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

急性髓系白血病是德国最致命的血液系统恶性肿瘤，年龄校正后的死亡率几乎没有改善。治疗方案包括静止期细胞毒化疗，变异小，复发难治性疾病和高危人群采用骨髓移植。更有针对性、毒性更低的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、导致恶性转化的变化以及疾病起始细胞的依赖性是至关重要的里程碑。然而，无论是遗传学还是转录学分析都不能解决AML的所有转化机制，转录后或翻译后效应有望为理解和治疗癌症开辟新的途径。这些过程的关键参与者是RNA结合蛋白(RBPs)，它可以调节RNA的丰度并将其翻译成蛋白质。在以前的研究中，我们已经研究了表观遗传调节因子EZH2及其在癌症中的作用。这些研究揭示了EZH2对过量的限制性商业惯例的负调控，以及在致癌过程中EZH2丢失时它们的上调。因此，我们假设，通过EZH2介导的RBP调控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循环。了解EZH2-RBP轴的这些调节功能，分离主要的RBP介体，以及它们在健康和疾病中的特征，不仅将为协调细胞命运决定提供洞察力，还可能为癌症治疗配备新的杠杆。基于我们的初步数据，我们确定了一系列受EZH2调控的限制性商业惯例，这些限制性商业惯例对EZH2缺失有很强的(10倍)放松管制。在这项研究中，我们将通过在健康和恶性细胞(WP1)中应用功能获得(增加RBP水平)和功能丧失(耗尽RBP)来从我们的候选列表中识别与癌症相关的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(WP2)中定义与功能相关的细胞和分子功能。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)。揭示所提议的EZH2-RBP转录组，重塑前馈循环，并使其可用于靶向治疗，可能因此成为晚期癌症治疗和细胞再生的StepStone。

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.