Name: a data integration workflow to identify drug combinations targeting synthetic lethal interactions
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开发数据一体化工作流程的资金是根据赠款协定从欧洲共同体第七框架方案获得的。279113（奥克西普斯）。本出版物中的数据改编已获得公共科学图书馆出版物和影响期刊有限责任公司的批准。

1. 检索合成致命基因对
<ol><li>从生物电网检索数据。<ol>
<li>使用 web 浏览器或直接从 Linux 命令行使用卷发或 wget18从 https://downloads.thebiogrid.org/Download/BioGRID/Latest-Release/BIOGRID-ALL-LATEST.tab2.zip 下载最新的 BioGRID 交互文件，以 tab2 格式下载。 ##download并拆开最新的生物GRID交互文件 使用卷发的#download最新的生物格栅交互文件 卷曲-o biogrid_latest.zip https://downloads.thebiogrid.org/...

<ol>
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我们概述了一个工作流程，以确定影响合成致命相互作用的药物组合。该工作流程利用（一） 模型生物合成致命相互作用的数据、（ii） 人类矫形器信息、（三） 药物靶向关联信息、（四） 癌症临床试验药物信息，以及从科学文献中提取的药物疾病和基因疾病关联信息（五） 。综合信息可用于评估正在调查的特定药物组合对合成致命基因对的影响。此外，综合数据可用于评估目前在癌症临床试...

合成杀伤力定义了两个基因的关联，其中一个基因的丧失不影响生存能力，但两个基因的丧失会导致细胞死亡。它第一次描述于1946年由多布赞斯基，同时分析各种表型的嗜血杆菌通过繁殖同源突变体1。没有产生可行后代的突变体虽然本身也可行，但与某些其他突变体交叉时表现出致命的表型，为建立合成杀伤力理论奠定了基础。哈特威尔和他的同事认为，这个概念可能适用于人类癌症治疗2。药理学引起的合成杀伤力可能只依赖于一个突变，因为突变基因的合成致命伙伴是药理化合物的目标。第一个能够使合成杀伤力的药理诱导基因对是BRCA（1/2）和PARP1。PARP1 作为 DNA 损伤的传感器，与双和单 DNA 链断裂、超线圈和交叉3的位点相连。BRCA1和2在修复DNA双链断裂通过同源重组4方面发挥着重要作用。农民和他的同事公布了发现，缺乏BRCA1/2的细胞易受PARP抑制，而在BRCA野生型细胞5中未观察到细胞毒性。最终，PARP抑制剂被批准用于治疗BRCA缺乏乳房和卵巢癌6，7。此外，合成致死基因对导致药理化合物的临床批准是备受期待的，是最近癌症研究的主要领域8。
合成致命基因相互作用在多种生物体中建模，包括果蝇、C.elegans和酵母2。近年来，利用RNA干扰和CRISPR/CAS-库淘汰等多种方法，发现了新的合成致命基因对9、10、11。胡斯登及其同事最近发表了一份关于RNAi与CRISPR/CAS结合的实验程序的议定书。同时，研究人员还在人细胞中进行了大屏幕，以确定合成的致命相互作用13、14。在生物网络分析和机器学习等硅学方法中，在发现合成致命相互作用方面也显示出了希望。
从概念上讲，在抗肿瘤治疗中利用合成致命相互作用的一种方法是识别肿瘤细胞中的变异或非功能性蛋白质，使其合成致命相互作用伙伴有望成为治疗干预的药物目标。由于大多数肿瘤类型的异质性，研究人员已经开始寻找所谓的合成致命中枢蛋白。这些合成致命中心有许多合成致命相互作用伙伴，这些伙伴要么发生变异，要么因此在肿瘤样本中不起作用或显著降低管制。解决这种合成致命中心在提高药物疗效或克服耐药性方面是有希望的，例如在耐葡萄抗神经母细胞瘤17的背景下可以证明这一点。利用合成致命相互作用的概念加强药物治疗的第二种方法是确定针对合成致命相互作用的药物组合。这可能导致已经批准的单一抗肿瘤疗法的新组合，并使药物从其他疾病领域重新定位到肿瘤学领域。
在本文中，我们提出了一个分步过程，以生成针对合成致命相互作用对的药物组合列表。在此工作流程中，我们（一） 使用来自 BioGRID 的合成致命相互作用的数据和 （ii） 来自 Ensembl 的同源基因信息，（iii） 从药物库检索药物靶向对，（iv） 从 ClinicalTrials.gov 建立疾病-药物关联，（v） 从而生成一组解决合成致命相互作用的药物组合。最后，我们在代表性结果部分提供卵巢癌和乳腺癌背景下的药物组合。

