RVG-9R

Objectives: Tumour necrosis factor (TNF)-α secreted by macrophages and dendritic cells (DCs) plays a predominant role in arthritis. Our previous studies suggest that a small peptide, RVG-9R (29-aa peptide derived from the rabies virus glycoprotein, fused to 9R residues), can deliver small interfering RNA (siRNA) to macrophages and DCs. We therefore tested whether knockdown of TNF-α expression in macrophages and DCs by RVG-9R/bound siRNA targeting TNF-α reduces the severity of collagen antibody-induced arthritis (CAIA) in mice. Method: Arthritis was induced in mice by injecting a combination of antibodies to collagen followed by lipopolysaccharide (LPS) treatment. Mice were also injected with TNF-α siRNA complexed with RVG-9R peptide or an irrelevant peptide RVMAT-9R on days 1, 3, 5, and 7. As a positive control, dexamethasone was injected intravenously. Paw thickness was measured every 2 days and the mice were killed on day 10 for testing synovial TNF-α levels and histological analysis of joints. Results: In control mice, arthritis developed on day 4 and reached its peak between day 7 and day 9. Treatment with siTNF-α bound to RVG-9R, but not to RVMAT-9R, resulted in reducing paw thickness scores to the same level as dexamethasone treatment, associated with reduced TNF-α level in synovial fluid. Histological analysis of joints in the control RVMAT-9R/TNF-α siRNA-treated mice showed marked pannus formation and destruction of cartilage and subchondrial bone, as well as severe infiltration of inflammatory cells into the synovium. By contrast, the joint pathology was markedly reduced in RVG-9R/TNF-α siRNA-treated mice resembling the dexamethasone-treated mice. Conclusions: Suppression of TNF-α expression in macrophages and DCs by RVG-9R-mediated siRNA delivery could potentially be a clinically viable strategy for treatment of arthritis.

We designed a delivery system to obtain an efficient and optimal nose-to-brain transport of BACE1 siRNA, potentially useful in the treatment of Alzheimer's disease. We selected a cell-penetrating peptide, the short peptide derived from rabies virus glycoprotein known as RVG-9R, to increase the transcellular pathway in neuronal cells. The optimal molar ratio between RVG-9R and BACE1 siRNA was elucidated. The complex between the two was then encapsulated. We propose chitosan-coated and uncoated solid lipid nanoparticles (SLNs) as a nasal delivery system capable of exploiting both olfactory and trigeminal nerve pathways. The coating process had an effect on the zeta potential, obtaining positively-charged nanoparticles, and on siRNA protection. The positive charge of the coating formulation ensured mucoadhesiveness to the particles and also prolonged residence time in the nasal cavity. We studied the cellular transport of siRNA released from the SLNs using Caco-2 as a model of epithelial-like phenotypes. We found that siRNA permeates the monolayer to a greater extent when released from any of the studied formulations than from bare siRNA, and primarily from chitosan-coated SLNs.

A major impediment in the treatment of neurological diseases is the presence of the blood-brain barrier, which precludes the entry of therapeutic molecules from blood to brain. Here we show that a short peptide derived from rabies virus glycoprotein (RVG) enables the transvascular delivery of small interfering RNA (siRNA) to the brain. This 29-amino-acid peptide specifically binds to the acetylcholine receptor expressed by neuronal cells. To enable siRNA binding, a chimaeric peptide was synthesized by adding nonamer arginine residues at the carboxy terminus of RVG. This RVG-9R peptide was able to bind and transduce siRNA to neuronal cells in vitro, resulting in efficient gene silencing. After intravenous injection into mice, RVG-9R delivered siRNA to the neuronal cells, resulting in specific gene silencing within the brain. Furthermore, intravenous treatment with RVG-9R-bound antiviral siRNA afforded robust protection against fatal viral encephalitis in mice. Repeated administration of RVG-9R-bound siRNA did not induce inflammatory cytokines or anti-peptide antibodies. Thus, RVG-9R provides a safe and noninvasive approach for the delivery of siRNA and potentially other therapeutic molecules across the blood-brain barrier.