In vivo imaging models for evaluating siRNA delivery to skin

描述（由申请人提供）：RNA干扰提供了治疗皮肤病的新治疗方法的潜力。设计、筛选和鉴定有效、选择性和稳定的小干扰RNA（siRNA）的技术现在相对简单。能够提供 这些对适当皮肤细胞的潜在治疗没有跟上步伐。该项目的长期目标是开发一种机制，以一种可以容易地转化为人类使用的方式，有效地将siRNA递送到适当的皮肤细胞。在第一阶段，我们调查了一些小鼠皮肤报告系统，其适用性监测siRNA交付角质形成细胞。这些模型中的几种使用报告基因，其可以在多个时间点使用体内免疫组织化学方法非侵入性地检测。 整体动物成像，而其他依赖于内源性基因靶标的实时PCR定量。还分析了具有或不具有导致疾病表型的突变角蛋白基因的人皮肤等同物作为将功能性siRNA递送至角质形成细胞的模型。此外，将在人和小鼠皮肤中测定siRNA的稳定性、半衰期和分布。第1阶段的目标非常简单;然而，这些报告系统对于为证明功能性siRNA的有效递送奠定基础至关重要。第二阶段，创新皮肤 在第1阶段确定的最合适的系统中筛选和优化所选siRNA的递送技术，并进行动物毒理学和药代动力学实验以准备临床试验。

DESCRIPTION (provided by applicant): RNA interference offers the potential of a novel therapeutic approach for treating skin disorders. The technology to design, screen and identify potent, selective, and stable small interfering RNAs (siRNAs) is now relatively straightforward. The ability to deliver these potential therapeutics to appropriate skin cells has not kept pace. The long-term goal of this project is to develop a mechanism to efficiently and effectively deliver siRNAs to appropriate skin cells in a way that can be readily translated to use in humans. In Phase I, we investigate a number of mouse skin reporter systems for their suitability to monitor siRNA delivery to keratinocytes. Several of these models use reporter genes that can be non-invasively detected at multiple timepoints using in vivo whole-animal imaging, while others rely on real-time PCR quantitation of endogenous gene targets. A human skin equivalent, with or without mutant keratin genes resulting in disease phenotype, is also analyzed as a model for delivery of functional siRNA to keratinocytes. In addition, the stability, half-lives, and distribution of siRNAs will be determined in both human and mouse skin. The proposed aims in Phase 1 are quite straightforward; however, these reporter systems are crucial for laying the groundwork for demonstrating effective delivery of functional siRNAs. In Phase 2, innovative skin delivery technologies for selected siRNAs are screened and optimized in the most appropriate systems identified in Phase 1, and animal toxicology and pharmacokinetic experiments are performed in preparation for clinical trials.