Discovery and characterization of EZH2-regulated RBP feed-forward mechanisms controlling cellular transformation

急性髓性白血病是德国最致命的造血恶性肿瘤，其年龄校正死亡率几乎没有改善。治疗方案由细胞毒性化疗组成，在安静的静态方案中变化很小，骨髓移植用于复发/难治性疾病和高危人群。更有针对性、毒性更小的治疗方案将是非常可取的。为了实现这一目标，了解AML的起源和功能，引起恶性转化的改变以及疾病启动细胞的依赖性是至关重要的里程碑。然而，AML的遗传和转录组学分析都不能解决所有的转化机制，转录后或翻译后效应有望为理解和治疗癌症开辟新的领域。这些过程的关键参与者是rna结合蛋白（rbp），它可以调节rna的丰度及其转化为蛋白质。在之前的研究中，我们研究了表观遗传调控因子EZH2及其在癌症中的作用。这些研究揭示了EZH2对过多rbp的负调控，以及在致癌过程中EZH2缺失时rbp的上调。因此，我们假设通过EZH2介导的RBP控制对转录组进行微调是调节细胞分化、再生和维持良性细胞状态的主要前馈回路。了解ezh2 -RBP轴的这些调节功能，分离主要的RBP介质，以及它们在健康和疾病中的特征，不仅将提供对细胞命运决定的编排的见解，而且可能为癌症治疗提供新的杠杆。根据我们的初步数据，我们确定了一系列受ezh2调控的rbp，这些rbp在ezh2缺失时具有强烈的（bb10倍）放松管制。在这项研究中，我们将通过在健康细胞和恶性细胞（WP1）中应用功能获得（增加RBP水平）和功能丧失（消耗RBP），从候选名单中确定与癌症相关的RBP。凭借在细胞和分子表征方面的成熟专业知识，将在健康细胞和恶性细胞中定义功能相关的细胞和分子功能（WP2）。这些数据将使我们能够开发和测试有针对性的、以rbp为中心的治疗方法，并将我们的发现转化为临床前模型（WP3）。揭示提出的EZH2-RBP转录组重塑前馈环并使其可用于靶向治疗，从而可能成为晚期癌症治疗和细胞再生的基石。

Acute myeloid leukemia is the most fatal hematopoietic malignancy in Germany with little improvements in age-corrected mortality. The treatment regimens are composed of cytotoxic chemotherapy in quiet static regimens with little variation, and bone marrow transplantation for recurrent/refractory disease and high-risk groups. More targeted –and less toxic- regimens would be highly desirable. Towards this goal, understanding the origin and function of AML, the alterations causing malignant conversion and the dependencies of the disease initiating cells are crucial milestones. However, neither the genetic nor the transcriptomic analyses of AML was able to resolve all mechanisms of transformation and post-transcriptional or post-translational effects are of high promise to open new venues to understand and treat cancer. Key players of these processes are RNA-binding-proteins (RBPs), which can modulate RNA-abundance and its translation into protein. In prior studies we have studied the epigenetic regulator EZH2 and its role in cancer. These studies unraveld the negative regulation of a plethora of RBPs by EZH2 and their upregulation upon loss of EZH2 during carcinogenesis. We thus hypothesize that finetuning of the transcriptome through EZH2 mediated RBP control is a major feed-forward loop regulating cellular differentiation, regeneration and maintenance of the benign cell state. Understanding these regulatory functions of the EZH2-RBP-axis, isolation of the major RBP mediators, and their characterization in health and disease will not only provide insights into the orchestration of cell fate decisions but may also equip cancer treatment with new levers. Based on our preliminary data, we identified a list of EZH2-regulated RBPs with strong (>10-fold) deregulation upon EZH2-loss. In this study, we will identify the cancer-relevant RBPs from our candidate list by applying gain-of-function (increasing RBP levels) and loss of function (depleting RBPs) in healthy and malignant cells (WP1). With proven expertise in cellular and molecular characterization, cellular and molecular function of the functionally relevant will be defined both in healthy and malignant cells (WP2). These data will enable us to develop and test targeted, RBP-centered therapeutic approaches and translate our findings to pre-clinical models (WP3). Uncovering the proposed EZH2-RBP transcriptome reshaping feed-forward loop and making it exploitable for targeted treatment may thereby become the stepstone for advanced cancer treatment and cellular regeneration.