SWI/SNF complex loss facilitates gene silencing during NSCLC development

基因表达的表观遗传学改变在人类非小细胞肺癌的发生发展中起着重要作用。最近的研究表明，具有一个或多个表观遗传沉默基因的肿瘤患者的总体存活率往往较低。大多数报告都集中在解释表观遗传修饰的两个主要机制上，DNA甲基化和组蛋白修饰的改变。然而，核小体在基因启动子上的位置改变代表了另一种重要的机制，通过这种机制可以对基因进行表观遗传调控。事实上，现在已经有几项研究表明染色质重塑复合体与人类肿瘤发展过程中表观遗传沉默的发生有关。特别是，SWI/SNF(交配型开关/蔗糖非发酵)染色质重塑复合体似乎是导致NSCLC表观遗传学改变的有力候选者。最初在酵母中发现的SWI/SNF复合体通过依赖于ATP的过程重塑核小体来改变染色质结构。 SWI2ATPase同源物BRG1和BRM由于启动子甲基化和/或两个基因的突变而丢失或表达，在约25%的人非小细胞肺癌细胞系和约10%的原代人非小细胞肺癌细胞中发生。重要的是，包括我们自己在内的几个小组已经表明，基因表达的缺失与 BRG1/BRM与非小细胞肺癌患者预后不良有关。我们已发表的报告和初步结果表明，BRG1或BRM在缺陷的NSCLC细胞中的重新表达诱导了许多表观遗传沉默的基因的表达。因此，我们假设SWI/SNF复合体活性的丧失代表了非小细胞肺癌发育过程中基因沉默的一种新机制。为了验证这一假设，我们提出了一个使用细胞培养和基因工程小鼠模型的协同研究计划。具体地说，我们将确定在缺陷的非小细胞肺癌细胞系中重新表达后哪些基因启动子被BRG1和/或BRM激活，发现在非小细胞肺癌细胞系中BRG1表达缺失后经历沉默的基因启动子，并评估BRG1和/或BRM缺失在非小细胞肺癌基因工程小鼠模型中对肿瘤发展的影响。这些研究的成功完成将为深入研究非小细胞肺癌发育过程中的表观遗传沉默机制和SWI/SNF染色质重塑机制提供有价值的见解，并为进一步的基础和翻译研究提供一种新的基因工程动物模型。此外，如果DNMT或HDAC抑制剂不能有效逆转缺乏BRG1和BRM表达的NSCLC亚群中的基因沉默，它们可能需要一种新的治疗和预防进展的方法。通过确定这些肿瘤中发生的独特的染色质变化以及它们与BRG1和/或BRM阳性肿瘤的不同之处，我们可以启动合理的药物设计研究，以找到治疗这种致命疾病的试剂。

Epigenetic changes in gene expression play an important role in the development and progression of human non-small cell lung carcinoma (NSCLC). Recent studies have shown that patients with tumors possessing one or more epigenetically silenced genes often show a poorer overall survival. Most reports have focused upon two major mechanisms to account for epigenetic modifications, DNA methylation and alterations in histone modifications. However, altered nucleosome positioning at gene promoters represent another important mechanism by which genes can be epigenetically regulated. Indeed, several studies have now implicated chromatin-remodeling complexes in the genesis of epigenetic silencing in human tumor development. In particular, the SWI/SNF (mating type switch/sucrose nonfermenting) chromatin remodeling complex appears like a strong candidate for contributing to epigenetic alterations in NSCLC. Originally identified in yeast, the SWI/SNF complex alters chromatin structure by remodeling nucleosomes through an ATP-dependent process. Loss or expression of the SWI2 ATPase homologs, BRG1 and BRM, by promoter methylation and/or mutations of either or both genes occurs in ~25% of human NSCLC cell lines and ~10% of primary human NSCLCs. Importantly, several groups including our own have shown a correlation between loss of expression of BRG1/BRM and poor prognosis in NSCLC patients. Our published reports and preliminary results demonstrate that reexpression of BRG1 or BRM in deficient NSCLC cells induces expression of many epigenetically silenced genes. Therefore, we hypothesize that loss of SWI/SNF complex activity represents a novel mechanism for gene silencing during NSCLC development. To test this hypothesis, we propose a synergistic research plan using cell culture and genetically engineered mouse models. Specifically, we will determine which gene promoters are activated by BRG1 and/or BRM after reexpression in deficient NSCLC cell lines, discover gene promoters that undergo silencing after loss of BRG1 expression in NSCLC cell lines and assess the effects of BRG1 and/or BRM loss on tumor development in a genetically engineered mouse model for NSCLC. The successful completion of the proposed studies will provide valuable insights into the mechanisms of epigenetic silencing during NSCLC development and of SWI/SNF chromatin remodeling as well as generate a novel genetically engineered animal model for further basic and translational studies. Furthermore, if DNMT or HDAC inhibitors are not effective in reversing gene silencing in the subset of NSCLCs that lack BRG1 and BRM expression, they may require a novel approach for treatment and prevention of progression. By identifying the unique chromatin changes that occur in these tumors and how they differ from BRG1 and/or BRM-positive tumors, we can initiate rational drug design studies to find the reagents to treat this deadly disease.