CRISPR/Cas9 screening identifies a kinetochore-microtubule dependent mechanism for Aurora-A inhibitor resistance in breast cancer.
CRISPR/Cas9 screening identifies a kinetochore-microtubule dependent mechanism for Aurora-A inhibitor resistance in breast cancer.
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CRISPR/Cas9 筛选确定了乳腺癌中 Aurora-A 抑制剂耐药性的动粒微管依赖性机制。
DOI：
10.1002/cac2.12125
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发表时间：
2021-03
期刊：
影响因子：
--
通讯作者：
Liu Q
中科院分区：
文献类型：
--
作者：
Chen A;Wen S;Liu F;Zhang Z;Liu M;Wu Y;He B;Yan M;Kang T;Lam EW;Wang Z;Liu Q
Aurora‐A（AURKA）的过度表达是乳腺癌的一个特征，与不良预后相关。选择性Aurora‐A抑制剂alisertib（MLN 8237）最近在各种癌症类型中作为单药显示出有希望的抗肿瘤反应，但其III期临床试验报告为失败，因为MLN 8237在延长患者生存期方面未显示出明显作用。因此，确定可增强MLN 8237活性的潜在靶点将为联合用药实现更好的治疗结局提供依据。在此，我们在乳腺癌细胞中使用MLN 8237对507种激酶进行了系统的合成致死性CRISPR/Cas9筛选，并鉴定了许多与MLN 8237显示合成致死性相互作用的靶向激酶。然后，我们进行了竞争性生长试验、集落形成试验、细胞活力试验、细胞凋亡试验和异种移植小鼠模型，以评价Haspin（GSG 2）耗竭或抑制与MLN 8237的协同治疗作用。对于机制研究，使用免疫荧光检测微管的状态以及Aurora B和有丝分裂着丝粒相关驱动蛋白（MCAK）的定位。在命中中，我们观察到Haspin耗竭或抑制略微抑制乳腺癌细胞生长，但可显著增强MLN 8237的杀伤作用。机制研究表明，Aurora‐A和Haspin抑制剂的共同处理消除了Aurora‐B和有丝分裂着丝粒相关驱动蛋白（MCAK）向着丝粒的募集，这与过度的微管解聚、动粒微管（KT‐MT）附着失败和严重的有丝分裂灾难有关。我们进一步表明，MLN 8237和Haspin抑制剂GAL-6494联合使用可协同降低乳腺癌细胞活力，并显著抑制体外和体内肿瘤生长。这些发现确立了Haspin是一种合成的致死靶点，并证明了MLN 8237是一种潜在的组合药物，可促进MLN 8237对乳腺癌的治疗作用。这些发现为促进MLN 8237的治疗作用提供了潜在的组合药物靶点，并揭示了着丝粒相关功能与Aurora‐A抑制剂敏感性之间的关系。
Overexpression of Aurora‐A (AURKA) is a feature of breast cancer and associates with adverse prognosis. The selective Aurora‐A inhibitor alisertib (MLN8237) has recently demonstrated promising antitumor responses as a single agent in various cancer types but its phase III clinical trial was reported as a failure since MLN8237 did not show an apparent effect in prolonging the survival of patients. Thus, identification of potential targets that could enhance the activity of MLN8237 would provide a rationale for drug combination to achieve better therapeutic outcome. Here, we conducted a systematic synthetic lethality CRISPR/Cas9 screening of 507 kinases using MLN8237 in breast cancer cells and identified a number of targetable kinases that displayed synthetic lethality interactions with MLN8237. Then, we performed competitive growth assays, colony formation assays, cell viability assays, apoptosis assays, and xenograft murine model to evaluate the synergistic therapeutic effects of Haspin (GSG2) depletion or inhibition with MLN8237. For mechanistic studies, immunofluorescence was used to detect the state of microtubules and the localization of Aurora‐B and mitotic centromere‐associated kinesin (MCAK). Among the hits, we observed that Haspin depletion or inhibition marginally inhibited breast cancer cell growth but could substantially enhance the killing effects of MLN8237. Mechanistic studies showed that co‐treatment with Aurora‐A and Haspin inhibitors abolished the recruitment of Aurora‐B and mitotic centromere‐associated kinesin (MCAK) to centromeres which were associated with excessive microtubule depolymerization, kinetochore‐microtubule (KT‐MT) attachment failure, and severe mitotic catastrophe. We further showed that the combination of MLN8237 and the Haspin inhibitor CHR‐6494 synergistically reduced breast cancer cell viability and significantly inhibited both in vitro and in vivo tumor growth. These findings establish Haspin as a synthetic lethal target and demonstrate CHR‐6494 as a potential combinational drug for promoting the therapeutic effects of MLN8237 on breast cancer. These findings provide a potential combinational drug target for promoting the therapeutic effects of MLN8237 and reveal a relationship between centromere‐related functions and the sensitivity of Aurora‐A inhibitor.
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DOI：
10.1186/s40880-018-0308-x
发表时间：
2018-07-03
期刊：
Cancer communications (London, England)
影响因子：
--
作者：
Gu X;Zheng R;Xia C;Zeng H;Zhang S;Zou X;Yang Z;Li H;Chen W
通讯作者：
Chen W
DOI：
10.1007/978-1-61779-080-5_34
发表时间：
2011-01-01
期刊：
CANCER CELL CULTURE: METHODS AND PROTOCOLS, SECOND EDITION
影响因子：
--
作者：
Bijnsdorp, Irene V.;Giovannetti, Elisa;Peters, Godefridus J.
通讯作者：
Peters, Godefridus J.
影响因子：
5.7
作者：
Fletcher, Graham C.;Brokx, Richard D.;Bray, Mark R.
通讯作者：
Bray, Mark R.
影响因子：
7.8
作者：
Goshima, Gohta;Mayer, Mirjam;Zhang, Nan;Stuurman, Nico;Vale, Ronald D.
通讯作者：
Vale, Ronald D.
影响因子：
16.6
作者：
Eot-Houllier G;Magnaghi-Jaulin L;Fulcrand G;Moyroud FX;Monier S;Jaulin C
通讯作者：
Jaulin C