CRISPR/Cas-mediated development of an RNAi rat model system

摘要 药物开发的成本已经飙升到估计每种FDA批准的药物26亿美元，主要是 由于缺乏有效性或安全性而失败，这表明我们目前的临床前验证过程 在预测人类的治疗潜力和毒性方面还不够。动物模型是黄金 用于解剖疾病机制和评估体内新药靶点的标准;然而， 由于开发这些药物的准备时间很长，因此无法在药物发现过程中进行常规使用。随着近期 CRISPR/Cas9基因组编辑的发展，以及RNA干扰技术的进步，我们现在 有能力快速开发具有精确基因组修饰和类人疾病的动物模型 病理学我们已经证明RNAi可以作为基因缺失的快速替代方法， 在基因工程小鼠模型中评估新的治疗潜力和预测毒性， 基因靶点这项提案的目标是将我们的能力扩展到mice之外，并开发一个平台， 在短短4个月内快速和具有成本效益的RNAi大鼠生产。尽管小鼠模型的实用性， 许多科学家认为，大鼠仍然是首选的啮齿动物，因为它们的体积更大，便于手术操作， 重复血液采样，以及他们的认知和生理特征，更接近于 人类比他们的老鼠同行。对于神经生物学、心脏生物学、免疫学和毒理学， 仍然是研究中的主要啮齿动物模型。我们目前的客户群中有近30%的人询问过利率 在过去的5年里，他们的化合物的大多数毒理学研究仍然是在大鼠中进行的， I期临床试验我们知道老鼠将再次成为药物研究中最重要的啮齿动物模型 发现，因为我们已经与一家大型制药公司签订了初步试点计划， 这些模型。我们打算站在这一转变范式的最前沿，回到大鼠模型。使用小说在 CRISPR/Cas9试剂的原位递送方法，我们现在将能够快速操纵大鼠基因组。 和成本效益的方式，并系统地产生RNAi大鼠模型，允许时间和可逆的 基因调控来模拟治疗方案。这些RNAi大鼠模型将改变临床前验证 评估体内潜在的药物反应和耐药机制，最终指导 开发更安全、更有效的药物。

Abstract The cost of drug development has skyrocketed to an estimated $2.6B for every FDA approved drug primarily due to failures from lack of efficacy or safety, suggesting that our current preclinical validation process has been insufficient in predicting therapeutic potential and toxicity in humans. Animal models are the gold standard for dissecting disease mechanisms and evaluating novel drug targets in vivo; however, the cost and long lead time to develop them has prevented their routine use in the drug discovery process. With the recent developments in CRISPR/Cas9 genome editing, and advances in RNA interference technologies, we now have the ability to rapidly develop animal models with precise genomic modifications and human-like disease pathologies. We have shown that RNAi serves as a fast alternative to gene deletion and can also be used within genetically engineered mouse models to assess the therapeutic potential and predict toxicities of novel gene targets. The goal of this proposal is to expand our capabilities beyond mice and develop a platform for rapid and cost-effective production of RNAi rats in as little as 4 months. Despite the utility of mouse models, for many scientists, the rat still remains the preferred rodent due to their larger size for surgical manipulation, repeat blood sampling, and their cognitive and physiological characteristics that more closely resemble humans than their mouse counterparts. For neurobiology, cardiobiology, immunology and toxicology, they are still the dominant rodent model in research. Nearly 30% of our current client base has inquired about rat models over the last 5 years, noting that most toxicology studies of their compounds are still done in rats prior to Phase I clinical trials. We know rats will gain popularity once again as the premier rodent model in drug discovery, as we have already been contracted a large pharmaceutical company for an initial pilot program for these models. We intend to be at the forefront of this shifting paradigm back to the rat model. Using novel in situ delivery methods of CRISPR/Cas9 reagents, we will now be able to manipulate the rat genome in a rapid and cost-efficient manner and systematically generate RNAi rat models that allow for temporal and reversible gene regulation to simulate therapeutic regimes. These RNAi rat models will transform the preclinical validation process with assessment of potential drug response and resistance mechanisms in vivo, ultimately guiding the development of safer and more effective drugs.