EWS-WT1 fusion isoforms establish oncogenic programs and therapeutic vulnerabilities in Desmoplastic Small Round Cell Tumors.. EWS-WT1 fusion isoforms establish oncogenic programs and therapeutic vulnerabilities in Desmoplastic Small Round Cell Tumors.

EWS fusion oncoproteins underlie the pathogenesis of several human malignancies including Desmoplastic Small Round Cell Tumor (DSRCT), an aggressive mesenchymal tumor driven by fusions between the disordered domain of EWS and the developmental transcription factor WT1. Here we combined chromatin occupancy and long-range interaction profiles to identify EWS-WT1-dependent gene regulation networks and directly controlled target genes. We show that EWS-WT1 operates primarily as a powerful activator of distal regulatory elements and controls an oncogenic gene expression program that characterize primary DSRCTs. Moreover, EWS-WT1 has two isoforms that differ by three amino acids in their DNA binding domain (+/- KTS), as observed for wild type WT1, and we show that each fusion isoform has a specific DNA binding profile that is distinct from its wild type counterparts and requires a functional EWSR1 prion like domain. Remarkably, xenograft experiments using human mesothelial cells, candidate cells of origin of DSRCT, reveal that both isoforms are required to generate viable tumors that resemble DSRCT. Finally, we identify new candidate EWS-WT1 target genes with potential therapeutic implications, including CCND1, whose inhibition by the clinically-approved drug Palbociclib leads to marked tumor burden decrease in DSRCT PDXs in vivo. Taken together, our studies identify gene regulation programs and therapeutic vulnerabilities in DSRCT and provide a mechanistic understanding of the complex isoform-dependent oncogenic activity of EWS-WT1. Overall design: ChIP-seq,for,histone,marks,in,DSRCT.

EWS融合癌蛋白是多种人类恶性肿瘤致病过程的核心驱动因素，其中包括促结缔组织增生性小圆细胞肿瘤（Desmoplastic Small Round Cell Tumor, DSRCT）——该侵袭性间叶源性肿瘤由EWS的无序结构域与发育性转录因子WT1发生融合所驱动。本研究整合染色质占位分析与长距离染色质相互作用谱数据，鉴定得到EWS-WT1依赖的基因调控网络及其直接调控的靶基因。研究显示，EWS-WT1主要作为远端调控元件的强效激活因子，调控原发性DSRCT所特有的致癌基因表达程序。此外，与野生型WT1一致，EWS-WT1存在两种剪接异构体，二者在DNA结合域内相差3个氨基酸（±KTS）；本研究证实，两种融合异构体均具有与其野生型对应体截然不同的特异性DNA结合谱，且其功能发挥依赖于具有活性的EWSR1样朊病毒结构域。值得注意的是，以人类间皮细胞——DSRCT的候选起源细胞——开展的异种移植实验表明，两种异构体均为生成模拟DSRCT的活体肿瘤所必需。最后，本研究鉴定得到一批具有潜在治疗价值的新型EWS-WT1候选靶基因，其中包括CCND1；经临床获批药物帕博西尼（Palbociclib）抑制该基因后，可在体内显著降低DSRCT患者来源异种移植瘤（PDXs）的肿瘤负荷。综上，本研究明确了DSRCT中的基因调控程序与治疗易感靶点，并从机制层面阐明了EWS-WT1异构体依赖性的复杂致癌活性。整体实验设计：针对DSRCT细胞的组蛋白修饰标记开展ChIP-seq测序。