Peptide Therapy for Pulmonary Arterial Hypertension

描述（由申请人提供）： 肺动脉高压（PH）是一种具有多种病因的毁灭性疾病，导致各种肺部和心脏疾病患者的高发病率和死亡率。PH的特征在于血管阻塞和血管收缩的可变存在，导致肺血管阻力持续增加、血管重塑、右心室（RV）衰竭和低心输出量。尽管其具有深远的临床后果，但过去25年的治疗进展已适度改善PH患者的生存率。联合使用现有药物以最大化临床获益是治疗PH的新兴策略。尽管RV衰竭是PH中常见的临床问题，但目前尚无RV特异性实验疗法。PH的主要生理机制之一表明内皮功能障碍，其导致肺循环中一氧化氮（NO）产生减少和NO/环鸟苷5'-磷酸（cGMP）依赖性血管舒张。或者，肺血管cGMP水平也可以通过抑制磷酸二酯酶5（PDE 5）来提高，以实现类似的反应。由于已显示PH患者的肺和RV中PDE 5表达增加，因此增加NO产生和PDE 5抑制的靶向努力可以抵消导致PH患者衰竭心脏中后负荷增加的血管收缩。我们已经鉴定了一种15个氨基酸（FKSRNSRWSCRIKSR）合成肽（P3），其增强内皮NO合酶（eNOS）的催化活性并抑制肺动脉内皮细胞以及肺动脉平滑肌细胞中cGMP特异性PDE 5。初步数据表明，P3刺激增加细胞内NO释放，抑制cGMP水解速率，并减弱肺动脉压（PAP）的增加，但在遥测植入的缺氧诱导的PH的清醒大鼠模型中监测的体循环动脉压（SAP）。基于这些初步数据，我们假设P3治疗以其作为NO释放PDE 5抑制剂的独特双重作用为PH的治疗提供了新的治疗方法。为了验证这一假设，这项研究的具体目标是：（I）鉴定与eNOS活性增加和NO产生相关的P3介导的分子事件，特别关注Ca 2+释放、信号传导模块[磷脂酰肌醇（PI）3-激酶（PI 3 K）、蛋白激酶B（PKB或Akt）、蛋白激酶A（PKA）和Src酪氨酸激酶（SrcK）]、小窝蛋白-1（Cav-1）和/或eNOS的磷酸化和Cav-1/eNOS解离，（II）阐明P3介导的PDE 5/cGMP水解的抑制是否与Ca 2+释放、PI 3 K的活化、蛋白激酶G（PKG）/PKA信号传导、PDE 5的磷酸化和/或PDE 5的cGMP结合的抑制相关，和（III）使用生理学相关的缺氧诱导的PH动物模型确定P3的治疗潜力，特别关注P3治疗对进行性和消退性肺血流动力学调节的生物利用度、给药方案、功效和毒性的评估（增加的PAP、SAP、心输出量/心脏指数、血细胞比容和RV肥大）和与PH相关的血管重塑以及毒性。在临床前动物模型中确认这种新型肽的NO释放PDE 5抑制剂功能的基于机制的生理方法对于治疗PH的I期临床试验的进展是创新的。 公共卫生相关性： 与退伍军人健康相关：肺血管疾病和高血压在VA患者人群中很常见，因为在该人群中吸烟相关慢性肺动脉高压的发生率很高。 阻塞性肺疾病（COPD）。肺损伤的血管并发症，包括PH，也经常在发展成成人呼吸窘迫综合征（ARDS）的战斗伤员中观察到。此外，退伍军人和现役人员暴露于多种危险因素，包括高海拔，药物，化学品，HIV感染，血吸虫病，可导致肺损伤和PH的进行性发展。识别基于机制的独特治疗剂与NO释放PDE 5抑制功能提供了一种新的策略治疗PH。

DESCRIPTION (provided by applicant): Pulmonary arterial hypertension (PH) is a devastating disease of diverse etiology that contributes to high morbidity and mortality of patients with various lung and heart diseases. PH is characterized by vascular obstruction and variable presence of vasoconstriction, leading to a sustained increased pulmonary vascular resistance, vascular remodeling, right ventricular (RV) failure, and low cardiac output. Despite its profound clinical consequences, therapeutic advances over the past 25 years have modestly improved survival of PH patients. The combined use of available drugs to maximize the clinical benefit is an emerging strategy for the treatment of PH. Although failing RV is a common clinical problem in PH, currently RV-specific experimental therapies are not available. One of the major physiologic mechanisms of PH suggests endothelial dysfunction that causes diminished nitric oxide (NO) production and NO/cyclic guanosine 5' monophosphate (cGMP)- dependent vasorelaxation in the pulmonary circulation. Alternatively, pulmonary vascular cGMP levels can also be elevated via inhibition of phosphodiesterase type 5 (PDE5) to achieve a similar response. Since PDE5 expression has been shown to be increased in the lungs and RV of PH patients, targeted efforts to increase NO production and PDE5 inhibition can counteract vasoconstriction that contributes to increased afterload in failing heart of PH patients. We have identified a fifteen amino