Primordial germ cells identified as one potential cell of origin of MYC rhabdoid tumors

Rhabdoid tumors (RT) are rare and highly aggressive pediatric neoplasms. Their epigenetically-driven intertumoral heterogeneity is well described; however, the cellular origin of RT remains an enigma. Here, we established and characterized different genetically engineered mouse models driven under the control of distinct promoters and being active in early progenitor cell types with diverse embryonic onsets. From all models only Sox2-positive progenitor cells give rise to murine RT. Using single-cell analyses, we identified distinct cells of origin for the SHH and MYC subgroups of RT, rooting in early stages of embryogenesis. Intra- and extracranial MYC tumors harbor common genetic programs and potentially originate from fetal primordial germ cells (PGCs). Using PGC specific Smarcb1 knockout mouse models we validated that MYC RT originate from these progenitor cells. We uncovered an epigenetic imbalance in MYC tumors compared to PGCs being sustained by epigenetically-driven subpopulations. Importantly, treatments with the DNA demethylating agent decitabine successfully impaired tumor growth in vitro and in vivo. Overall design: We used a mouse model, Rosa26-creERT2::Smarcb1fl/fl , to profile the transcriptome of rhabdoid tumors of the MYC subgroup, by single cell RNA-sequencing (scRNA-seq). We crossed the Smarcb1 fl/fl strain with a mouse line harboring the CreERT2 coding region under the control of ubiquitous (Rosa26) promoter. Pregnant females were induced with a single intraperitoneal dose (50 mg/kg) of Tamoxifen at 6.5 days post-coitum (post-coital plug observation was considered as day 0.5). All mice were intensively monitored until neurological symptoms indicating tumor growth were observed. Tumors were dispersed into single cells and further processed for scRNA-seq (10X Genomics Platform).

横纹肌样瘤（Rhabdoid Tumor, RT）是一类罕见且极具侵袭性的儿童肿瘤。其表观遗传驱动的瘤间异质性已被充分阐明，但横纹肌样瘤的细胞起源仍是未解之谜。本研究构建并表征了多种基因工程小鼠模型，这些模型受不同启动子调控，且在具有不同胚胎发生起始阶段的早期祖细胞中具有活性。在所有模型中，仅Sox2阳性祖细胞可诱发小鼠横纹肌样瘤。通过单细胞分析，我们确定了横纹肌样瘤SHH亚型与MYC亚型各自独特的细胞起源，其溯源至胚胎发育早期阶段。颅内与颅外MYC亚型横纹肌样瘤具有共同的遗传程序，且可能起源于胎儿原始生殖细胞（Primordial Germ Cell, PGCs）。利用原始生殖细胞特异性Smarcb1敲除小鼠模型，我们验证了MYC亚型横纹肌样瘤确起源于此类祖细胞。我们发现，相较于原始生殖细胞，MYC亚型肿瘤存在表观遗传失衡，且该失衡由表观遗传驱动的亚群维持。值得注意的是，使用DNA去甲基化药物地西他滨（decitabine）处理可在体外与体内有效抑制肿瘤生长。

实验设计概述：我们采用Rosa26-creERT2::Smarcb1fl/fl小鼠模型，通过单细胞RNA测序（Single Cell RNA-Sequencing, scRNA-seq）对MYC亚型横纹肌样瘤的转录组进行分析。我们将Smarcb1fl/fl品系与携带受泛在性（Rosa26）启动子调控的CreERT2编码区的小鼠品系进行杂交。在妊娠第6.5天（以观察到交配栓记为妊娠0.5天），对孕鼠单次腹腔注射他莫昔芬（Tamoxifen），剂量为50 mg/kg。对所有小鼠进行密切监测，直至出现提示肿瘤生长的神经系统症状。分离肿瘤组织并制备单细胞悬液，随后通过10X Genomics平台进行单细胞RNA测序。