<table>
	<tbody>
		<tr>
			<td>BioGRID</td>
			<td>n/a</td>
			<td>n/a</td>
			<td>thebiogrid.org</td>
		</tr>
		<tr>
			<td>ClinicalTrials.gov</td>
			<td>n/a</td>
			<td>n/a</td>
			<td>ClinicalTrials.gov</td>
		</tr>
		<tr>
			<td>DrugBank</td>
			<td>n/a</td>
			<td>n/a</td>
			<td>drugbank.ca</td>
		</tr>
		<tr>
			<td>Ensembl BioMart</td>
			<td>n/a</td>
			<td>n/a</td>
			<td>ensembl.org</td>
		</tr>
		<tr>
			<td colspan="4">for alternative computational databases please refer to the manuscript</td>
		</tr>
		<tr>
			<td>7-AAD</td>
			<td>ebioscience</td>
			<td>00-6993-50</td>
			<td></td>
		</tr>
		<tr>
			<td>AnnexinV-APC</td>
			<td>BD Bioscience</td>
			<td>550474</td>
			<td></td>
		</tr>
		<tr>
			<td>celecoxib</td>
			<td>Sigma-Aldrich</td>
			<td>PZ0008-25MG</td>
			<td></td>
		</tr>
		<tr>
			<td>CellTiter-Blue Viability Assay</td>
			<td>Promega</td>
			<td>G8080</td>
			<td></td>
		</tr>
		<tr>
			<td>FACS Canto II</td>
			<td>BD Bioscience</td>
			<td>n/a</td>
			<td></td>
		</tr>
		<tr>
			<td>fetal bovine serum</td>
			<td>Fisher Scientific/Gibco</td>
			<td>16000044</td>
			<td></td>
		</tr>
		<tr>
			<td>FloJo Software</td>
			<td>FloJo LLC</td>
			<td>V10</td>
			<td></td>
		</tr>
		<tr>
			<td>McCoy&#39;s 5a Medium Modified</td>
			<td>Fisher Scientific/Gibco</td>
			<td>16600082</td>
			<td></td>
		</tr>
		<tr>
			<td>penicillin G/streptomycin sulfate</td>
			<td>Fisher Scientific/Gibco</td>
			<td>15140122</td>
			<td></td>
		</tr>
		<tr>
			<td>SKBR-3 cells</td>
			<td>American Type Culture Collection (ATCC)</td>
			<td>ATCC HTB-30</td>
			<td></td>
		</tr>
		<tr>
			<td>zoledronic acid</td>
			<td>Sigma-Aldrich</td>
			<td>SML0223-50MG</td>
			<td></td>
		</tr>
		<tr>
			<td colspan="4">further materials or equipment will be made available upon request</td>
		</tr>
	</tbody>
</table>

a data integration workflow to identify drug combinations targeting synthetic lethal interactions

当两个基因中的任何一个被击倒时，两个基因之间的合成致命相互作用不会影响细胞的生存能力，但两个合成致命相互作用器的淘汰会导致细胞生存能力丧失或细胞死亡。研究最好的合成致命相互作用是BRCA1+2和PARP1之间，PARP1抑制剂用于临床实践治疗BRCA1/2突变肿瘤患者。模型生物体中以及人细胞系中的大型基因筛选导致识别出许多额外的合成致命相互作用对，所有这些都成为开发新型肿瘤疗法的潜在目标。一种方法是用一种合成的致命相互作用器对基因进行治疗，这种相互作用器在感兴趣的肿瘤中发生变异或显著降低调节。第二种方法是制定药物组合，解决合成致命相互作用问题。在本文中，我们概述了数据集成工作流程，以评估和识别针对合成致命相互作用的药物组合。我们利用现有的合成致命相互作用对数据集、同源映射资源、专用数据库中的药物目标链接，以及有关相关疾病领域临床试验中正在调查的药物信息。我们进一步强调了我们小组最近关于卵巢癌和乳腺癌药物组合评估的两项研究的主要发现。

我们的小组最近发表了两项研究，应用本手稿中描述的工作流程，以确定针对卵巢癌和乳腺癌24，25背景下合成致命相互作用的药物组合。在第一项研究中，我们评估了目前在晚期临床试验（第三阶段和第四阶段）中测试的药物组合，或在临床实践中已经用于治疗卵巢癌患者对合成致命相互作用的影响。此外，我们确定了目前尚未在临床试验中测试的药?...

