Oligotherapeutics to enhance CFTR correction

项目总结 囊性纤维化(CF)是一种致命的肺部疾病，每3500名儿童中就有一人患病。F508del CFTR， 最常见的突变存在于90%的CF患者中，已被证明难以治疗。两个人 FDA批准的药物Orkambi®(Lumacaftor和iVacaftor)和Symdeko®(tezacaftor和 IVacaftor)，仅能改善2-4%的肺功能。改进F508del CFTR仍然是一个关键 慢性充血性心力衰竭治疗中未满足的需求。 F508del CFTR校正的一个障碍是遗传修饰物的表达增加 肺和呼吸道上皮细胞表达转化生长因子-β。转化生长因子-β抑制cftr功能并使 Orkambi®和Symdeko®的优势。我们实验室发现microRNA miR-145 介导转化生长因子-β抑制cftr校正。转化生长因子-β促进miR-145直接与 降解cftr转录本和减少蛋白质的3‘-非翻译区(3’-UTR) 表情。CFTR底物的丢失消除了治疗反应。MIR-145拮抗作用 克服这一障碍，提高Orkambi®的收益。 该项目寻求一种新的方法来增强F508del疗法。我们将利用一个 反义寡核苷酸(ASO)阻断miR-145与CFTR的结合。我们假设ASO- 定向的miR-145目标位置封锁提高了F508del CFTR的校正能力。 特定目标#1：通过反义选择性地增强F508del CFTR校正 寡核苷酸阻断miR-145结合位点。 特定目标2：测试反义寡核苷酸的体内传递、安全性和有效性 人源化CFTR小鼠miRNA靶向特异性阻断。 这些目标将为寡聚疗法的发展提供必要的“下一步”。目标1 提供体外机制、剂量和疗效数据，以建立 寡核苷酸靶向miRNA结合位点以改进F508del CFTR校正。目标2 利用最近开发的全长人源化CFTR小鼠来建立分娩 可行性、安全性和体内治疗反应。

PROJECT SUMMARY Cystic Fibrosis (CF) is a fatal lung disease that affects 1 in 3500 children. F508del CFTR, the most common mutation present in 90% of CF patients, has proven difficult to treat. The two FDA approved drugs, Orkambi® (lumacaftor and ivacaftor) and Symdeko® (tezacaftor and ivacaftor), only improve lung function by 2-4%. Improving F508del CFTR remains a critical unmet need in CF therapeutics. One barrier to F508del CFTR correction is increased expression of the genetic modifier TGF-β in CF lung and airway epithelia. TGF-β suppresses CFTR function and nullifies the benefit of Orkambi® and Symdeko®. Our laboratory has discovered that the microRNA miR-145 mediates TGF-β inhibition of CFTR correction. TGF-β increases miR-145 which directly binds to the 3'-untranslated region (3'-UTR) to degrade CFTR transcripts and diminish protein expression. Loss of CFTR substrate eliminates therapeutic response. miR-145 antagonism overcomes this barrier to improve Orkambi® benefit. The project pursues a novel approach to augment F508del therapeutics. We will utilize an antisense oligonucleotide (ASO) to block miR-145 binding to CFTR. We hypothesize that ASO- directed miR-145 target site blockade improves F508del CFTR correction. Specific Aim #1: To selectively augment F508del CFTR correction through antisense oligonucleotide blockade of the miR-145 binding site. Specific Aim #2: To test in vivo delivery, safety, and efficacy of antisense oligonucleotide miRNA target specific blockade in humanized CFTR mice. These Aims will provide the necessary “next steps” in oligotherapeutic development. Aim 1 provides the in vitro mechanism, dosing, and efficacy data to establish the rationale for oligonucleotide targeting of the miRNA binding site to improve F508del CFTR correction. Aim 2 utilizes the recently developed full-length humanized CFTR mouse to establish delivery feasibility, safety profile, and therapeutic response in vivo.