Development of siRNA delivery methodologies using in vivo imaging models

DESCRIPTION (provided by applicant): RNA interference offers the potential of a novel therapeutic approach for treating skin disorders. The ability to design, screen and identify potent, selective and stable small interfering RNAs (siRNAs) is now relatively straightforward. Unfortunately, methodologies to deliver these potential therapeutics to appropriate cells (including skin cells) have not kept pace. The long-term goal of this project is to develop effective and efficient siRNA skin delivery technologies, facilitating translation of siRNA therapeutics to the clinic. In Phase I, we investigated and identified a number of animal models that readily allow monitoring (including in vivo whole animal imaging) of functional siRNA activity in skin keratinocytes, characterized the stability of siRNAs under a variety of conditions related to delivery and analyzed the distribution of fluorescently-tagged siRNAs in mouse and human skin. In Phase 2, we utilize innovative siRNA skin delivery technologies and monitor their effectiveness in the model systems developed in Phase 1. Furthermore, the development and application of a dual-axes confocal (DAC) intravital imaging system allows noninvasive monitoring of the effectiveness of functional siRNA in individual cells in real time by visualization of reporter gene expression in transgenic murine skin. To take full advantage of this imaging system, functional siRNAs that are effective in the mouse model systems will be labeled with near infrared dyes that allow visualization in mouse skin at the cellular level. These tools now afford the opportunity in Phase 2 to efficiently screen, develop and optimize delivery technologies in a way that has not been possible previously. PUBLIC HEALTH RELEVANCE: Despite the exciting discoveries of the underlying mutations responsible for a large number of genodermatoses, few if any novel clinical treatments have emerged. The purpose of this proposal is to harness the potential of RNA interference, utilizing short interfering RNAs (siRNAs) that have picomolar efficacy, single nucleotide specificity, and relatively low toxicity profiles to target mutant disease-causing genes, particularly for dominant negative genetic skin disorders. The largest remaining hurdle for bringing siRNAs to the clinic is an effective delivery platform. In Phase 1, we set up appropriate animal models for evaluating siRNA delivery and in Phase 2, we utilize these animal models to optimize existing genecream formulations to topically delivery the inhibitors. In theory, the platform technology developed should be applicable to a large number of the approximately 2,000 known monogenic skin disorders currently afflicting patients.