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A Data Integration Workflow to Identify Drug Combinations Targeting Synthetic Lethal Interactions

模型生物体中的大型基因筛选导致对负基因相互作用的识别。在这里，我们使用模型生物体中遗传筛选的数据来描述数据集成工作流，以描述针对癌症中合成致命相互作用的药物组合。

数据集成工作流，以识别针对合成致命相互作用的药物组合

A synthetic lethal interaction between two genes is given when knock-out of either one of the two genes does not affect cell viability but knock-out of ...

当两个基因的单次击倒不影响细胞生存能力，但两个合成致命伙伴的击倒会导致细胞生存能力丧失或细胞死亡时，就会发生两个基因之间的合成致命相互作用。结合异构数据集可识别合成致命相互作用，这些相互作用可成为癌症等疾病药物组合的目标，目的是阻止细胞增殖。合成杀伤力的概念对于任何细胞增殖是一个问题的疾病都感兴趣。至于癌症，合成致命基因相互作用已经通过使用PARP抑制剂在BRCA突变肿瘤中成为目标。像我们这样的努力可以扩大其他基因相互作用和癌症实体的概念的使用。在我们的实验室中，我们测试了针对乳腺癌背景下合成致命相互作用的计算预测药物组合。首先从生物电网检索数据。使用 Web 浏览器以 tab2 格式下载最新的 BioGRID 交互文件。筛选列仅保留相关用于后续分析步骤的列。其次，过滤合成杀伤力和负基因相互作用。使用实验系统列中的信息将数据集限制为具有负遗传或合成杀伤力值的条目。通过确定合成致命相互作用伙伴税项 ID 的数量来确定报告合成致命相互作用的物种，这将提供每个有机体可用的这些相互作用数量的估计值。要从 Ensembl BioMart 中检索相关模型生物体的人类矫型细胞，请选择相应的模型生物数据集，单击属性，并选择基因名称。然后单击数据集，选择人类基因。再次单击属性，然后选择基因名称。然后单击"结果"。仅检查唯一的结果，然后单击 Go.自动化检索过程，并将查询直接发送到 BioMart RESTful 访问以检索正交基因对。如果通过 Ensembl BioMart Web 界面手动检索数据，请重命名并注意自动添加标头行。为了为其他模型生物检索直观人类基因，请用各自的 Ensembl 数据集名称替换第一个数据集元素的名称属性值。接下来，丢弃任何没有同源基因的输入，在单个文件中收集所有同源映射，并为人类基因添加虚拟映射。此外，为人类基因添加人工条目。通过为每个交互伙伴添加一个持有税务 ID 和基因符号组合的新列，为后续加入准备合成文件。加入基于生物体税ID和基因符号的合成致命相互作用与检索回的矫直对。从药物银行从下载部分检索药物目标对，如有必要，首先创建一个帐户。将药品银行药品目标文件限制在相关栏目。仅保留人类分子实体的条目。然后，提取相关列，并过滤人类物种的数据。由于药物名称和药物靶点信息在两个单独的 CSV 文件中提供，必须合并两个文件中的信息。为此，必须首先使药品名称条目正常化。然后使药物靶点文件正常化，使每个药物有一条线。继续使用药物银行词汇文件，并提取相关列。由于药物名称和药物靶点信息在两个单独的CSV文件中提供，因此使用通用的药物银行 ID 合并两个文件中的信息。最后，从 ClinicalTrials.gov 检索有关临床试验的信息。为了便于访问，请使用临床试验转化倡议提供的关系数据库，如有必要，请先创建一个帐户。将建议的药物溶解在适当的溶剂中，如DMSO或PBS，至少四种不同的浓度。将药物添加到96井板中播种的人类乳腺癌细胞中，并在所需的时间内孵育，以确定抑制浓度值。使用选择的生存能力或凋亡检测，以确定细胞生存能力与各种药物组合和浓度，从先前建立的IC50开始。然后，通过计算药物组合的组合指数来确定药物组合的协同细胞毒性作用。这种方法用于识别针对卵巢癌合成致命相互作用的药物组合。研究发现，21种独特的药物促成了84种已查明的药物组合，这些药物组合针对的是一组39种合成致命相互作用器。同样的工作流程被用来识别243个有前途的药物组合，针对乳腺癌背景下的166个合成致命基因对。精选组合被测试对两个乳腺癌细胞系的细胞生存能力和凋亡的影响。对西勒科西布、佐勒德龙酸以及SK-BR-3乳腺癌细胞系中佐勒德龙酸和西勒科西布的结合的可行性检测结果表明，当合并后，这些药物对细胞的生存能力有显著的协同作用。附件五和7-AAD染色SK-BR-3细胞自行治疗，并结合表明，晚凋亡细胞和坏死细胞的百分比增加后，治疗与药物组合。在尝试此协议时，需要花时间确定所有药物和从临床试验中检索到的干预措施的相应药物库药物名称。将基因表达特征或研究中该疾病合成致命相互作用者的注释程度等额外数据纳入研究中的疾病，可进一步用于确定合成致命相互作用的优先级。由于可用生物医学数据的数量不断增加，与计算生物学家联手是一种有益的相互作用，导致新的假设在实验室中测试。在与细胞静止剂合作时，请务必遵循您当地的指南，处理实验室安全设备和危险试剂的处理。在任何时候，避免直接接触，并确保在开始实验之前收集有关所用物质的信息。

