Department of Neurological Surgery, Northwestern University Feinberg School of Medicine, Chicago, Illinois.
Department of Neurological Surgery, Juntendo University, Tokyo, Japan.
Neuro Oncol. 2021 Aug 2;23(8):1348-1359. doi: 10.1093/neuonc/noab009.
Diffuse intrinsic pontine glioma (DIPG) is associated with transcriptional dysregulation driven by H3K27 mutation. The super elongation complex (SEC) is required for transcriptional elongation through release of RNA polymerase II (Pol II). Inhibition of transcription elongation by SEC disruption can be an effective therapeutic strategy of H3K27M-mutant DIPG. Here, we tested the effect of pharmacological disruption of the SEC in H3K27M-mutant DIPG to advance understanding of the molecular mechanism and as a new therapeutic strategy for DIPG.
Short hairpin RNAs (shRNAs) were used to suppress the expression of AF4/FMR2 4 (AFF4), a central SEC component, in H3K27M-mutant DIPG cells. A peptidomimetic lead compound KL-1 was used to disrupt a functional component of SEC. Cell viability assay, colony formation assay, and apoptosis assay were utilized to analyze the effects of KL-1 treatment. RNA- and ChIP-sequencing were used to determine the effects of KL-1 on gene expression and chromatin occupancy. We treated mice bearing H3K27M-mutant DIPG patient-derived xenografts (PDXs) with KL-1. Intracranial tumor growth was monitored by bioluminescence image and therapeutic response was evaluated by animal survival.
Depletion of AFF4 significantly reduced the cell growth of H3K27M-mutant DIPG. KL-1 increased genome-wide Pol II occupancy and suppressed transcription involving multiple cellular processes that promote cell proliferation and differentiation of DIPG. KL-1 treatment suppressed DIPG cell growth, increased apoptosis, and prolonged animal survival with H3K27M-mutant DIPG PDXs.
SEC disruption by KL-1 increased therapeutic benefit in vitro and in vivo, supporting a potential therapeutic activity of KL-1 in H3K27M-mutant DIPG.
弥漫性内在脑桥神经胶质瘤（DIPG）与 H3K27 突变驱动的转录失调有关。超级延伸复合物（SEC）是通过释放 RNA 聚合酶 II（Pol II）促进转录延伸所必需的。SEC 破坏导致转录延伸受阻，可能是 H3K27M 突变 DIPG 的一种有效治疗策略。在这里，我们测试了 SEC 药理学破坏对 H3K27M 突变 DIPG 的影响，以深入了解分子机制，并为 DIPG 提供新的治疗策略。
短发夹 RNA（shRNA）用于抑制 H3K27M 突变 DIPG 细胞中 AF4/FMR2 4（AFF4）这一 SEC 核心成分的表达。肽模拟物 KL-1 用于破坏 SEC 的一个功能成分。细胞活力测定、集落形成测定和凋亡测定用于分析 KL-1 处理的影响。RNA 和 ChIP-seq 用于确定 KL-1 对基因表达和染色质占据的影响。我们用 KL-1 治疗携带 H3K27M 突变的 DIPG 患者来源异种移植（PDX）的小鼠。通过生物发光图像监测颅内肿瘤生长，并通过动物存活评估治疗反应。
AFF4 的耗竭显著降低了 H3K27M 突变的 DIPG 细胞生长。KL-1 增加了全基因组 Pol II 占据，并抑制了涉及多个促进 DIPG 细胞增殖和分化的细胞过程的转录。KL-1 治疗抑制 DIPG 细胞生长、增加凋亡，并延长 H3K27M 突变 DIPG PDX 动物的存活。
KL-1 通过破坏 SEC 增加了体外和体内的治疗益处，支持 KL-1 在 H3K27M 突变 DIPG 中的潜在治疗活性。
