Role of TOP2B in generating complex genomic rearrangements in human cancer

描述（由申请人提供）：尽管最近的研究表明复杂的基因组重排在前列腺癌的发生和发展中的重要性，但产生这些重排的机制尚不清楚。我们的长期目标是了解前列腺癌和其他癌症中这些复杂重排的原因和后果。这里的目的是确定多重、协调、双链断裂（DSB）可能参与产生这些复杂重排的机制。我们的中心假设是，肿瘤前列腺细胞中雄激素诱导的转录程序的启动涉及高度复杂拓扑环的形成，该拓扑环部分由拓扑异构酶II β (TOP2B)介导；这种酶的功能障碍可以同时产生多个断裂，这些断裂可以重新组合形成复杂的基因组后排。我们基于强有力的初步数据提出了这一假设，这些数据显示，在人类前列腺癌中，andro- gen2诱导的，top2b介导的DSB是重组的，并且出现在重排断点附近。这项工作的基本原理是，了解这些机制可以为预防这些重排的形成提供策略，潜在地允许癌症预防和拦截，也可能促进基于重排的前列腺癌风险分层生物标志物的发展。我们将通过三个具体目标来验证我们的假设：1)确定TOP2B在雄激素受体（AR）介导的转录程序启动过程中在染色体构象重塑中的作用。2)确定TOP2B结合和催化活性在雄激素诱导的双链断裂和重组/重排事件中的作用。3)确定top2b相关重排信号在人类前列腺癌中的相关性和实用性。为了实现这些目标，我们已经创新和/或在我们手中建立了可行性：i)遗传和药理学抑制TOP2B的稳健方法；Ii)明确定义的前列腺癌细胞系模型系统，研究雄激素受体信号；Iii)利用低通对端测序技术鉴定人类前列腺癌重排断点；iv)测量top2b介导的染色体环相互作用、基因表达变化、DSB和重组事件的方法；iv)强大的统计方法，用于评估这些TOP2B相关事件彼此之间的空间相关性以及在人类癌症中确定的重排断点。利用这种与这里收集的数据和公开可用数据的相关性，我们将能够评估TOP2B/AR重排特征是否与人类前列腺癌重排相关，以及它是否可以用于前列腺癌风险分层。该方法具有创新性和高度重要性，因为它使用了强大的新实验和计算方法来检验一个创新的假设，解释了在人类前列腺癌中观察到的高度复杂的、特定环境的重排的形成。这种理解最终可以为前列腺癌的预防和拦截提供信息。

DESCRIPTION (provided by applicant): Although recent studies have shown the importance of complex genomic rearrangements in prostate cancer initiation and progression, the mechanisms that generate these rearrangements are unknown. Our long-term goal is to understand the causes and consequences of these complex rearrangements in prostate and other cancers. The objective here is to determine the mechanism by which multiple, coordinate, double strand breaks (DSB) likely involved in generating these complex rearrangements can form. Our central hypothesis is that initiation of androgen-induced transcriptional programs in neoplastic prostate cells involves the formation of highly complex topological loops that are mediated in part by topoisomerase II beta (TOP2B); dysfunction of this enzyme can produce multiple simultaneous breaks that can recombine to form complex genomic rear- rangements. We have formulated this hypothesis on the basis of our strong preliminary data showing andro- gen-induced, TOP2B-mediated DSB that are recombinogenic and arise near rearrangement breakpoints ob- served in human prostate cancers. The rationale for this work is that understanding these mechanisms can inform strategies to prevent formation of these rearrangements, potentially allowing cancer prevention and interception, and may also spur development of rearrangement-based biomarkers for prostate cancer risk stratification. We will test our hypothesis through three specific aims: 1) Determine the role of TOP2B in chromosomal conformational remodeling during initiation of androgen receptor (AR) mediated transcriptional programs. 2) Identify the role of TOP2B binding and catalytic activity in generating androgen-induced double strand breaks and recombination/rearrangement events. 3) Determine the relevance and utility of a TOP2B-related rearrangement signature in human prostate cancer. To facilitate these aims, we have innovated and/or established feasibility in our hands for: i) robust methods for genetic and pharmacologic inhibition of TOP2B; ii) well defined prostate cancer cell line model systems to study androgen receptor signaling; iii) identification o rearrangement breakpoints in human prostate cancers with low-pass paired-end sequencing; iv) methods for measuring TOP2B-mediated chromosomal looping interactions, gene expression changes, DSB, and recombination events; and iv) powerful statistical methods for assessing the spatial correlations of these TOP2B- related events with each other and with rearrangement breakpoints identified in human cancers. Using such correlations with data collected here and with publically available data, we will be able to assess whether a TOP2B/AR rearrangement signature has relevance to human prostate cancer rearrangements and whether it can have utility for prostate cancer risk stratification. The approach is innovative and highly significant because it uses powerful new experimental and computational methodologies to test an innovative hypothesis explaining the formation of highly complex, context-specific rearrangements observed in human prostate cancer. This understanding can ultimately inform approaches for prostate cancer prevention and interception.