Mechanism of Prostone Activation During CFTR Modulation

CFTR 调节过程中前列酮激活的机制

• 批准号: 10687435
• 负责人: Pamela L. Zeitlin
• 金额: $ 42.68万
• 依托单位: NATIONAL JEWISH HEALTH
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-19 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10687435
• 关键词: Acute; Address; Adrenergic Agonists; Adrenergic Receptor; Albuterol; Amplifiers; Anions; Automobile Driving; Cell Culture Techniques; Cell surface; Cells; Chloride Channels; Chlorides; Chronic; Combination Medication; Coupling; Cyclic AMP; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Data; Delta F508 mutation; Dinoprostone; Disease; Drug Targeting; EP4 receptor; Epithelial Cells; Exposure to; Forskolin; G-Protein-Coupled Receptors; Genes; Grant; Homeostasis; In Vitro; Inflammation; Inhalation; Ion Transport; Ions; Knowledge; Measurement; Measures; Mediating; Mutation; Pathway interactions; Permeability; Persons; Pharmaceutical Preparations; Probability; Prostaglandin E Receptor; Prostaglandins; Proteins; Protocols documentation; Regulator Genes; Secretory Cell; Signal Pathway; Signal Transduction; Surface; Testing; Treatment Efficacy; airway epithelium; airway surface liquid; base; beta-2 Adrenergic Receptors; cell bank; cell type; cystic fibrosis infection; cystic fibrosis patients; gastrointestinal epithelium; improved; mutant; novel; prospective; receptor; recruit; repaired; response; single-cell RNA sequencing; trafficking

PROJECT SUMMARY/ABSTRACT Cystic fibrosis (CF) is an autosomal recessive disease resulting from mutations in the CF transmembrane conductance regulator (CFTR) gene that disrupts the functions of CFTR. Approximately 90% of people with CF have at least one copy of F508del CFTR. F508del CFTR function can be repaired to a significant extent by double and triple combination medications on the market in the USA. The other 10% of people with CF do not have F508del and many of their mutations do not have a medication to restore CFTR. The GOAL of this grant is to explore the efficacy of a new class of CFTR modulator—a prostone—for the ability to activate wild-type and mutant CFTR. Our HYPOTHESIS is that lubiprostone and related prostones activate many forms of CFTR— normal CFTR, F508del CFTR, or other rare mutations, especially those that have been rescued to the cell surface by the marketed modulators. The prostones were developed as chloride channel 2 (CLCN2) activators but interact with receptor-mediated pathways present in airway and gastrointestinal epithelia to boost CFTR- mediated chloride secretion. It is likely that select prostones are dual modulators of both CFTR and CLCN2. CFTR-mediated chloride transport is a meaningful and quantifiable measure to assess efficacy of channel correctors, activators, potentiators, and amplifiers. Correctors, stabilizers, and amplifiers function by increasing CFTR protein levels on the cellular surface, and potentiators increase the open probability of the CFTR channel to increase anion permeability. Current activators that raise cAMP in vitro, forskolin and IBMX, are not clinically safe or available, which means that the double and triple combination modulators on the market are relying on endogenous levels of cAMP. Specific Aim 1: To quantify the effects of lubiprostone and other novel CFTR activators on ion transport in non-CF, F508del CFTR, and non-F508del mutant CFTR-expressing airway epithelial cell cultures. Aim 1A tests the hypothesis that acute lubiprostone stimulates CFTR-mediated chloride transport. Aim 1B tests the hypothesis that chronic exposure to CFTR activators recruits CFTR function and can stimulate CLCN2-mediated chloride secretion. Aim 1C tests the hypothesis that increased CFTR function leads to increased airway surface liquid depth. Specific Aim 2: To study the PGE2 signaling pathway during activation with lubiprostone. The hypothesis is that one or more EP receptor subtypes are target(s) of lubiprostone during activation of chloride secretion by wild-type and mutant CFTR. Aim 2A addresses GPCR coupling from lubiprostone to CFTR and differentiates from lubiprostone activation of CLCN2. Aim 2B tests the hypothesis that EP2 and EP4 receptors are expressed in the same cells as CFTR.

项目概要/摘要 囊性纤维化（CF）是一种常染色体隐性遗传疾病，由 CF 跨膜突变引起 大约 90% 的 CF 患者会破坏 CFTR 的功能。 至少有一份 F508del CFTR 功能可以通过 F508del CFTR 进行很大程度的修复。 美国市场上有双重和三重组合药物，另外 10% 的 CF 患者则没有。 患有 F508del 并且他们的许多突变没有药物可以恢复 CFTR 此项资助的目标。 的目的是探索一类新型 CFTR 调节剂（前石）的功效，以激活野生型和 我们的假设是鲁比前列酮和相关前列酮激活多种形式的 CFTR—— 正常 CFTR、F508del CFTR 或其他罕见突变，尤其是那些已被拯救到细胞中的突变 前列酮被开发为氯离子通道 2 (CLCN2) 激活剂。 但与气道和胃肠道上皮细胞中存在的受体介导途径相互作用，以增强 CFTR- 介导的氯离子分泌。选择的前石可能是 CFTR 和 CLCN2 的双重调节剂。 CFTR 介导的氯离子转运是评估通道功效的一种有意义且可量化的措施 校正器、激活器、增强器和放大器通过增加来发挥作用。 细胞表面的 CFTR 蛋白水平和增效剂增加 CFTR 通道的开放概率 目前在体外提高 cAMP 的激活剂，福司可林 (forskolin) 和 IBMX，尚未用于临床。 安全或可用，这意味着市场上的双组合和三组合调制器都依赖于 具体目标 1：量化鲁比前列酮和其他新型 CFTR 的作用。 非 CF、F508del CFTR 和非 F508del 突变体 CFTR 表达气道中离子转运的激活剂 目标 1A 测试急性鲁比前列酮刺激 CFTR 介导的氯的假设。 目标 1B 测试了以下假设：长期接触 CFTR 激活剂会募集 CFTR 功能，并且可以增强 CFTR 功能。 刺激 CLCN2 介导的氯离子分泌，目标 1C 测试了 CFTR 功能增加导致的假设。 增加气道表面液体深度 具体目标 2：研究激活过程中的 PGE2 信号通路。 假设一种或多种 EP 受体亚型是鲁比前列酮的靶标。 野生型和突变型 CFTR 激活氯离子分泌，Aim 2A 解决了 GPCR 偶联问题。 鲁比前列酮与 CFTR 的关系，并与 CLCN2 的鲁比前列酮激活区分开来，测试以下假设： EP2 和 EP4 受体与 CFTR 在同一细胞中表达。