Epigenetic reprogramming of malignant stem cells of the testes

描述（申请人提供）：胚胎癌（EC）是睾丸生殖细胞肿瘤（TGCT）的干细胞，具有多能性，与人ES细胞具有显著的遗传和生物学相似性。我们认为，EC可以被看作是一个原型模型的成功治疗，恶性肿瘤干细胞。我们的努力，以确定维甲酸（RA）介导的肿瘤细胞分化的分子机制，使用全球基因表达分析和一个独特的系统RA敏感和RA耐药的人类EC导致我们的假设，即一个关键的决定因素损失的EC的致瘤性是表观遗传重编程。这一证据部分基于一组候选多能性基因，包括参与Hox基因抑制的polycomb组基因PHC 1/EDR 1，我们发现该基因在谱系定型窗口被RA快速下调。我们提供的证据表明，EDR 1和其他多能性基因可能通过RA介导的主多能性因子Oct 4的下调由RA调节。有趣的是，许多这些RA下调的多能性基因位于染色体12 p上，其在TGCT中均匀扩增。因此，我们认为正常原始生殖细胞向EC细胞的恶性转化可能涉及“强制性”多能性和相关的染色质重塑。待检验的假设是RA诱导的干细胞更新丧失部分是由于EDR 1的转录调节导致EC细胞的表观遗传重编程和谱系定型。该R21干细胞和癌症应用的目标是开始表征EC细胞中与致瘤性和治疗潜力相关的组蛋白修饰。我们的重点是多梳复合物，因为已知该复合物的改变与多种人类癌症密切相关，并且我们已经证明RA下调了特定的多梳基因EDR 1，该基因定位于TGCT中关键的12 p扩增子。这将通过分子、遗传和细胞生物学方法来实现，以完成以下具体目标。1.确定RA敏感EC和RA耐药EC的多梳特异性染色质修饰，并确定RA是否是调节多梳特异性抑制和癌症干细胞更新的主要形态发生信号。2.确定RR 12基因EDR 1在胚胎癌中RA介导的多梳特异性癌症干细胞更新重编程中的作用。这些研究很重要，因为人类EC独特的“生殖细胞样”表观遗传学可能与TGCT的可治愈性有关。睾丸生殖细胞肿瘤（TGCT）是少数几种可以用化疗治愈的实体肿瘤之一，即使是高度晚期，这表明TGCT的自我更新的癌症干细胞可能是治疗过程中的靶点。完成这项资助的目标将增加对TGCT遗传学和生物学进展的理解，并可能增加对为什么这些肿瘤如此可治愈的理解，而绝大多数晚期实体瘤则不然。这反过来可能导致新的疗法，有效地靶向肿瘤内的关键细胞，癌症干细胞。

DESCRIPTION (provided by applicant): Embryonal carcinoma (EC) are the stem cells of testicular germ cell tumors (TGCTs), are pluripotent, and share remarkable genetic and biologic similarity to human ES cells. We contend that EC can be seen as an archetypal model of successfully treated, malignant cancer stem cells. Our efforts to define the molecular mechanisms of retinoic acid (RA) mediated tumor cell differentiation using global gene expression analysis and a unique system of RA-sensitive and RA-resistant human EC has led to our hypothesis that a critical determinant of loss of tumorigenicity of EC is epigenetic reprogramming. This evidence is based in part on a set of candidate pluripotency genes, including a polycomb group gene involved in Hox gene repression, PHC1/EDR1, that we discovered to be rapidly downregulated by RA at the lineage commitment window. We provide evidence that EDR1 and other pluripotency genes may be regulated by RA via RA-mediated downregulation of the master pluripotency factor Oct4. Interestingly, many of these RA-downregulated pluripotency genes reside on chromosome 12p, which is uniformly amplified in TGCTs. Hence, we propose that the malignant conversion of the normal primordial germ cell to the EC cell may involve "enforced" pluripotency with associated chromatin remodeling. The hypothesis to be tested is that RA-induced loss of stem cell renewal is in part due to transcriptional modulation of EDR1 leading to epigenetic reprogramming and lineage commitment of EC cells. The goal of this R21 Stem Cells and Cancer application is to begin to characterize histone modifications in EC cells associated with tumorigenic and curative potential. Our focus is the polycomb complex since alterations in this complex are known to be closely associated with a variety of human cancers and since we have shown that RA downregulates a specific polycomb gene EDR1 that is localized to the critical 12p amplicon in TGCTs. This will be accomplished through molecular, genetic and cell biologic approaches to complete the following specific aims. 1. To define polycomb-specific chromatin modifications of RA-sensitive EC and RA-resistant EC and to determine whether RA is a major morphogenic signal regulating polycomb-specific repression and cancer stem cell renewal. 2. To determine the role of the RR12 gene, EDR1, in RA-mediated, polycomb-specific reprogramming of cancer stem cell renewal in embryonal carcinoma. These studies are important because the unique "germ cell-like" epigenetics of human EC may be linked to the curability of TGCTs. Testicular germ cell tumors (TGCTs) are one of the few solid tumors curable with chemotherapy even when highly advanced, suggesting that the self-renewing, cancer stem cells of TGCTs are likely targeted during therapy. Completion of the aims of this grant will increase understanding of the genetics and biologic progression of TGCTs and may increase understanding of why these tumors are so curable when the vast majority of advanced solid tumors are not. This in turn may lead to new therapies that effectively target crucial cells within tumors, cancer stem cells.