an emerging class of carriers for drug delivery. Investigations on how the physicochemical properties (e.g., size, surface charge, shape, and density of targeting ligands) of nanoparticles enable their ability to overcome biological barriers and reach designated cellular destinations in sufficient amounts to elicit biological efficacy are of interest. Despite proven success in nanoparticle accumulation at cellular locations and occurrence of downstream therapeutic effects (e.g., target gene inhibition) in a selected few organs such as tumor and liver, reports on effective delivery of engineered nanoparticles to other organs still remain scarce. Here, we show that nanoparticles of ~75 ± 25-nm diameters target the mesangium of the kidney. These data show the effects of particle diameter on targeting the mesangium of the kidney. Because many diseases originate from this area of the kidney, our findings establish design criteria for constructing nanoparticle-based therapeutics for targeting diseases that involve the mesangium of the kidney.

Despite being engineered to avoid renal clearance, many cationic polymer (polycation)-based siRNA nanoparticles that are used for systemic delivery are rapidly eliminated from the circulation. Here, we show that a component of the renal filtration barrier—the glomerular basement membrane (GBM)—can disassemble cationic cyclodextrin-containing polymer (CDP)-based siRNA nanoparticles and, thereby, facilitate their rapid elimination from circulation. Using confocal and electron microscopies, positron emission tomography, and compartment modeling, we demonstrate that siRNA nanoparticles, but not free siRNA, accumulate and disassemble in the GBM. We also confirm that the siRNA nanoparticles do not disassemble in blood plasma in vitro and in vivo. This clearance mechanism may affect any nanoparticles that assemble primarily by electrostatic interactions between cationic delivery components and anionic nucleic acids (or other therapeutic entities).