Molecular Manipulation to Enhance Anti-Myeloma Response

多发性骨髓瘤(MM)是一种异质性疾病。尽管已经有了几个新的特工 和可用于治疗的组合，肿瘤细胞的基因组适应性导致它们 持续的生长和不良的生存结果。因此，有必要对小说进行鉴定 以沙子为靶点的治疗。这一方向的一个重要研究领域是 最近的研究进展突出了跨越的长非编码RNA(LncRNAs)的功能意义 人类基因组的80%。这些RNA分子通过以下途径控制各种细胞和分子功能 机制目前还没有得到很好的描述。在我们的初步调查中，我们利用了我们的RNA- 来自308名新诊断和统一治疗的患者的CD138+MM细胞的SEQ数据，以及16名 正常浆细胞，并描述了MM中异常的lncRNA景观。 与正常浆细胞相比，多发性骨髓瘤中差异表达的lncRNAs。我们鉴定了14个LncRNA 与PFS相关，并计算风险分数，对患者进行分层并报告他们的重要作用 作为临床结果的独立风险预测因子1。这些结果为进一步 研究LncRNA在MM中的生物学和分子活性。我们对913个表达的LncRNA进行了评估 在基于CRISPRi的初步CRISPR干扰(CRISPRi)筛查中用于对MM细胞活性的影响。一个主要的 在3个MM细胞系中筛选出20个影响MM细胞活性的LncRNA。使用RNA-SEQ进行评估 数据显示，这20个LncRNAs显著上调。值得注意的是，发现了特定的lncRNA 在基因定义的患者亚群中选择性上调，包括携带t(4；14)的高危MM。 二次筛选(最丰富或最贫乏的sgRNAs)确定的MIR17HG(RNA调节因子 在随后的实验中，作为MM中的一种新的lncRNA， 抑制RROL显著损害MM细胞的生长和存活。我们也 观察到新生脂肪生成的限速酶--乙酰辅酶A羧基酶1 (DNL)途径，是RROL的主要靶点之一。这一代谢途径将营养物质转化为 (葡萄糖、谷氨酰胺等)转化为用于储存能量或膜生物合成的脂肪酸 信号分子。我们还开始研究lncRNA RROL的抑制剂以及 ACc1，并观察其抗MM活性。基于这些观察结果，我们假设失控 LncRNAs通过控制多个基因，显著影响MM的病理生物学。 有可能成为治疗靶点。为了进一步了解LncRNAs在多发性骨髓瘤中的作用并确定 那些与进展相关的，并评估它们的治疗潜力，我们将识别功能 骨髓瘤中的lncRNA依赖性(目标1)，验证功能性lncRNA在多发性骨髓瘤中的作用(目标2)和 利用小分子和转基因技术评价MIR17HG(RROL)抑制剂及其下游靶点 MM中的操作(目标3)。这些研究将确定lncRNA的独特功能格局。 MM，并允许开发翻译应用程序。

Multiple myeloma (MM) is a heterogenous disease. Although there have been several novel agents and combinations available for treatment, the genomic adaptability of the tumor cells lead to their continued growth and and adverse survival outcome. Therefore there is need for identification of novel target sand directed therapies. One of the emrging areas of research in this direction has been the recent advances highlighting the functional significance of long noncoding RNAs (lncRNAs) that span > 80% of human genome. These RNA molecules control variety of cellular and molecular functions via mechanisms that are as yet not well described. In our preliminary investigation we utilized our RNA- seq data from CD138+ MM cells from 308 newly diagnosed and uniformly treated patients, and 16 normal plasma cells and described the aberrant lncRNA landscape in MM. We identified 869 differentially expressed lncRNAs in MM compared to normal plasma cells. We identified 14 lncRNAs associated with PFS and calculated a risk score that stratified patients and report their significant role as an independent risk predictor for clinical outcome1. These results provided the rationale to further investigate biological and molecular activity of lncRNA in MM. We evaluated 913 expressed lncRNAs for impact on MM cell viability in a preliminary CRISPR interference (CRISPRi)-based screen. A primary screen in 3 MM cell lines identified 20 lncRNAs impacting MM cell viability. Evaluation using RNA-seq data showed a significant upregulation of these 20 lncRNAs. Of note, specific lncRNAs were found selectively upregulated in genetically-defined patient subsets, including high-risk MM carrying t(4;14). A secondary screen (of the most enriched or depleted sgRNAs) identified MIR17HG (RNA Regulator of Lipogenesis; RROL), as one of the top hits as a novel lncRNA in MM. In subsequent experiments, suppression of RROL significantly impaired MM cell growth and survival in vitro and in vivo. We also observed that Acetyl Co-A Carboxylase 1 (ACC1), the rate-limiting enzyme of de novo lipogenesis (DNL) pathway, is one of the primary targets of RROL. This metabolic pathway converts nutrients (glucose, glutamine, etc.) into fatty acids serving for energy storage or biosynthesis of membranes and signaling molecules. We have also begun to investigate inhibitors of both lncRNA RROL as well as ACC1 and observed anti-MM activity. Based on these observations, we hypothesize that dysregulated lncRNAs significantly impact the pathobiology of MM by their ability to control multiple genes, with potential to serve as therapeutic targets. To further understand the role of lncRNAs in MM and identify those associated with progression, and to evaluate their therapeutic potential, we will identify functional lncRNA dependencies in myeloma (Aim 1), validate the role of functional lncRNAs in MM (Aim 2) and evaluate inhibitors of MIR17HG (RROL) and its downstream target using small molecule and transgenic manipulations in MM (Aim 3). These studies will define the unique functional landscape of lncRNA in MM and allow development of translational applications.