Research

JoVE Journal

Cancer Research

Utilizing Functional Genomics Screening to Identify Potentially Novel Drug Targets in Cancer Cell Spheroid Cultures

利用功能基因组筛选识别在肿瘤细胞球体培养潜在的新药物靶标

The identification of functional driver events in cancer is central to furthering our understanding of cancer biology and indispensable for the discovery ...

科研

癌症研究

In Vitro Imaging and Quantification of the Drug Targeting Efficiency of Fluorescently Labeled GnRH Analogues

In Vitro Imaging and Quantification of the Drug Targeting Efficiency of Fluorescently Labeled GnRH Analogues

体外成像和荧光标记的GnRH类似物的药物靶向性的定量分析

GnRH analogues are effective targeting moieties and able to deliver anticancer agents selectively into malignant tumor cells which highly express GnRH ...

Genome-wide RNAi Screening to Identify Host Factors That Modulate Oncolytic Virus Therapy

全基因组的 rna 干扰筛查以确定调节溶瘤病毒治疗的宿主因素

High-throughput genome-wide RNAi (RNA interference) screening technology has been widely used for discovering host factors that impact&#160;virus ...

A GPC3-targeting Bispecific Antibody, GPC3-S-Fab, with Potent Cytotoxicity

GPC3-targeting 双特异性抗体, GPC3-S-Fab, 具有强大的细胞毒性

This protocol describes the construction and functional studies of a bispecific antibody (bsAb), GPC3-S-Fab. bsAbs can recognize two different epitopes ...

Synthesis and Characterization of Placental Chondroitin Sulfate A (plCSA)-Targeting Lipid-Polymer Nanoparticles

Synthesis and Characterization of Placental Chondroitin Sulfate A plCSA-Targeting Lipid-Polymer Nanoparticles

胎盘硫酸软骨素 A (plCSA) 靶向脂质高分子纳米粒的合成与表征

An effective cancer therapeutic method reduces and eliminates tumors with minimal systemic toxicity. Actively targeting nanoparticles offer a promising ...

A Chromatin Immunoprecipitation Assay to Identify Novel NFAT2 Target Genes in Chronic Lymphocytic Leukemia

一种检测慢性淋巴细胞白血病新 nfat2 靶基因的染色质免疫沉淀法

Chronic lymphocytic leukemia (CLL) is characterized by the expansion of malignant B cell clones and represents the most common leukemia in western ...

A Simple Migration/Invasion Workflow Using an Automated Live-cell Imager

使用自动活单元成像仪的简单迁移/入侵工作流

Cancer cell mobility is crucial for the initiation of metastasis. Therefore, investigation of the cell movement and invasive capacity is of great ...

Integration of Bioinformatics Approaches and Experimental Validations to Understand the Role of Notch Signaling in Ovarian Cancer

生物信息学方法与实验验证的集成，了解诺奇信号在卵巢癌中的作用

Notch signaling is a highly conserved regulatory pathway involved in many cellular processes. Dysregulation of this signaling pathway often leads to ...

Establishing a Competing Risk Regression Nomogram Model for Survival Data

为生存数据建立竞争风险回归 Nomogram 模型

The Kaplan&#8211;Meier method and Cox proportional hazards regression model are the most common analyses in the survival framework. These are relatively ...

In Vivo Targeting of Xenografted Human Cancer Cells with Functionalized Fluorescent Silica Nanoparticles in Zebrafish

斑马鱼中具有功能化荧光硅纳米粒子的异种移植人类癌细胞的Vivo靶向

Developing nanoparticles capable of detecting, targeting, and destroying cancer cells is of great interest in the field of nanomedicine. In vivo animal ...