Pathogenesis and Treatment of NUT midline carcinoma

项目摘要摘要。坚果癌(NC，前身为NUT中线癌)是一种侵袭性肿瘤 鳞癌，影响所有年龄段，具有高度(90%)致命性。这项提案的总体目标是 通过机制驱动的关键致癌靶点的识别来提高这些患者的存活率。NC IS 由NUTM1-融合癌基因定义，最常见的(~78%)编码BRD4-NUT蛋白。BRD4-螺母 通过阻止分化和维持增殖通过形成非常大的 (100kb-2MB)，我们描述的富含乙酰基组蛋白的超级增强子，称为巨蛋白(Megadomain，MD)。MDS出现 从NUT重新募集组蛋白乙酰转移酶(HAT)的p300到结合的乙酰化组蛋白 BET家族蛋白BRD4的双溴结构域。我们已经证明了BRD4-NUT驱动KEY转录 BRD4通过额外招募大量转录激活子来达到致癌靶点。 我们的证据表明，使用竞争性抑制结合的BET溴域抑制剂(Beti)治疗 BET溴结构域对染色质的抑制作用，可抑制NC在人体内的生长导致新领域的研究作用 BRD4在癌症中的作用。不幸的是，贝蒂单一疗法的疗效受到毒性的限制。 在下一阶段，我们正在研究新的致癌机制，其靶向可以与 贝蒂在治疗上。我们的初步数据，加上已经建立的数据，使我们制定了一个 NC增长如何由表观遗传驱动的机械双域模型。该模型将在AIMS进行测试 下面的总体目标是确定一种治疗组合，最大限度地优化 选择性、协同的肿瘤抑制作用，毒性最小。该模型提出，第一，转录 由PRC2形成的抑制结构域通过抑制促分化的表达而使NC生长， 肿瘤抑制基因(在AIM 1中测试)。 第二个物理上分离的允许MD由与乙酰化结合的BRD4-NUT-P300组成 驱动致癌基因转录的组蛋白。在该模型中，MD的形成需要协同绑定 BRD4-NUT溴结构域(Bd1和BD2)到乙酰化组蛋白(在Aim 2中测试)； 能够形成MD的组蛋白允许状态需要DOT1L对H3K79进行单甲基化(目标3)。 抑制这两个结构域将1.抑制致癌基因的转录，如myc；2.去... 抑制对分化、衰老和退出细胞周期至关重要的基因的转录。事实上，我们有 体外和体内研究发现，EZH2和BRD4-NUT具有协同抑制作用。 具体目的1.确定EZH2在NUT癌中的致癌作用。 特定目的2.确定BRD4溴结构域1和2在NUT癌中的致癌作用。 具体目的3.DOT1L在BRD4-NUT巨噬细胞的形成和致癌中的作用 功能呢？

Project Summary Abstract. NUT carcinoma (NC, formerly NUT midline carcinoma) is an aggressive squamous carcinoma, affecting all ages, that is highly (>90%) lethal. The over-arching goal of this proposal is to improve survival of these patients through mechanism-driven identification of key oncogenic targets. NC is defined by NUTM1-fusion oncogenes, most commonly (~78%) encoding the BRD4-NUT protein. BRD4-NUT drives NC growth through the blockade of differentiation and maintenance of proliferation by forming very large (100kb-2MB), acetyl-histone-rich super-enhancers called megadomains (MD) that we described. MDs arise from the recruitment of p300, a histone acetyl-transferase (HAT), by NUT to acetylated histones bound by the dual bromodomains of BRD4, a BET family protein. We have shown that BRD4-NUT drives transcription of key oncogenic targets through the additional recruitment of numerous transcriptional activators by BRD4. Our demonstration that treatment with BET bromodomain inhibitors (BETi) that competitively inhibit binding of BET bromodomains to chromatin, can inhibit growth of NC in humans led to a new field investigating the role of BRD4 in cancer. Unfortunately, the efficacy of BETi monotherapy is limited by toxicity. In this next phase, we are investigating novel oncogenic mechanisms whose targeting can synergize with BETi therapeutically. Our preliminary data, together with what is established, has led us to formulate a mechanistic two-domain model of how NC growth is epigenetically driven. The model will be tested in the aims below with the overall goal of identifying a therapeutic combination of compounds optimized for maximally selective, synergistic tumor inhibition with minimized toxicity. The model proposes that the first, transcriptionally repressive domain formed by PRC2 enables NC growth by repressing the expression of pro-differentiation, tumor suppressive genes (tested in aim 1). The second, physically separate permissive MD is comprised of BRD4-NUT-p300 bound to acetylated histones that drive transcription of oncogenic genes. In this model, MD formation requires cooperative binding of both BRD4-NUT bromodomains (BD1 and BD2) to acetylated histones (tested in aim 2); and the acetyl- histone-permissive state that enables MD formation requires mono-methylation of H3K79 by DOT1L (aim 3). Inhibition of both of these domains would 1. repress transcription of oncogenic genes, such as MYC, and 2. de- repress transcription of genes critical for differentiation, senescence, and exit from cell cycle. Indeed, we have found that co-inhibition of EZH2 and BRD4-NUT is synergistic in vitro and in vivo. Specific Aim 1. Determine the oncogenic role of EZH2 in NUT carcinoma. Specific Aim 2. Determine the oncogenic roles of BRD4 bromodomains 1 and 2 in NUT carcinoma. Specific Aim 3. What is the role of DOT1L in BRD4-NUT megadomain formation and oncogenic function?