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.

项目摘要/摘要 囊性纤维化（CF）是由CF跨膜突变引起的常染色体隐性疾病 电导调节剂（CFTR）基因破坏了CFTR的功能。大约90％的CF患者 至少具有F508DEL CFTR的副本。 F508DEL CFTR功能可以在很大程度上通过 美国市场上的双重组合药物。其他10％的患有CF的人没有 具有F508DEL，其许多突变没有恢复CFTR的药物。这笔赠款的目标 是探索新的CFTR调制器（Prostone）的效率，以激活野生型和 突变CFTR。我们的假设是，脂肪动酮和相关前列腺激活了许多形式的CFTR- 正常的CFTR，F508DEL CFTR或其他罕见突变，尤其是对细胞响应的突变 由市场调节器表面表面。前列腺是作为氯化物通道2（CLCN2）激活剂开发的 但是与气道和胃肠道上皮中存在的受体介导的途径相互作用，以提高CFTR- 介导的氯化物分泌。选择的前列腺可能是CFTR和CLCN2的双调节剂。 CFTR介导的氯化物转运是评估通道效率的有意义且可量化的措施 校正器，激活器，电势器和放大器。校正器，稳定器和放大器通过增加而起作用 细胞表面上的CFTR蛋白水平，潜在主义者增加了CFTR通道的开放概率 增加阴离子的渗透性。当前在体外升高营地的激活剂，福斯科林和IBMX在临床上都不是 安全或可用，这意味着市场上的双重和三重组合调制器正在依靠 内源性营地。特定目的1：量化脂肪支酮和其他新型CFTR的影响 在非CF，F508DEL CFTR和非F508DEL突变体CFTR的气道中的离子传输的激活剂 上皮细胞培养。 AIM 1A检验了急性脂肪支酮刺激CFTR介导的氯化物的假设 运输。 AIM 1B检验了以下假设：长期暴露于CFTR激活剂募集CFTR功能，并且可以 刺激CLCN2介导的氯化物分泌。 AIM 1C检验CFTR功能引导的假设 增加气道表面液体深度。特定目标2：在激活过程中研究PGE2信号通路 与lubiprostone。假设是一种或多种EP受体亚型是lubiprostone的靶标。 野生型和突变体CFTR激活氯化物分泌。 AIM 2A解决了GPCR耦合 lubiprostone to CFTR，并与ClCN2的脂蛋白激活区分开。 AIM 2B检验以下假设 EP2和EP4受体在与CFTR相同的细胞中表达。