EWS-FLI1 in embryonic hMSCs induces DNA damage and Ewing sarcoma tumorigenesis [ChIP-seq]

Ewing sarcoma (ES) is an aggressive bone and soft tissue neoplasm, unique to humans, characterized by EWSR1/ETS rearrangements and whose cellular origin remains unclear. Expression of EWS-FLI1 in pediatric human mesenchymal stem cells (hMSCs) induced a transcriptional profile more resembling ES than its expression in adult hMSCs, but did not confer tumorigenic capacity. We have isolated human mesenchymal stem-like cells (hMSLCs) from experimental human embryonic stem cell (hESC)-derived teratomas. EWS-FLI1 expression in hMSLCs results in the acquisition of an ES signature through its preferential binding to microsatellites of intergenic and intronic regions. In hMSLCs, EWS-FLI1 directly regulates BRCA1 expression, although oncogene-expressing cells show defects in DNA damage repair (DDR). Xenografting in mice of EWS-FLI1-transduced hMSLCs resulted in the formation of tumors expressing characteristic ES markers. In summary, EWS-FLI1 enforces an aberrant transcriptome and endows in vivo transforming capacity when expressed in an undifferentiated early embryonic hMSC. Our approach represents an innovative experimental method for understanding critical aspects of the biology of developmental tumors, from leukemia to sarcomas, in which few (even single) genetic alterations are able to transform a fetal stem cell. Overall design: human Mesenchymal Stem Like Cells (hMSLCs) line 1 was infected with flag-tagged EWS-FLI1 or with control lentivirus

尤因肉瘤（Ewing sarcoma, ES）是一种仅人类罹患的侵袭性骨与软组织肿瘤，以EWSR1/ETS基因重排为特征，其细胞起源至今尚未明确。在儿童人间充质干细胞（pediatric human mesenchymal stem cells, hMSCs）中表达EWS-FLI1时，其诱导产生的转录谱更贴近尤因肉瘤的特征，而在成人hMSCs中表达该基因则无法产生类似效果，且未赋予细胞致瘤能力。本研究从实验性人类胚胎干细胞（human embryonic stem cell, hESC）来源的畸胎瘤中分离得到人间充质干细胞样细胞（human mesenchymal stem-like cells, hMSLCs）。在hMSLCs中表达EWS-FLI1，可通过优先结合基因间区与内含子区的微卫星序列，使细胞获得尤因肉瘤特征性的转录谱。在hMSLCs中，EWS-FLI1可直接调控BRCA1的表达，即便表达该致癌基因的细胞存在DNA损伤修复（DNA damage repair, DDR）缺陷。将转导EWS-FLI1的hMSLCs进行小鼠异种移植，可形成表达尤因肉瘤特征性标志物的肿瘤。综上，当EWS-FLI1在未分化的早期胚胎hMSCs中表达时，可诱导异常转录组，并赋予细胞体内转化能力。本研究方法为理解从白血病到肉瘤等发育源性肿瘤的关键生物学特征提供了创新实验手段，此类肿瘤仅需少量（甚至单个）遗传改变即可使胎儿干细胞发生恶性转化。总体实验设计：将flag标签标记的EWS-FLI1或对照慢病毒感染人间充质干细胞样细胞系1（human Mesenchymal Stem Like Cells, hMSLCs line 1）。