Development of human thioredoxin as a mucolytic therapy for Cystic Fibrosis

描述（由申请人提供）：阻塞性肺疾病（尤其是囊性纤维化（CF））的有效管理代表了一个显著未满足的医疗需求。这类疾病的特征是粘液增厚，导致肺功能受损。这种粘液屏障主要由与作为持续性肺部炎症和感染的结果而释放的核酸复合的蛋白质（粘蛋白）的交联链组成。在CF（一种影响美国约30，000名患者和全球约70，000名患者的常染色体隐性疾病）的情况下，目前设计用于增加粘液流动性和促进清除的疗法涉及使用高渗盐水吸入以稀释分泌物和使用DNA酶（α-脱氧核糖核酸酶，商标为“Pulmozyme”）消化粘液核酸。此外，一半的CF患者服用四种不同的药物，而大约四分之一的患者服用六种以上的药物，包括短效β受体激动剂，粘液溶解剂，抗生素以及吸入和鼻内给药。 皮质类固醇因此，非常需要通过开发与现有疗法和递送装置相容并且在更广泛百分比的CF患者中有效的制剂和递送方案来减轻治疗负担。OrPro Therapeutics，Inc.正在开发硫氧还蛋白，这是一类具有高功效和安全性的新型粘液溶解蛋白药物，其特异性破坏交联粘蛋白之间的巯基键。在实验室研究中，已证明还原型重组人硫氧还蛋白-1（rhTrx）是人CF痰液的高效净化剂。Trx是一种小的、稳定的非糖基化氧化还原蛋白，适用于在一系列原核和真核表达宿主中生产，并且不需要哺乳动物细胞表达。我们的总体目标是建立rhTrx的制剂策略，以支持该分子作为囊性纤维化的新的更有效的粘液溶解疗法的开发。这种疗法的关键组成部分是药物制剂，其实现将蛋白质维持在稳定的化学还原状态（对于其粘液溶解和抗蛋白酶功能至关重要）以及促进与现代高效气雾剂递送系统的相容性的目标，所述现代高效气雾剂递送系统最小化CF患者的治疗负担。对于第一阶段的应用，我们计划将重点放在实验室规模的纯化生产。 蛋白质（天然rhTrx对照和单个氨基酸突变变体r（Cys）hTrx，具有增强安全性和功效的潜力），这些蛋白质的物理表征，以顺序 以评估其化学性质和相对活性，并进行初始预制剂研究以评估硫氧还蛋白通过一系列雾化装置递送的适用性。如果实现了该项目的Ais，将建立对用于粘液溶解药物开发的二硫键还原概念的关键验证，从而实现II期项目，其目标是重点开发特定制剂，这些制剂将支持临床前研究，从而实现利用改进的患者友好型递送装置的潜在更安全和更有效的治疗方案的人体试验。 公共卫生相关性：对于囊性纤维化（CF）的安全和有效治疗仍然存在关键的未满足的需求，囊性纤维化是高加索人群中最常见的遗传性致死性疾病，在美国影响超过30，000人。我们的总体目标是开发一种新的和改进的CF粘液破坏（粘液溶解）疗法，该疗法非常有效，不会给CF患者带来过度的治疗负担。我们将通过开发吸入性人硫氧还蛋白的最佳制剂来实现这一目标，硫氧还蛋白是一种具有新作用机制的重组蛋白粘液溶解剂，与目前的治疗方法相比，它有可能使更广泛的CF患者受益。

DESCRIPTION (provided by applicant): Effective management of obstructive pulmonary diseases, especially Cystic Fibrosis (CF), represents a significant unmet medical need. Such diseases are characterized by thickened mucus that contributes to impaired lung function. This mucus barrier is composed primarily of cross-linked chains of proteins (mucins) complexed with nucleic acids released as a consequence of persistent lung inflammation and infection. In the case of CF, an autosomal recessive disease affecting approximately 30,000 patients in the US and 70,000 world-wide, current therapies designed to increase the fluidity of mucus and facilitate clearance involve the use of hypertonic saline inhalation to thin secretions and DNase enzymes (dornase alfa, trademarked as Pulmozyme") to digest mucus nucleic acids. In addition, half of patients with CF take four different medications, while about a quarter take upwards of six drugs including short-acting beta agonists, mucolytics, antibiotics, and inhaled and intranasal corticosteroids. There is consequently a great need to reduce treatment burden by developing formulations and delivery regimes that are compatible with existing therapies and delivery devices and that are effective across a broader percentage of CF patients. OrPro Therapeutics, Inc. is developing thioredoxins, a new class of mucolytic protein drugs with high efficacy and safety which specifically disrupt the thiol linkages between crosslinked mucin proteins. Reduced recombinant human thioredoxin-1 (rhTrx) has been demonstrated to be a highly potent liquefier of human CF sputum in laboratory studies. Trx is a small, stable non-glycosylated redox protein suitable for manufacturing in a range of prokaryotic and eukaryotic expression hosts and does not require mammalian cell expression. Our overall goal is to establish a formulation strategy for rhTrx that will support development of this molecule as a new and more efficacious mucolytic therapy for cystic fibrosis. A key component of such a therapy is a drug formulation that accomplishes the objectives of maintaining the protein in a stable, chemically reduced state (essential for its mucolytic and anti-protease functions) as well as facilitating compatibility wit modern, highly efficient aerosol delivery systems that minimize the treatment burden on CF patients. For this Phase I application, we plan to focus on laboratory-scale production of purified proteins (both native rhTrx control and a single amino-acid mutant variant, r(Cys)hTrx, having the potential to enhance safety and efficacy), physical characterization of these proteins in order to assess their chemical properties and relative activities, and initial preformulation studies to evaluate the suitability of thioredoxin for delivery via a range of nebulization devices. If the Ais of this project are achieved, key validation of the concept of disulfide reduction for mucolytic drug development will be established, enabling a Phase II project with the goal of focused development of specific formulations that will support preclinical studies leading to human trials of potentially safer and more efficacious treatment regimens that utilize improved, patient-friendly delivery devices. PUBLIC HEALTH RELEVANCE: There remains a critical unmet need for safe and effective treatment for Cystic Fibrosis (CF), the most common inherited lethal disease in Caucasian populations affecting more than 30,000 individuals in the United States. Our overall goal is to develop a new and improved mucus-disrupting (mucolytic) therapy for CF that is highly efficacious and which does not impose excessive treatment burdens on CF patients. We will accomplish this through development of optimal formulations of inhaled human thioredoxin, a recombinant protein mucolytic with a novel mechanism of action that has the potential to benefit a broader range of CF patients than current therapies.