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）是一种常染色体隐性遗传疾病，由CF跨膜突变引起。 传导调节因子（CFTR）基因，其破坏CFTR的功能。大约90%的CF患者 具有至少一个F508 del CFTR拷贝。F508 del CFTR功能可以通过以下方式修复到显著程度： 在美国市场上的双重和三重组合药物。另外10%的CF患者则没有 有F508 del和他们的许多突变没有药物来恢复CFTR。本补助金的目的 是探索一类新的CFTR调节剂-前列腺素-激活野生型和 突变CFTR。我们的假设是鲁比前列酮和相关的前列酮激活多种形式的CFTR- 正常CFTR、F508 del CFTR或其他罕见突变，特别是那些已被拯救到细胞中的突变 表面由市售调制器。前列腺素被开发为氯离子通道2（CLCN 2）激活剂 但与存在于气道和胃肠道上皮中的受体介导的途径相互作用，以增强CFTR。 介导的氯化物分泌。选择的前列腺素可能是CFTR和CLCN 2两者的双重调节剂。 CFTR介导的氯离子转运是评估通道功效的有意义且可量化的措施。 校正剂、活化剂、增效剂和放大剂。校正器、稳定器和放大器通过增加 细胞表面CFTR蛋白水平，增强剂增加CFTR通道的开放概率 以增加阴离子渗透性。目前的激活剂，提高cAMP在体外，毛喉素和IBMX，不是临床上 安全或可用，这意味着市场上的双重和三重组合调制器依赖于 内源性cAMP水平。具体目标1：量化鲁比前列酮和其他新型CFTR的作用 非CF、F508 del CFTR和非F508 del突变CFTR表达气道中离子转运的激活剂 上皮细胞培养物。目的1A检验急性鲁比前列酮刺激CFTR介导的氯化物的假设 运输目的1B检验慢性暴露于CFTR激活剂招募CFTR功能， 刺激CLCN 2介导氯化物分泌。目的1C检验CFTR功能增加导致 增加气道表面液体深度。具体目标2：研究活化过程中的PGE 2信号通路 鲁比前列酮假设是一种或多种EP受体亚型是鲁比前列酮治疗期间的靶标。 野生型和突变型CFTR对氯化物分泌的激活。目标2A解决了气相化学还原偶联问题， 鲁比前列酮与CFTR的关系，并与鲁比前列酮对CLCN 2的活化不同。目标2B检验假设， EP 2和EP 4受体在与CFTR相同的细胞中表达。