3' tsRNAs: biologic function and pre-clinical targeting for treating human disease

摘要 在过去的十年里，我们表征和研究了各种tRNA衍生物的性质， 小RNA（TSRNA）。近年来，我们一直专注于一个类通常被称为 作为3 'tsRNA（来自成熟tRNAs的3'末端），因为它们是最不好的 但似乎在组织再生中起作用（例如，肝再生）和 包括癌症的过度增殖状态。在这里，我们计划建立潜在的目标， 用于治疗目的的3 'TSRNA。最近，我们确定了一种特定的RNA， 22 nt CAG-亮氨酸3 'tsRNA，当通过加入反义寡核苷酸下调时， 在快速分裂而非静止细胞中的寡核苷酸抑制核糖体生物合成。损失 这种特异性的tsRNA限制了至少一个核糖体的翻译（在延伸步骤）， 蛋白mRNA。这导致rRNA加工的阻断和快速细胞凋亡。在 相比之下，添加3 'tsRNA模拟物增加了细胞增殖，并可以补充 细胞中的核糖体生物合成缺陷。我们建议进一步鉴定其他3 'tsRNA-mRNA 相互作用，建立其生物学和分子功能，发展基因治疗 和寡核苷酸反义递送技术，以追求 在动物中操纵3 '-tsRNA。虽然tsRNA在许多组织中表达，但我们 将把这些研究集中在肝脏上，包括肝癌。这项工作将提供新的 健康人基因调控和细胞内稳态中3 'tsRNA功能相关信息 和疾病状态，以及建立其潜在的治疗价值， 临床前人异种移植鼠动物模型。在修改后的申请中，我们 删除了与筛选先前特定研究中概述的改良LNP相关的所有研究， 目标3，如审查者所建议的。

ABSTRACT Over the last decade we characterized and studied the properties of various tRNA derived small RNAs (tsRNAs). In recent years, we have focused on one class commonly referred to as 3’tsRNAs (derived from the 3’end of mature tRNAs) because they are the least well studied but appear to play a role in tissue regeneration (e.g., liver regeneration) and hyperproliferative states including cancer. Here we plan to establish the potential of targeting the 3’tsRNAs for therapeutic purposes. Recently, we established that one specific RNA, the 22nt CAG-Leucine 3’tsRNA, which when down regulated by the addition of antisense oligonucleotides in rapidly dividing but not quiescent cells inhibit ribosome biogenesis. Loss of this specific tsRNA limits the translation (at the elongation step) of at least one ribosomal protein mRNA. This results in a block in rRNA processing and rapid cellular apoptosis. In contrast, the addition of a 3’tsRNA mimic increases cellular proliferation and can complement the ribosome biogenesis defect in cells. We propose to further identify other 3’tsRNA-mRNA interactions and establish their biologic and molecular function and develop gene therapy and oligonucleotide antisense delivery technologies to pursue the therapeutic potential of manipulating 3’tsRNAs in animals. Although the tsRNAs are expressed in many tissues, we will focus these studies on the liver including liver cancer. This work will provide new information related to 3’tsRNA function in gene regulation and cellular homeostasis in health and disease states, as well as establish their potential therapeutic value by the proposed preclinical human xenotransplant murine animal models. In the amended application, we removed all the studies related to screening for improved LNPs outlined in previous specific aim 3 as suggested by the reviewers.