MicroSCALE screening reveals genetic modifiers of therapeutic response in melanoma.
MicroSCALE screening reveals genetic modifiers of therapeutic response in melanoma.
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微观筛查显示黑色素瘤中治疗反应的遗传修饰剂。
DOI：
10.1126/scisignal.2002612
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发表时间：
2012-05-15
影响因子：
7.3
通讯作者：
Sabatini DM
中科院分区：
文献类型：
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作者：
Wood KC;Konieczkowski DJ;Johannessen CM;Boehm JS;Tamayo P;Botvinnik OB;Mesirov JP;Hahn WC;Root DE;Garraway LA;Sabatini DM
细胞微阵列是以合理的成本在哺乳动物细胞中进行大规模功能性基因组筛选的一种有前途的工具，但由于技术限制，一直被限制在狭窄的细胞系和短期分析中使用。在这里，我们介绍MicroScale(空间受限粘附性慢病毒特征微阵列)，这是一种基于细胞微阵列的平台，使这项技术能够应用于广泛的细胞类型和更长期的分析。我们使用微尺度揭示了当过度表达部分不敏感的B-RAFV600E突变黑色素瘤细胞对丝裂原激活蛋白激酶(MAPKKK)RAF、MAPKKK1和2、mTOR(雷帕霉素的哺乳动物靶标)或PI3K(磷脂酰肌醇3-激酶)的抑制剂。这些筛选表明，通过共同机制作用的抑制剂处理的细胞受到类似的过度表达蛋白质的影响。相比之下，涉及通过不同机制作用的抑制剂的筛查产生了独特的特征，这一发现对小分子靶标识别和联合药物研究具有潜在的相关性。此外，通过结合大规模的功能筛选结果与癌细胞系的基因表达和药理学敏感性数据，我们验证了核因子κB(NF-κB)途径是潜在的MAPK途径抑制剂耐药的介导物。这里描述的微尺度平台可以实现以前不可行的大规模、资源高效的新类别的筛选，包括涉及细胞系、扰动和分析输出的组合的筛选，或者涉及有限数量的细胞和有限或昂贵的试剂的筛选。
Cell microarrays are a promising tool for performing large-scale functional genomic screening in mammalian cells at reasonable cost, but due to technical limitations, have been restricted for use with a narrow range of cell lines and short-term assays. Here, we describe MicroSCALE (Microarrays of Spatially Confined Adhesive Lentiviral Features), a cell microarray-based platform that enables application of this technology to a wide range of cell types and longer term assays. We used MicroSCALE to uncover kinases that when overexpressed partially desensitized B-RAFV600E-mutant melanoma cells to inhibitors of the mitogen-activated protein kinase kinase kinase (MAPKKK) RAF, the MAPKKs MEK1 and 2, mTOR (mammalian target of rapamycin), or PI3K (phosphatidylinositol 3-kinase). These screens indicated that cells treated with inhibitors acting through common mechanisms were affected by a similar profile of overexpressed proteins. In contrast, screens involving inhibitors acting through distinct mechanisms yielded unique profiles, a finding that has potential relevance for small molecule target identification and combination drugging studies. Further, by integrating large-scale functional screening results with cancer cell line gene expression and pharmacological sensitivity data, we validated the nuclear factor κB (NF-κB) pathway as a potential mediator of resistance to MAPK pathway inhibitors. The MicroSCALE platform described here may enable new classes of large-scale, resource-efficient screens that were not previously feasible, including those involving combinations of cell lines, perturbations, and assay outputs or those involving limited numbers of cells and limited or expensive reagents.
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影响因子：
11.2
作者：
Gray-Schopfer, Vanessa C.;Karasarides, Maria;Marais, Richard
通讯作者：
Marais, Richard
影响因子：
48
作者：
Collins, Sean R.;Weissman, Jonathan S.;Krogan, Nevan J.
通讯作者：
Krogan, Nevan J.
影响因子：
82.9
作者：
通讯作者：
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影响因子：
7
作者：
Conrad, C;Erfle, H;Eils, R
通讯作者：
Eils, R
影响因子：
64.8
作者：
通讯作者：
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