title: ROCK1 is a potential combinatorial drug target for BRAF mutant melanoma.
abstract: Treatment of BRAF mutant melanomas with specific BRAF inhibitors leads to tumor remission. However, most patients eventually relapse due to drug resistance. Therefore, we designed an integrated strategy using (phospho)proteomic and functional genomic platforms to identify drug targets whose inhibition sensitizes melanoma cells to BRAF inhibition. We found many proteins to be induced upon PLX4720 (BRAF inhibitor) treatment that are known to be involved in BRAF inhibitor resistance, including FOXD3 and ErbB3. Several proteins were down-regulated, including Rnd3, a negative regulator of ROCK1 kinase. For our genomic approach, we performed two parallel shRNA screens using a kinome library to identify genes whose inhibition sensitizes to BRAF or ERK inhibitor treatment. By integrating our functional genomic and (phospho)proteomic data, we identified ROCK1 as a potential drug target for BRAF mutant melanoma. ROCK1 silencing increased melanoma cell elimination when combined with BRAF or ERK inhibitor treatment. Translating this to a preclinical setting, a ROCK inhibitor showed augmented melanoma cell death upon BRAF or ERK inhibition in vitro. These data merit exploration of ROCK1 as a target in combination with current BRAF mutant melanoma therapies.
authors.affiliation: Division of Molecular Oncology, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
authors.forename: Marjon A
authors.initials: MA
authors.lastname: Smit
authors.affiliation: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Science, Utrecht University, Utrecht, The Netherlands Netherlands Proteomics Centre, Utrecht, The Netherlands Center for Biomembrane Research, Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden.
authors.forename: Gianluca
authors.initials: G
authors.lastname: Maddalo
authors.affiliation: Division of Molecular Oncology, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
authors.forename: Kylie
authors.initials: K
authors.lastname: Greig
authors.affiliation: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Science, Utrecht University, Utrecht, The Netherlands Netherlands Proteomics Centre, Utrecht, The Netherlands.
authors.forename: Linsey M
authors.initials: LM
authors.lastname: Raaijmakers
authors.affiliation: Division of Molecular Oncology, The Netherlands Cancer Institute, Amsterdam, The Netherlands.
authors.forename: Patricia A
authors.initials: PA
authors.lastname: Possik
authors.affiliation: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Science, Utrecht University, Utrecht, The Netherlands Netherlands Proteomics Centre, Utrecht, The Netherlands.
authors.forename: Bas
authors.initials: B
authors.lastname: van Breukelen
authors.affiliation: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Science, Utrecht University, Utrecht, The Netherlands Netherlands Proteomics Centre, Utrecht, The Netherlands.
authors.forename: Salvatore
authors.initials: S
authors.lastname: Cappadona
authors.affiliation: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Science, Utrecht University, Utrecht, The Netherlands Netherlands Proteomics Centre, Utrecht, The Netherlands.
authors.forename: Albert J R
authors.initials: AJ
authors.lastname: Heck
authors.affiliation: Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Science, Utrecht University, Utrecht, The Netherlands Netherlands Proteomics Centre, Utrecht, The Netherlands m.altelaar@uu.nl d.peeper@nki.nl.
authors.forename: A F Maarten
authors.initials: AF
authors.lastname: Altelaar
authors.affiliation: Division of Molecular Oncology, The Netherlands Cancer Institute, Amsterdam, The Netherlands m.altelaar@uu.nl d.peeper@nki.nl.
authors.forename: Daniel S
authors.initials: DS
authors.lastname: Peeper
mesh_terms: D045744:Cell Line, Tumor; D002853:Chromatography, Liquid; D015536:Down-Regulation; D051858:Forkhead Transcription Factors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007191:Indazoles / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D008545:Melanoma / Q000235:genetics*; D058990:Molecular Targeted Therapy; D009154:Mutation; D010879:Piperazines / Q000494:pharmacology; D040901:Proteomics; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D020893:Receptor, ErbB-3 / Q000235:genetics / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology; D053719:Tandem Mass Spectrometry; D000077484:Vemurafenib; D054460:rho-Associated Kinases / Q000235:genetics / Q000378:metabolism*
chemical_list: C503514:FOXD3 protein, human; D051858:Forkhead Transcription Factors; D007191:Indazoles; D007211:Indoles; C528407:PLX 4720; D010879:Piperazines; D034741:RNA, Small Interfering; C587178:SCH772984; D013449:Sulfonamides; D000077484:Vemurafenib; C581292:ERBB3 protein, human; D020893:Receptor, ErbB-3; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; C515851:ROCK1 protein, human; D054460:rho-Associated Kinases
keywords: PLX4720; ROCK1; kinome shRNA genomic screening; proteomics