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。这里的目的是确定在产生这些复杂重排过程中可能涉及的多个配位双链断裂(DSB)的形成机制。我们的中心假设是，雄激素诱导的前列腺癌细胞转录程序的启动涉及形成高度复杂的拓扑环，这些拓扑环部分由拓扑异构酶IIβ(TOP2B)介导；该酶的功能障碍可以产生多个同时断裂，这些断裂可以重组形成复杂的基因组后排列。我们提出这一假说的基础是，我们的初步数据显示，在人类前列腺癌中，雄激素诱导的、TOP2B介导的DSB是重组的，并出现在重排断裂点附近。这项工作的基本原理是，了解这些机制可以提供预防这些重排形成的策略，潜在地允许癌症预防和拦截，还可能刺激基于重排的生物标记物的发展，用于前列腺癌风险分层。我们将通过三个特定的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期间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/或建立了以下方面的可行性：i)TOP2B基因和药物抑制的可靠方法；ii)明确的前列腺癌细胞系模型系统来研究雄激素受体信号；iii)利用低通配对末端测序确定人类前列腺癌中的重排断裂点；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；以及iv)强大的统计学方法来评估这些TOP2B相关事件彼此之间的空间相关性以及与在人类癌症中确定的重排断点之间的空间相关性。利用这些与本文收集的数据和公开可用的数据的相关性，我们将能够评估TOP2B/AR重排标记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。这种方法是创新的，意义重大，因为它使用了强大的新的实验和计算方法来测试一个创新的假说，该假说解释了在人类前列腺癌中观察到的高度复杂的、特定于背景的重排的形成。这一认识最终可以为前列腺癌的预防和拦截提供参考。

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.