acid (FKSRNSRWSCRIKSR) synthetic peptide (P3) that enhances the catalytic activity of endothelial NO synthase (eNOS) and inhibits cGMP-specific PDE5 in pulmonary artery endothelial cells as well as in pulmonary artery smooth muscle cells. Preliminary data indicate that P3 stimulation increases intracellular NO release, inhibits rate of cGMP hydrolysis, and attenuates increase in pulmonary artery pressure (PAP) but not systemic artery pressure (SAP) monitored in telemetry implanted conscious rat model of hypoxia-induced PH. Based on these preliminary data, we hypothesize that P3 treatment with its unique dual action as NO releasing PDE5 inhibitor offers novel therapeutic approach for the treatment of PH. To test this hypothesis, the specific aims of this study are to: (I) identify P3-mediated molecular events associated with increased eNOS activity and NO production with specific focus on Ca2+ release, activation of signaling modules [phospatidylinositide (PI) 3-kinase (PI3K), protein kinase B (PKB or Akt), protein kinase A (PKA), and Src tysorine kinase (SrcK)], phosphorylation of caveolin-1 (Cav-1) and/or eNOS, and Cav-1/eNOS dissociation, (II) elucidate whether P3 mediated inhibition of PDE5/cGMP hydrolysis is associated with Ca2+ release, activation of PI3K, protein kinase G (PKG)/PKA signaling, phosphorylation of PDE5, and/or inhibition of cGMP binding of PDE5, and (III) determine therapeutic potential of P3 using a physiologically-relevant hypoxia-induced animal model of PH with specific focus on assessment of bioavailability, dosing regimen, efficacy, and toxicity of P3 therapy on the progressive and regressive pulmonary hemodynamic modulations (increased PAP, SAP, cardiac output/cardiac index, hematocrit, and RV hypertrophy) and vascular remodeling associated with PH as well as toxicity. Confirmation of the mechanism-based physiological approach for NO releasing PDE5 inhibitor function of this novel peptide in preclinical animal model is innovative for progression towards Phase I clinical trial for treatment of PH. PUBLIC HEALTH RELEVANCE: Relevance to Veterans Health: Pulmonary vascular disease and hypertension are common in the VA patient population because of the high incident in this population of cigarette smoking-related chronic obstructive pulmonary disease (COPD). Vascular complications of pulmonary injury, including PH, are also frequently observed in combat casualty victims who develop adult respiratory distress syndrome (ARDS). In addition, exposure of veterans and current active duty personnel to multiple risk factors including high altitude, drugs, chemicals, HIV infection, and schistosomiasis can cause lung injury and progressive development of PH. Identification of mechanism-based unique therapeutic agent with NO releasing PDE5 inhibitory function offers a novel strategy for treatment of PH.