Development of algorithm for identification of functional self-delivering RNAi co

描述（由申请人提供）：用转染试剂将小干扰rna （sirna）引入细胞可实现有效的基因沉默。不幸的是，尽管基于sirna的功能基因组学在体外被广泛应用，但将该技术应用于原代细胞和体内靶标验证的能力受到了有效和无毒递送系统的限制。我们与RXi制药公司合作，开发了一类新型共价修饰的RNAi化合物，不需要递送载体进入细胞，与传统sirna相比，其药理学性能得到了改善。我们称这些化合物为“自我传递RNA”或sdRNA。sdRNA是一种疏水修饰的RNAi-反义杂交种，已被证明在体外原代细胞和体内局部给药时具有很高的疗效。在包括真皮、肌肉、肿瘤、肺泡巨噬细胞、脊髓、视网膜等组织中，已证实其具有强大的吸收和/或无毒性的沉默作用。在真皮层和视网膜中，以5g剂量皮内和玻璃体内注射sdRNA可诱导有效和持久的沉默。RXi已经利用广泛的筛选成功地鉴定了针对许多靶标的功能性sdRNA化合物，并正在积极采购其中一些化合物用于临床开发。与传统sirna相比，sdRNA作为一种优越的功能基因组学工具，在原代细胞和体内实现RNAi商业化的一个主要障碍是相对较低的命中率。虽然需要筛选每个基因的大量序列并不是治疗应用的限制因素，但它严重限制了sdRNA技术在功能基因组学中的适用性，在功能基因组学中，需要针对数千个基因进行成本有效的化合物选择。本课题旨在通过对sdRNA的结构、化学、靶向位置、序列偏好等方面的优化，建立sdRNA效价预测算法。该项目的完成将为开发第一批基于sdrna的商业化产品奠定基础，这将作为第二阶段提案的一部分进行。这些在所有细胞类型和体内都有活性的sdRNA试剂的可用性将使功能基因组学和靶标分层/验证研究成为可能，并对药物发现过程产生重大影响。

DESCRIPTION (provided by applicant): Introduction of small interfering RNAs (siRNAs) into cells with transfection reagents results in efficient gene silencing. Unfortunately, while siRNA-based functional genomics is widely used in vitro, the ability to apply this technology to primary cells and in vivo target validation has been impeded limited availability of efficient and non-toxic delivery systems. We, in collaboration with RXi Pharmaceuticals, have developed a novel class of covalently modified RNAi compounds that do not require a delivery vehicle to enter cells and have improved pharmacology compared to traditional siRNAs. We term these compounds "self- delivering RNA" or sdRNA. sdRNA is a hydrophobically modified RNAi- antisense hybrid, which has been demonstrated to be highly efficacious in vitro in primary cells and in vivo upon local administration. Robust uptake and/or silencing without toxicity has been demonstrated in several tissues including dermal, muscle, tumors, alveolar macrophages, spinal cord, retina etc. In dermal layer and retina, intradermal and intravitreal injections of sdRNA at 5g doses induce potent and long lasting silencing. RXi has utilized extensive screening to successfully identify functional sdRNA compounds against a number of targets and are actively procuring some of these compounds toward clinical development. A major hurdle to commercialization of sdRNA as a superior functional genomics tool, enabling RNAi in primary cells and in vivo, is the relatively low hit rate as compared to conventional siRNAs. While the need to screen large number of sequences per gene is not a limiting factor for therapeutic applications, it severely limits the applicability of sdRNA technology to functional genomics, where cost effective compound selection against thousands of genes is required. The objective of this proposal is to optimize sdRNA structure, chemistry, targeting position, sequence preferences to develop an algorithm for sdRNA potency prediction. Completion of this project will create a foundation for developing the first line of sdRNAs based products for commercialization, which will be done as part of phase II proposal. Availability of these sdRNA reagents that are active in all cell types and in vivo will enable functional genomics and target stratification/validation studies and have a major impact on the drug discovery process. PUBLIC HEALTH RELEVANCE: The utility of RNAi technology is severely limited by limited availability of efficient delivery of RNAi compounds to primary cells and in vivo. We have recently developed a new class of RNAi compounds, "self- delivering rxRNA" or sdRNA. sdRNA is a hydrophobically modified RNAi- antisense hybrid, which has been demonstrated to be highly efficacious in vitro in primary cells and in vivo upon local administration. Current proposal will focus on further optimizing this new class of compounds to enable rapid design and synthesis.