Targeting TET DNA Dioxygenases as Therapeutic Principle in Myeloid Neoplasms

项目总结/摘要 TET 1、TET 2和TET 3是α-酮戊二酸（αKG）和Fe 2+依赖的DNA- 双加氧酶是表观遗传景观的关键调节因子。这些酶逐渐氧化5- 甲基胞嘧啶转化为5-羟甲基胞嘧啶，并进一步转化为5-甲酰基胞嘧啶和5-羧基胞嘧啶 最终导致DNA去甲基化，这对于大时间尺度上的有效转录至关重要。功能丧失 TET 2突变（TET 2 MT）是MDS最常见的致病性病变之一， 人类的恶性肿瘤TET 2 MT存在于所有疾病阶段和侵袭性水平中。此外，最近 研究表明，体细胞TET 2 MT非常频繁地发现于“健康”老年人中。的存在 CHIP中的TET 2 MT意味着它是血液系统疾病形成的早期事件。它还支持鼠 研究结论TET 2 MT引起造血干细胞和祖细胞（HSPC）的扩增。早期 缺陷克隆进化中的事件是预防策略的合理目标， 阶段可能取决于这些事件。然而，它们在所有细胞中的存在也可以被利用， 疾病的晚期。例如，小鼠造血细胞中TET 1和TE 2的联合缺失， 相对于TET 2单独损失，模型显著延长了寿命。此外，还推导了原理性证明 根据对IDH 1/2 MT患者的观察，这些新变体突变导致产生一种新的 弱泰特抑制剂，2-羟基谷氨酸（2-HG），已知的双加氧酶抑制剂。我们观察到IDH 1/2 MT 与TET 2 MT相互排斥，强烈支持我们的假设，即泰特抑制剂2-HG， TET 2 MT克隆的进化。这一观察结果在骨髓恶性肿瘤的细胞模型中得到了进一步的证实。 细胞和支持物，可以通过消除剩余的必需TET活性来实现合成致死性 用于在TET-双加氧酶缺陷型TET 2 MT克隆中有效转录增殖。 我们的总体假设是，合理设计和合成的小分子TETi 76可以被利用， 阻碍来源于TET 3和TET 1的补偿性泰特双加氧酶活性， 在TET 2 MT失活的情况下致死或谱系重定向。我们的目标是开发一种新的治疗方法 通过评估TETi 76在临床前模型中作为靶向治疗的潜在用途，研究TET 2 MT MDS的治疗方法。 更具体地说，我们的目标是：i）研究泰特抑制在正常和恶性肿瘤中的机械后果， 体外造血。ii）使用人和鼠细胞培养物在体外建立TETi化合物的作用。 iii）表征TETi预防和治疗功效以及在临床前 鼠模型。 我们的建议，如果成功，将导致一类新的治疗剂TET 2 MT相关的造血 疾病，也许还有其他双加氧酶突变。这项工作也可能对癌症有影响 在组蛋白去甲基化酶KDM 6A中突变，其类似于TET 2并且在膀胱癌中频繁突变。

PROJECT SUMMARY/ABSTRACT Ten Eleven Translocation (TET1, TET2 and TET3) are α-ketoglutarate (αKG) and Fe2+ dependent DNA- dioxygenases that is a key regulator of epigenetic landscape. These enzymes progressively oxidize 5- methylcytosine to 5-hydroxymethylcytosine and further to 5-formylcytosine and 5-carboxylcytosine in DNA culminating into DNA demethylation essential for efficient transcription over large time scales. Loss-of-function TET2 mutations (TET2MT), is one of the most frequent pathogenic lesion in MDS and related hematologic malignancies in humans. TET2MT are found in all disease stages and levels of aggressiveness. In addition, recent studies have demonstrated that somatic TET2MT are very frequently found in “healthy” elderly. The presence of TET2MT in CHIP implies that it is an early event in the creation of hematologic disorders. It also supports murine study conclusions that TET2MT cause expansions of hematopoietic stem and progenitor cells (HSPC). Early events in the evolution of defective clones are rational targets for preventative strategies since clones in such stages are likely to be dependent on these events. Their presence in all cells can, however, also be exploited in late stages of the disease. For example, the combined loss of TET1 and TE2 in hematopoietic cells in murine models extends life substantially relative to TET2 loss alone. In addition, the proof of principle is also derived from observation in patients with IDH1/2MT whereby, these neomorphic mutations lead to the production of a weak TET inhibitor, 2-hydroxyglutyrate (2-HG), known inhibitor of dioxygenases. Our observations that IDH1/2MT are mutually exclusive with TET2MT, strongly supports our hypotheses that the TET inhibitor, 2-HG, prevents evolution of TET2MT clones. This observation was further substantiated in a cellular model of myeloid malignant cells and supports that synthetic lethality can be achieved through elimination of remaining TET-activity essential for efficient transcription for proliferation in TET-dioxygenase deficient TET2MT clones. Our overarching hypothesis is that rationally designed and synthesized small molecules TETi76 can be utilized to impede compensatory TET dioxygenase activity originating from TET3 and TET1, to cause either synthetic lethality or lineage redirection in cases with TET2MT inactivation. Our goal is to develop a novel therapeutic approach for TET2MT MDS by evaluating the potential use of TETi76 as targeted treatments in preclinical models. More specifically, we aim to: i) Study the mechanistic consequences of TET inhibition in normal and malignant hematopoiesis in vitro. ii) Establish the effects of TETi compounds in vitro using human and murine cellular models and iii) Characterize TETi preventative and therapeutic efficacy as well as tolerability in pre-clinical murine models. Our proposal, if successful, will lead to a novel class of therapeutic agents for TET2MT associated hematopoietic disorders, and perhaps also other dioxygenase mutations. This work could also have implications for cancers mutated in histone demethylase KDM6A, which is similar to TET2 and frequently mutated in bladder cancer.