Cargando…
Target identification of small molecules using large-scale CRISPR-Cas mutagenesis scanning of essential genes
Unraveling the mechanism of action and molecular target of small molecules remains a major challenge in drug discovery. While many cancer drugs target genetic vulnerabilities, loss-of-function screens fail to identify essential genes in drug mechanism of action. Here, we report CRISPRres, a CRISPR-C...
Autores principales: | , , , , , , , , , , , , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Nature Publishing Group UK
2018
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5799254/ https://www.ncbi.nlm.nih.gov/pubmed/29402884 http://dx.doi.org/10.1038/s41467-017-02349-8 |
_version_ | 1783297958075170816 |
---|---|
author | Neggers, Jasper Edgar Kwanten, Bert Dierckx, Tim Noguchi, Hiroki Voet, Arnout Bral, Lotte Minner, Kristien Massant, Bob Kint, Nicolas Delforge, Michel Vercruysse, Thomas Baloglu, Erkan Senapedis, William Jacquemyn, Maarten Daelemans, Dirk |
author_facet | Neggers, Jasper Edgar Kwanten, Bert Dierckx, Tim Noguchi, Hiroki Voet, Arnout Bral, Lotte Minner, Kristien Massant, Bob Kint, Nicolas Delforge, Michel Vercruysse, Thomas Baloglu, Erkan Senapedis, William Jacquemyn, Maarten Daelemans, Dirk |
author_sort | Neggers, Jasper Edgar |
collection | PubMed |
description | Unraveling the mechanism of action and molecular target of small molecules remains a major challenge in drug discovery. While many cancer drugs target genetic vulnerabilities, loss-of-function screens fail to identify essential genes in drug mechanism of action. Here, we report CRISPRres, a CRISPR-Cas-based genetic screening approach to rapidly derive and identify drug resistance mutations in essential genes. It exploits the local genetic variation created by CRISPR-Cas-induced non-homologous end-joining (NHEJ) repair to generate a wide variety of functional in-frame mutations. Using large sgRNA tiling libraries and known drug–target pairs, we validate it as a target identification approach. We apply CRISPRres to the anticancer agent KPT-9274 and identify nicotinamide phosphoribosyltransferase (NAMPT) as its main target. These results present a powerful and simple genetic approach to create many protein variants that, in combination with positive selection, can be applied to reveal the cellular target of small-molecule inhibitors. |
format | Online Article Text |
id | pubmed-5799254 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | Nature Publishing Group UK |
record_format | MEDLINE/PubMed |
spelling | pubmed-57992542018-02-08 Target identification of small molecules using large-scale CRISPR-Cas mutagenesis scanning of essential genes Neggers, Jasper Edgar Kwanten, Bert Dierckx, Tim Noguchi, Hiroki Voet, Arnout Bral, Lotte Minner, Kristien Massant, Bob Kint, Nicolas Delforge, Michel Vercruysse, Thomas Baloglu, Erkan Senapedis, William Jacquemyn, Maarten Daelemans, Dirk Nat Commun Article Unraveling the mechanism of action and molecular target of small molecules remains a major challenge in drug discovery. While many cancer drugs target genetic vulnerabilities, loss-of-function screens fail to identify essential genes in drug mechanism of action. Here, we report CRISPRres, a CRISPR-Cas-based genetic screening approach to rapidly derive and identify drug resistance mutations in essential genes. It exploits the local genetic variation created by CRISPR-Cas-induced non-homologous end-joining (NHEJ) repair to generate a wide variety of functional in-frame mutations. Using large sgRNA tiling libraries and known drug–target pairs, we validate it as a target identification approach. We apply CRISPRres to the anticancer agent KPT-9274 and identify nicotinamide phosphoribosyltransferase (NAMPT) as its main target. These results present a powerful and simple genetic approach to create many protein variants that, in combination with positive selection, can be applied to reveal the cellular target of small-molecule inhibitors. Nature Publishing Group UK 2018-02-05 /pmc/articles/PMC5799254/ /pubmed/29402884 http://dx.doi.org/10.1038/s41467-017-02349-8 Text en © The Author(s) 2018 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. |
spellingShingle | Article Neggers, Jasper Edgar Kwanten, Bert Dierckx, Tim Noguchi, Hiroki Voet, Arnout Bral, Lotte Minner, Kristien Massant, Bob Kint, Nicolas Delforge, Michel Vercruysse, Thomas Baloglu, Erkan Senapedis, William Jacquemyn, Maarten Daelemans, Dirk Target identification of small molecules using large-scale CRISPR-Cas mutagenesis scanning of essential genes |
title | Target identification of small molecules using large-scale CRISPR-Cas mutagenesis scanning of essential genes |
title_full | Target identification of small molecules using large-scale CRISPR-Cas mutagenesis scanning of essential genes |
title_fullStr | Target identification of small molecules using large-scale CRISPR-Cas mutagenesis scanning of essential genes |
title_full_unstemmed | Target identification of small molecules using large-scale CRISPR-Cas mutagenesis scanning of essential genes |
title_short | Target identification of small molecules using large-scale CRISPR-Cas mutagenesis scanning of essential genes |
title_sort | target identification of small molecules using large-scale crispr-cas mutagenesis scanning of essential genes |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5799254/ https://www.ncbi.nlm.nih.gov/pubmed/29402884 http://dx.doi.org/10.1038/s41467-017-02349-8 |
work_keys_str_mv | AT neggersjasperedgar targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT kwantenbert targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT dierckxtim targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT noguchihiroki targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT voetarnout targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT brallotte targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT minnerkristien targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT massantbob targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT kintnicolas targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT delforgemichel targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT vercruyssethomas targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT balogluerkan targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT senapediswilliam targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT jacquemynmaarten targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes AT daelemansdirk targetidentificationofsmallmoleculesusinglargescalecrisprcasmutagenesisscanningofessentialgenes |