Oncogenic pathway-induced fragile sites: a new paradigm for understanding genome instability in cancer

摘要 癌症研究中的重大挑战之一是对治疗的反应存在巨大的异质性 不同的癌症类型。我们在治疗一些癌症方面取得了长足的进步，而另一些癌症的预后 仍然令人沮丧。大规模全基因组序列(WGS)分析已经识别出“断点签名” 不同的癌症类型，可能反映了它们潜在的干扰途径的异质性，但是，药物- 只有在断裂产生致癌融合蛋白的情况下，才会出现Gable靶标。有能力 使用WGS预测通路受阻将对癌症诊断和治疗产生很大影响。色度- 在培养的细胞中，一些脆性部位(FSS)在中期染色体上表现为缝隙和断裂。 ENS复制应激。然而，许多FS不是癌症类型特异性的，并且在癌症中有许多断点热点 都是未知的金融稳定系统，这严重限制了金融稳定领域的影响。因为培养的细胞提供了 前瞻性地剖析启动染色体断裂的机制，并追踪它们向复合体的扩展 随着结构的变化，迫切需要使这些体外系统与癌症更相关。我们的长期计划 目标是建立一种持续的研究策略，该策略可以预测在给定的时间内哪些已知路径被干扰 从他们的断点模式判断癌症类型。这项提案的总体目标是通过以下方式揭示机制 在细胞培养中，哪种与癌症相关的细胞通路的扰动会产生FSS的通路特异性模式 并确定这些模式是否可以在癌症中识别。我们的中心假设是 在体外系统中分析导致癌症断点特征的机制只有在以下情况下才会变得明显 从化学诱导的FSS转向通过干扰已知的癌症相关途径而诱导的FSS。 我们已经证明，不同癌基因的过度表达会导致FSS和Pre-Pre癌基因的特异性谱。 有限的数据表明了这种情况发生的一些机制。我们的理论基础是了解癌症- 指定FSS的相关机制将填补在体外将FSS与癌症断裂点联系起来的空白。这将是 从癌症WGS数据预测中断的通路的策略迈出的重要一步，从而建议治疗- 治疗以前难以治愈的癌症的药物。AIM1将使用高吞吐量/分辨率复制和寡色 识别下游机制的分析，以区分许多癌基因诱导的复制位点中的哪些 延迟显示为FSS。AIM2将在特定情况下阐明导致复制延迟和FS的上游机制 网站。AIM3将以前所未有的分辨率绘制FSS图，并挖掘肿瘤测序数据库中的签名 与癌基因特异性FSS的基因序列相匹配。这一贡献将是重大的，因为识别受影响的能力 仅来自断点信号的通路将暴露出精确癌症的肿瘤特异性脆弱性 医药。拟议的研究具有创新性，因为它将恢复FS研究的影响，结束 体外系统和体内癌症之间的差距，同时利用新技术操纵细胞，MAP 它们的基因组不稳定特征，并将它们与癌症断裂点特征进行匹配。

ABSTRACT One of the grand challenges in cancer research is the vast heterogeneity in responsiveness to treatment for different cancer types. We have made great strides in treating some cancers, while the prognosis for others remains dismal. Large scale whole genome sequence (WGS) analyses have identified “breakpoint signatures” of different cancer types that presumably reflect heterogeneity in their underlying disrupted pathways but, drug- gable targets have emerged only in cases where the breaks create oncogenic fusion proteins. The ability to predict disrupted pathways using WGS would be highly impactful for cancer diagnosis and treatment. Chromo- some fragile sites (FSs) manifest as gaps and breaks in metaphase chromosomes when cultured cells experi- ence replication stress. However, many FSs are not cancer-type specific and many breakpoint hotspots in cancer are not known FSs, which has severely limited the impact of the FS field. Since cultured cells offer the potential to prospectively dissect mechanisms initiating chromosome breaks and to track their expansion into complex structural variation, there is a critical need to make these in vitro systems more cancer relevant. Our longterm goal is to establish a sustained research strategy that can predict which known pathways are disrupted in a given cancer type from their breakpoint patterns. The overall objective of this proposal is to reveal mechanisms by which perturbation of cancer-relevant cellular pathways produce pathway-specific patterns of FSs in cell culture and determine whether those patterns can be identified in cancers. Our central hypothesis is that the power of in vitro systems to dissect mechanisms leading to cancer breakpoint signatures will become evident only when shifted away from chemically-induced FSs towards FSs induced by perturbing known cancer-relevant pathways. We have shown that overexpression of different oncogenes leads to oncogene-specific spectra of FSs and pre- liminary data suggest some of the mechanisms by which this occurs. Our rationale is that understanding cancer- relevant mechanisms that specify FSs will fill the gap in linking in vitro FSs to cancer breakpoints. This would be a major step toward a strategy to predict disrupted pathways from cancer WGS data, thereby suggesting treat- ments for previously intractable cancers. Aim1 will use high throughput/resolution replication and Oligopaints assays to identify downstream mechanisms distinguishing which of many sites of oncogene-induced replication delay manifest as FSs. Aim2 will elucidate upstream mechanisms causing replication delays and FS at specific sites. Aim3 will map FSs at unprededented resolution and mine tumor sequencing databases for signatures that match those of oncogene-specific FSs. This contribution will be significant because the ability to identify affected pathways solely from breakpoint signatures would expose tumor-specific vulnerabilities for precision cancer medicine. The proposed research is innovative because it will rejuvenate the impact of FS research, closing the gap between in vitro systems and in vivo cancers while leveraging novel technologies to manipulate cells, map their signatures of genome instability and match them to cancer breakpoint signatures.