Cardioprotective mechanisms of novel noncoding RNA in myocardial infarction

项目摘要/摘要 这项提议试图了解一种新的非编码RNA(NcRNA)实体TY4的心脏保护作用。 这是受到外体内容的生物启发。我们的重点是一类小的ncRNA，而不是以前的 被公认为与心脏病特别相关的。Y RNA吸引了我们的注意力，因为分子 由YRNA4基因编码的蛋白在心球来源的细胞分泌的外切体中异常丰富。 这种主要的ncRNA，EV-YF1，当它自己递送时具有有益的特性：EV-YF1增加 分泌IL-10，具有心脏保护和抗肥大作用。在这里，我们提供了TY4的初步数据，这是一种新的 EV-YF1生物启发的化学实体(NCE)。TY4在全球转录水平上发生变化，涉及炎症 和纤维化途径，并在保存的心力衰竭小鼠模型中发挥疾病修改生物活性 射血分数、杜氏肌营养不良和心肌梗死。值得注意的是，当TY4与母乳一起配方时 蛋白质酪蛋白在所有三种模型中都有口服作用。在机制方面，初步数据显示，TY4结合到 TPR是一种核孔复合体蛋白，它充当信使RNA从细胞核输出的守门人。我们 他们还发现，巨噬细胞的耗尽破坏了TY4介导的心脏保护。作为一种小的化学物质- 修饰突变体NCE，TY4具有合成RNA药物所需的关键特征。但其他RNA药物，无论是 FDA批准的或正在开发中的所有药物都有已知的作用机制：它们是反义寡核苷酸，小 干扰RNA、适配子或miR修饰物。TY4不属于这些类，因此是 新型核糖核酸类药物。这里的重点是了解TY4限制梗塞范围的机制。 我们将检验以下机制假设：TY4通过与TPR和TPR结合而对心肌梗死具有心脏保护作用 改变靶细胞(特别是巨噬细胞)的基因表达。我们将追求以下目标：1. 探讨TY4改变巨噬细胞基因表达的机制(S)。子目标寻求 确定TY4结合伙伴，表征TY4诱导的转录变化，并优先考虑生物 进一步审讯的目标。2.优化TY4口服液对小鼠心肌梗死的体内疗效。子目标寻求 优化急性心肌梗死口服TY4给药方案，测定口服TY4在小鼠体内的生物分布 急性心肌梗死前后，确定急性心肌梗死后TY4疗效的机会窗，测试TY4 老年小鼠的心脏保护作用，以及优化口服TY4对慢性心肌梗死的影响 MI模型。3：测试巨噬细胞是否是解释急性心肌梗死TY4疗效的必要条件和充分条件。 子目标寻求比较+/-TY4在有和没有先前巨噬细胞耗竭的情况下的心肌梗死结果，探索相对的 组织驻留巨噬细胞与循环巨噬细胞的作用，并确定心肌梗死后是否过继转移EX 体内TY4条件下的巨噬细胞减少了巨噬细胞耗竭小鼠的梗塞面积。虽然高度机械化， 我们的建议，专注于合成ncRNA nce生物灵感来自外体货物，也提供了翻译价值 正在开发一种新的心脏保护剂，即使在再灌流后也是有效的。

PROJECT SUMMARY/ABSTRACT This proposal seeks to understand the cardioprotective effects of a new noncoding RNA (ncRNA) entity, TY4, that is bioinspired by the contents of exosomes. Our focus is on one class of small ncRNA, not previously recognized as particularly relevant to cardiac disease. Y RNAs captured our attention because molecules encoded by the YRNA4 gene are exceptionally plentiful in exosomes secreted by cardiosphere-derived cells. The predominant such ncRNA, EV-YF1, has salutary properties when delivered on its own: EV-YF1 increases IL-10 secretion, is cardioprotective and anti-hypertrophic. Here we present preliminary data on TY4, a new chemical entity (NCE) bioinspired by EV-YF1. TY4 exerts global transcriptomic changes involving inflammatory and fibrotic pathways, and exerts disease-modifying bioactivity in murine models of heart failure with preserved ejection fraction, Duchenne muscular dystrophy, and MI. Remarkably, TY4, when formulated with the breast milk protein casein, works orally in all three models. In terms of mechanism, preliminary data show that TY4 binds to TPR, a nuclear pore complex protein that acts as a gatekeeper for messenger RNA export from the nucleus. We have also found that macrophage depletion abrogates TY4-mediated cardioprotection. As a small chemically- modified mutant NCE, TY4 has key features desirable in a synthetic RNA drug. But other RNA drugs, whether FDA-approved or in development, all have known mechanisms of action: they are antisense oligos, small interfering RNAs, aptamers, or miR-modifiers. TY4 fits into none of those classes and thus is the prototype for a new class of RNA drugs. The focus here is on understanding the mechanism whereby TY4 limits infarct size. We will test the following mechanistic hypothesis: TY4 is cardioprotective against MI by binding to TPR and altering gene expression in target cells (and specifically, in macrophages). We will pursue the following Aims: 1: Investigate the mechanism(s) whereby TY4 alters gene expression in macrophages. Sub-aims seek to identify TY4 binding partners, characterize the transcriptomic changes induced by TY4, and prioritize biological targets for further interrogation. 2: Optimize in vivo efficacy of oral TY4 in murine MI. Sub-aims seek to optimize the dosage paradigm for oral TY4 in acute MI, measure biodistribution of TY4 after oral delivery in mice pre- and post- acute MI, determine the window of opportunity for TY4 efficacy post-MI in acute MI, test TY4 cardioprotective efficacy in aged mice, and characterize the effects of optimized oral TY4 delivery in a chronic MI model. 3: Test whether macrophages are necessary and sufficient to explain TY4 efficacy in acute MI. Sub-aims seek to compare MI outcomes +/- TY4 with and without prior macrophage depletion, probe the relative roles of tissue-resident versus circulating macrophages, and determine whether post-MI adoptive transfer of ex vivo TY4-conditioned macrophages reduces infarct size in macrophage-depleted mice. While highly mechanistic, our proposal, focusing on a synthetic ncRNA NCE bioinspired by exosomal cargo, also offers translational value in its development of a novel cardioprotective agent that is effective even when given after reperfusion.