AntimiR-199a for cardiac ischemia and failure

描述（由申请人提供）：保护心脏免受缺血或缺血/再灌注损伤是医疗保健的首要目标之一。目前，最有效的机制之一是缺血预处理，这是通过球囊膨胀间歇性闭塞冠状动脉来实现的。然而，不幸的是，它的临床应用受到其有效性是短暂的这一事实的阻碍，因此，需要事先知道缺血发作的时间。此外，它是一种侵入性技术，有动脉粥样硬化栓塞的风险。因此，识别能够模拟这一过程的因素将对缺血性心脏病大有裨益。microrna （miRNA）是一种转录后调节因子，在细胞的命运中起着重要作用。虽然该领域的大多数研究都集中在参与心脏器官发生和发病机制的miRNA上，但我们已经将注意力转向了参与缺血性损伤保护的miRNA。我们最近发现miR-199a是缺氧诱导因子1 α (Hif- 1a)和Sirt1的强制性直接调节剂，这两个关键分子分别是细胞对抗缺氧和长寿所必需的。缺氧诱导miR-199a下调，这是Hif-1a和Sirt1上调所必需的。通过在培养的肌细胞或转基因小鼠模型中敲除miR-199a，我们能够概括缺氧预处理的作用，并保护细胞免受缺氧损伤。调节miRNA在体内的表达以获得治疗优势仍然是一个挑战。在本研究的第一阶段，我们将在小鼠体内试验不同形式、剂量、持续时间和递送方法的anti - mir -199a，以确定其在降低内源性miRNA-199a水平和上调其靶标方面的功效和特异性。这些实验是第二阶段的前奏，第二阶段将测试其在保护心脏免受缺血性损伤方面的有效性。总体目标是评估antiir -199a作为缺血性心脏预防和预处理措施的潜力，从而确定拟议研发的科学价值和可行性。我们的假设是，1)将anti - miR-199a全身递送到心脏会诱导内源性miR-199a的下调，从而导致包括心脏在内的所有相关器官中Hif-1a和Sirt1的上调，2)注射的anti - miR-199a寡核苷酸的不同修饰将影响其在心脏中敲低内源性miR-199a所需的有效性、剂量和持续时间。3)局部递送的anti - miR-199a将有效地敲低miR-199a并诱导其靶点在注射心肌及其他部位上调，但在其他器官中不起作用。我们的目的是在小鼠中测试不同形式、剂量、持续时间和递送方法的anti - mir -199a，以确定其在降低内源性miRNA-199a水平和上调其靶标方面的功效和特异性。

DESCRIPTION (provided by applicant): Protecting the heart against ischemia or ischemia/reperfusion injury is one of the top health care objectives. Currently, one of the most effective mechanisms is ischemia preconditioning, which is achieved by intermittent occlusion of the coronary artery via balloon inflation. Unfortunately, though, its clinical applicability is hampered by the fact that its effectiveness is short-lived and, thus, requires prior knowledge of the time of an ischemic episode. Moreover, it is an invasive technique that carries the risk of atherosclerotic emboli. Thus, identification of factors that can mimic this process would be a great benefit to ischemic heart disease. MicroRNAs (miRNA) are posttranscriptional regulators that play a fundamental role in a cell's fate. While most of the studies in the field have focused on miRNA that are involved in cardiac organogenesis and pathogenesis, we have turned our attention to miRNA that are involved in protection against ischemic injury. We have recently identified miR-199a as an obligatory direct regulator of hypoxia-inducible factor 1 alpha (Hif- 1a) and Sirt1, two key molecules that are required for preconditioning of cells against hypoxia, and longevity, respectively. Hypoxia induces downregulation of miR-199a, which is required for upregulation of Hif-1a and Sirt1. By knocking down miR-199a in cultured myocytes or in a transgenic mouse model we were able to recapitulate the effects of hypoxia preconditioning and protect the cells against hypoxic damage. Modulating the expression of miRNA in vivo for therapeutic advantages remains a challenge. In phase I of this study we will experiment with different forms, doses, duration, and methods of delivery, of antimiR-199a in mice to establish its efficacy and specificity in reducing the levels of endogenous miRNA-199a and upregulation of its targets. These experiments are a prelude to phase II that will involve testing its effectiveness in protection of the heart against ischemic damage. The overall objective is to evaluate the potential of antimiR-199a delivery to the heart as a, preventive, preconditioning measure for the ischemic heart and, thus, establish the scientific merit and feasibility of the proposed R&D. Our hypotheses are, 1) systemic delivery of antimiR-199a to the heart will induce downregulation of endogenous miR-199a that will result in upregulation of Hif-1a and Sirt1 in all organs involved, including the heart, 2) the different modifications of the injected antimiR-199a oligo will impact its effectiveness, dose, and duration required for knockdown of endogenous miR-199a in the heart, and 3) locally delivered antimiR-199a will effectively knockdown miR-199a and induce upregulation of its targets in the injected myocardium and beyond, but not in any other organs. Our aim is to test different forms, doses, duration, and methods of delivery, of antimiR-199a in mice to establish its efficacy and specificity in reducing the levels of endogenous miRNA-199a and upregulation of its targets. PUBLIC HEALTH RELEVANCE: Protecting the heart against ischemia or ischemia/reperfusion injury is one of the top health care objectives. Currently, one of the most effective mechanisms is ischemia preconditioning, which is achieved by intermittent occlusion of the coronary artery via balloon inflation. Unfortunately, though, its clinical applicability is hampered by the fact that its effectiveness is short-lived and, thus, requires prior knowledge of the time of an ischemic episode. Moreover, it is an invasive technique that carries the risk of atherosclerotic emboli. Thus, identification of factors that can mimic this process would be a great benefit to ischemic heart disease. We have recently identified miR-199a that has the potential to fulfill that purpose, which will be investigated in this proposal.