BET Inhibitors Target the SCLC-N subtype of Small Cell Lung Cancer by Blocking NEUROD1 Transactivation

Small cell lung cancer can be divided into several molecular subtypes based on the expression of four master transcription factors (ASCL1, NEUROD1, POU2F3, and YAP1). These master factors have not been directly druggable, and we hypothesized that targeting their transcriptional coactivator(s) could provide an alternative approach. Here, we identify that BET bromodomain proteins physically interact with NEUROD1 and function as transcriptional coactivators. Using CRISPR knockout and ChIP-seq, we demonstrate that NEUROD1 plays a critical role in defining the landscapes of BET bromodomain proteins in the SCLC genome. Targeting BET bromodomain proteins by BET inhibitors leads to broad suppression of the NEUROD1-target genes, especially those associated with superenhancers, and reduces SCLC growth in vitro and in vivo. LSAMP, a membrane protein in the IgLON family, was identified as one of the NEUROD1-target genes mediating BET inhibitor sensitivity in SCLC. Altogether, our study reveals that targeting transcriptional coactivators could be a novel approach to blocking the master transcription factors in SCLC for therapeutic purposes. The goal of this experiment was to characterize the genome-wide binding profiles of BET proteins, RPII, and H3K27Ac upon NEUORD1 knockout in H446 cells. Overall design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for NEUROD1, BRD2, BRD3, BRD4, histone H3K27Ac, and RNA polymerase II in H446 NEUROD1-KO cells (#297, cl.2-1) and control cells (NTgRNA cl.10).

小细胞肺癌（Small cell lung cancer, SCLC）可依据四种核心转录因子（ASCL1、NEUROD1、POU2F3及YAP1）的表达特征划分为多种分子亚型。这类核心转录因子目前尚无直接可成药的靶点，我们推测靶向其转录共激活因子可提供替代治疗策略。本研究证实BET溴结构域蛋白（BET bromodomain proteins）可与NEUROD1发生物理相互作用，并作为其转录共激活因子发挥功能。通过CRISPR基因敲除（CRISPR knockout）与染色质免疫沉淀测序（ChIP-seq），我们证明NEUROD1在定义SCLC基因组中BET溴结构域蛋白的结合图谱方面发挥关键作用。利用BET抑制剂靶向BET溴结构域蛋白，可广泛抑制NEUROD1靶基因的表达，尤其是与超级增强子（superenhancers）相关的靶基因，并在体外及体内抑制SCLC的增殖。IgLON家族膜蛋白LSAMP被鉴定为介导SCLC中BET抑制剂敏感性的NEUROD1靶基因之一。综上，本研究揭示靶向转录共激活因子可作为阻断SCLC核心转录因子的新型治疗手段。
本实验的研究目标为：解析H446细胞中NEUROD1敲除后，BET蛋白、RNA聚合酶II（RNA polymerase II, RPII）及组蛋白H3K27Ac的全基因组结合图谱。
实验整体设计：对H446 NEUROD1基因敲除细胞（#297，克隆2-1）及对照细胞（NTgRNA克隆10）开展NEUROD1、BRD2、BRD3、BRD4、组蛋白H3K27Ac及RNA聚合酶II的染色质免疫沉淀测序（ChIP-seq）。