Systematic screening of isogenic cancer cells identifies DUSP6 as context-specific synthetic lethal target in melanoma
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Stephanie Wittig-Blaich1,2,*, Rainer Wittig1,3,*, Steffen Schmidt4,5,*, Stefan Lyer1,6, Melanie Bewerunge-Hudler1,7, Sabine Gronert-Sum1, Olga Strobel-Freidekind1, Carolin Müller4,5, Markus List4,5, Aleksandra Jaskot4,5, Helle Christiansen4,5, Mathias Hafner8, Dirk Schadendorf9, Ines Block10,11,**, Jan Mollenhauer11,12,**
1Department of Molecular Genome Analysis, German Cancer Research Center (DKFZ), 69118 Heidelberg, Germany
2Institute for Comparative Molecular Endocrinology, Ulm University, 89081 Ulm, Germany
3Institute for Laser Technologies in Medicine and Metrology, Ulm University, 89081 Ulm, Germany
4Lundbeckfonden Center of Excellence NanoCAN, Institute of Molecular Medicine, University of Southern Denmark, 5000 Odense, Denmark
5Molecular Oncology, Institute of Molecular Medicine, University of Southern Denmark, 5000 Odense, Denmark
6Department of Otorhinolaryngology, Section for Experimental Oncology and Nanomedicine (SEON), University Hospital Erlangen, 91054 Erlangen, Germany
7Genomics and Proteomics Core Facility, German Cancer Research Center (DKFZ), 69118 Heidelberg, Germany
8Department of Biotechnology, Mannheim University of Applied Sciences, 68163 Mannheim, Germany
9Department of Dermatology, University Hospital Duisburg-Essen, 45147 Essen, Germany and German Cancer Consortium, 69118 Heidelberg, Germany
10Department of Clinical Genetics, Odense University Hospital, 5000 Odense, Denmark
11Lundbeckfonden Center of Excellence NanoCAN, Institute of Molecular Medicine, University of Southern Denmark, 5000 Odense, Denmark
12Molecular Oncology, Institute of Molecular Medicine, University of Southern Denmark, 5000 Odense, Denmark
Ines Block, email: firstname.lastname@example.org
Keywords: cancer, isogenic cell line libraries, functional genomics, melanoma, synthetic lethal
Received: September 14, 2016 Accepted: February 06, 2017 Published: March 02, 2017
Next-generation sequencing has dramatically increased genome-wide profiling options and conceptually initiates the possibility for personalized cancer therapy. State-of-the-art sequencing studies yield large candidate gene sets comprising dozens or hundreds of mutated genes. However, few technologies are available for the systematic downstream evaluation of these results to identify novel starting points of future cancer therapies.
We improved and extended a site-specific recombination-based system for systematic analysis of the individual functions of a large number of candidate genes. This was facilitated by a novel system for the construction of isogenic constitutive and inducible gain- and loss-of-function cell lines. Additionally, we demonstrate the construction of isogenic cell lines with combinations of the traits for advanced functional in vitro analyses. In a proof-of-concept experiment, a library of 108 isogenic melanoma cell lines was constructed and 8 genes were identified that significantly reduced viability in a discovery screen and in an independent validation screen. Here, we demonstrate the broad applicability of this recombination-based method and we proved its potential to identify new drug targets via the identification of the tumor suppressor DUSP6 as potential synthetic lethal target in melanoma cell lines with BRAF V600E mutations and high DUSP6 expression.
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