Novel Therapeutics for Interstitial Lung Disease: Modified, Water-Soluble Caveolin-1 Scaffolding Domain Peptides with Improved Characteristics for Drug Development

我们的长期目标是改善治疗许多毁灭性疾病的有限选择 它们共同构成间质性肺疾病（ILD），其中最普遍的是硬皮病， 特发性肺纤维化。由于ILD影响那些试图保持活跃的中年晚期人群， 治疗将迅速增长。Caveolin-1是一个很有前途的纤维化疾病治疗靶点。促纤维化 小窝蛋白-1缺乏在细胞和小鼠模型中的作用被与其活性相当的肽抑制 位点（小窝蛋白-1支架结构域，CSD）。然而，CSD缺乏合适的药物药理学性质， 发展为了克服这个问题，我们开发了新的，修改版本的CSD。我们首先将CSD 三个亚区，发现它们都抑制博莱霉素诱导的肺纤维化。为了改善制药业， 生物学特性，然后我们修改CSD和每个亚区是水溶性的，并保护免受蛋白质， 溶解这种修饰增强了细胞对所有四种修饰肽的摄取，并增加了它们的能力， 在体外抑制几种纯化的激酶。到目前为止，我们只在体内测试了四种修饰肽中的一种， 它在抑制博来霉素诱导的肺纤维化方面具有显著的活性。这些初步研究证明， 支持我们的建议，从四个候选人中确定一个铅化合物，然后评估其Thera- 毒性指数（毒性和有益剂量之间的比率）。我们的研究和文献也表明， 与FDA批准的重磅炸弹药物尼达尼布相比， （品牌名Ofev）。总之，要进行药物开发，我们必须确定先导化合物。由于 由于其独特的药理和功能特点，我们必须对我们的四种药物进行并排比较， 修饰肽。1)使用两种模型系统选择先导化合物：全身博来霉素治疗和 异硫氰酸异黄绿素（FITC）处理。我们将选择一种先导化合物，然后展示其 特异性和活性，与对照肽（乱序Lead）、尼达尼布和吡非尼酮进行比较 （商标名Esbriet，一种FDA批准的ILD药物，据报道会影响caveolin-1水平）。肽将 皮下注射在治疗方案中，在诱导纤维化后7天开始。主要读数将是 肺功能和纤维化标志物定量、微血管渗漏和组织形态学。成功 定义为先导化合物对肺功能有害作用的抑制>50% 和>75%的对纤维化和微血管渗漏的有害作用。2)确定治疗指数 铅化合物。将确定先导化合物有益作用的剂量依赖性 使用高于和低于我们目前标准剂量的剂量。其毒性将在单次治疗中进行评价 使用1X、5X、25 X和125 X当前标准剂量的最大耐受剂量实验。我们将 如果治疗指数>50，则认为这些研究成功。总之，这些研究将提供 在ILD抑制和安全性方面均符合我们的成功标准的新型先导化合物。

Abstract Our long-term goal is to improve on the limited choices for treating the many devastating diseases that together constitute the Interstitial Lung Diseases (ILD), the most prevalent of which are scleroderma and Idiopathic Pulmonary Fibrosis. As ILD affects those in late middle age trying to remain active, the market for a treatment will grow rapidly. Caveolin-1 is a promising therapeutic target in fibrotic diseases. The profibrotic effects of caveolin-1 deficiency in cells and in mouse models is suppressed by a peptide equivalent to its active site (caveolin-1 scaffolding domain, CSD). However, CSD lacks suitable pharmacologic properties for drug development. To overcome this problem, we developed novel, modified versions of CSD. We first divided CSD into three subregions and found they all suppressed bleomycin-induced lung fibrosis. To improve the pharma- cological properties, we then modified CSD and each subregion to be water soluble and protected from proteo- lysis. This modification enhanced the uptake by cells of all four modified peptides and increased their ability to inhibit several purified kinases in vitro. We have so far tested only one of the four modified peptides in vivo and it was outstandingly active in inhibiting bleomycin-induced lung fibrosis. These initial studies justify and strongly support our proposal to identify a Lead Compound from among the four candidates, then evaluate its Thera- peutic Index (ratio between toxic and beneficial doses). Our studies and the literature also suggest that our peptides will be more effective and have fewer side effects than the FDA-approved blockbuster drug nintedanib (brand name Ofev). In summary, to proceed with drug development we must identify a Lead Compound. Due to their distinct pharmacological and functional features, we must do a side-by-side comparison of our four modified peptides. 1) Select a Lead Compound using two model systems: Systemic Bleomycin Treatment and Fluorescein Isothiocyanate (FITC) Treatment. We will choose a Lead Compound, then demonstrate its specificity and activity by comparing it to a control peptide (scrambled Lead), nintedanib, and pirfenidone (brand name Esbriet, an FDA-approved drug for ILD reported to affect caveolin-1 levels). Peptides will be delivered s.c. in a Therapeutic Protocol, beginning 7 days after fibrosis is induced. Primary Readouts will be lung function and quantification of fibrosis markers, microvascular leakage, and tissue morphology. Success will be defined as the suppression by the Lead Compound of >50% of the deleterious effect on lung function and >75% of the deleterious effect on fibrosis and microvascular leakage. 2) Determine the Therapeutic Index of the Lead Compound. The dose-dependence of the Lead Compound’s beneficial effects will be determined using doses above and below our current standard dose. Its toxicity will be evaluated in a Single-Treatment Maximum Tolerated Dose Experiment using 1X, 5X, 25X, and 125X our current standard dose. We will consider these studies to be a success if the Therapeutic Index is >50. In summary, these studies will provide a novel Lead Compound that meets our Criteria for Success, both in terms of suppression of ILD and of safety.