Cardioprotective mechanisms of novel noncoding RNA in myocardial infarction

项目总结/摘要 该提案旨在了解一种新的非编码RNA（ncRNA）实体TY 4的心脏保护作用， 它是由外来体的内容物所启发的。我们的重点是一类小的ncRNA，而不是以前 被认为与心脏病特别相关。Y RNA吸引了我们的注意力， 由YRNA 4基因编码的外泌体在由心球来源的细胞分泌的外泌体中异常丰富。 主要的这种ncRNA，EV-YF 1，在其自身递送时具有有益的性质：EV-YF 1增加 IL-10分泌，具有心脏保护和抗肥大作用。在这里，我们提出了TY 4的初步数据，一个新的 由EV-YF 1生物启发的化学实体（NCE）。TY 4发挥涉及炎症的整体转录组学变化 和纤维化途径，并在心力衰竭的小鼠模型中发挥疾病修饰生物活性， 射血分数、Duchenne肌营养不良和MI。值得注意的是，TY 4与母乳一起配制时， 蛋白质酪蛋白，在所有三种模型中口服起作用。在机制方面，初步数据显示TY 4结合至 TPR是一种核孔复合物蛋白，作为信使RNA从细胞核输出的看门人。我们 还发现巨噬细胞消耗消除TY 4介导的心脏保护。作为一个小的化学- 修饰的突变体NCE，TY 4具有合成RNA药物中所需的关键特征。但是其他RNA药物， FDA批准的或正在开发的，都有已知的作用机制：它们是反义寡核苷酸，小 干扰RNA、适体或miR-修饰剂。TY 4不适合这些类，因此是 新型RNA药物这里的重点是了解TY 4限制梗死面积的机制。 我们将检验以下机制假设：TY 4通过与TPR结合而对MI具有心脏保护作用， 改变靶细胞（特别是巨噬细胞）中的基因表达。我们将追求以下目标：1： 研究TY 4改变巨噬细胞中基因表达的机制。次级目标旨在 鉴定TY 4结合配偶体，表征TY 4诱导的转录组学变化， 进一步审讯的目标2：优化口服TY 4在鼠MI中的体内功效。次级目标旨在 优化TY 4在急性MI中的口服剂量模式，测量口服递送后TY 4在小鼠中的生物分布 急性MI前和后，确定TY 4在急性MI中MI后疗效的机会窗口，试验TY 4 在老年小鼠中的心脏保护功效，并表征优化的口服TY 4递送在老年小鼠中的作用。 MI模型。3：测试巨噬细胞是否是解释TY 4在急性MI中的功效所必需的和充分的。 子目标旨在比较MI结局+/-TY 4伴和不伴既往巨噬细胞耗竭，探索相对 组织驻留与循环巨噬细胞的作用，并确定是否MI后过继转移的前 体内TY 4-调节的巨噬细胞减少巨噬细胞耗尽小鼠的梗死面积。虽然高度机械化， 我们的建议，集中在一个合成的ncRNA NCE生物启发外泌体货物，也提供了翻译价值 在其开发一种新的心脏保护剂，即使在再灌注后给予也是有效的。

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.