Epigenetic reprogramming of malignant stem cells of the testes

描述（由申请人提供）：胚胎癌（EC）是睾丸生殖细胞肿瘤（tgct）的干细胞，具有多能性，与人类胚胎干细胞具有显著的遗传和生物学相似性。我们认为，EC可以被视为成功治疗的原型模型，恶性肿瘤干细胞。我们利用全局基因表达分析和独特的RA敏感和RA耐药人类EC系统来定义视黄酸（RA）介导的肿瘤细胞分化的分子机制，这导致了我们的假设，即EC致瘤性丧失的关键决定因素是表观遗传重编程。这一证据部分基于一组候选多能性基因，包括参与Hox基因抑制的多梳组基因PHC1/EDR1，我们发现该基因在谱系承诺窗口被RA快速下调。我们提供的证据表明，EDR1和其他多能性基因可能通过RA介导的主多能性因子Oct4的下调而受到RA的调控。有趣的是，许多这些ra下调的多能性基因位于12p染色体上，在tgct中均匀扩增。因此，我们提出正常原始生殖细胞向EC细胞的恶性转化可能涉及与染色质重构相关的“强制”多能性。待验证的假设是，ra诱导的干细胞更新丧失部分是由于EDR1的转录调节导致EC细胞的表观遗传重编程和谱系承诺。这项R21干细胞和癌症应用的目标是开始表征与致瘤性和治疗潜力相关的EC细胞中的组蛋白修饰。我们的重点是多梳复合体，因为已知该复合体的改变与多种人类癌症密切相关，并且我们已经证明RA下调了位于tgct中关键12p扩增子上的特定多梳基因EDR1。这将通过分子，遗传和细胞生物学的方法来完成以下具体目标。1. 定义RA敏感EC和RA抗性EC的polycomb特异性染色质修饰，并确定RA是否是调节polycomb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RR12基因EDR1在ra介导的胚胎癌干细胞再生的多梳特异性重编程中的作用。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（TGCTs）是为数不多的可以通过化疗治愈的实体肿瘤之一，即使是高度晚期，这表明TGCTs的自我更新的癌症干细胞可能是治疗的目标。完成这项拨款的目的将增加对tgct的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是可治愈的，而绝大多数晚期实体瘤是不可治愈的理解。这反过来可能会导致新的治疗方法，有效地针对肿瘤中的关键细胞，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.