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Cancer Drug Resist 2018;1:266-302 I http://dx.doi.org/10.20517/cdr.2018.18 Page 275

19. Withdrawal of MEK1/2 inhibitor reverses acquired resistance driven by BRAF amplification
but drives EMT and chemoresistance in cells with amplified KRAS

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Matthew J. Sale , Kathryn Balmanno , Jayeta Saxena , Eiko Ozono , Rebecca E. McIntyre ,
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1
Katarzyna Wojdyla , Rebecca Gilley , Anna Woroniuk , Karen D. Howarth , Gareth Hughes ,
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Jonathan R. Dry , Mark J. Arends , David Oxley , Susan Ashton , David J. Adams , Julio Saez-
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Rodriguez , Paul D. Smith , Simon J. Cook 1
1 Signalling Programme, The Babraham Institute, Babraham Research Campus, Cambridge CB22 3AT, UK
2 Experimental Cancer Genetics, Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cam-
bridgeshire CB10 1SA, UK
3 Proteomics Facility, The Babraham Institute, Babraham Research Campus, Cambridge CB22 3AT, UK
4 Hutchison-MRC Research Centre, Department of Pathology, University of Cambridge, Hills Road, Cam-
bridge CB2 0XZ, UK
5 Oncology Bioscience, Innovative Medicines and Early Development Biotech Unit, AstraZeneca, CRUK Cam-
bridge Institute, Robinson Way, Cambridge CB2 0RE, UK
6 Oncology Bioscience, Innovative Medicines and Early Development Biotech Unit, AstraZeneca, 35 Gatehouse
Drive, Waltham, Massachusetts 02451, USA
7 Division of Pathology, Centre for Comparative Pathology, Cancer Research UK Edinburgh Centre, MRC In-
stitute of Genetics & Molecular Medicine, University of Edinburgh, Western General Hospital, Crewe Road
South, Edinburgh EH4 2XR, UK
8 Oncology Bioscience, Innovative Medicines and Early Development Biotech Unit, AstraZeneca, Alderley
Park, Macclesfield SK10 4TG, UK
9 European Molecular Biology Laboratory, European Bioinformatics Institute (EMBL-EBI), Wellcome Genome
Campus, Hinxton, Cambridgeshire CB10 1SD, UK
Acquired resistance to MEK1/2 inhibitors can arise through amplification of BRAF V600E or KRAS G13D to
reinstate ERK1/2 signalling. Here we show that BRAF V600E amplification and selumetinib resistance are
fully reversible following drug withdrawal. Resistant cells with BRAF V600E amplification become addicted to
selumetinib to maintain a precise level of ERK1/2 signalling (2%-3% of total ERK1/2 active, here quantified
by mass spectrometry), that is optimal for cell survival and proliferation. The magnitude of ERK1/2 activa-
tion following selumetinib withdrawal (~20% active) drives a p57 KIP2 -dependent G1 cell cycle arrest and se-
nescence or expression of NOXA and cell death, which selects against those cells with amplified BRAF V600E .

ERK1/2-dependent p57 KIP2 expression is required for loss of BRAF V600E amplification and determines
the rate of reversal of selumetinib resistance. Furthermore, growth of selumetinib-resistant cells with
BRAF V600E amplification as tumour xenografts is inhibited in mice that do not receive selumetinib. Thus,
BRAF V600E amplification confers a selective disadvantage during drug withdrawal, providing a rationale for
intermittent dosing to forestall resistance. In striking contrast, selumetinib resistance driven by KRAS G13D
amplification is not reversible. In these cells ERK1/2 reactivation does not inhibit proliferation but drives a
ZEB1-dependent epithelial-to-mesenchymal transition that increases cell motility and promotes resistance
to traditional chemotherapy agents arguing strongly against the use of ‘drug holidays’ in cases of KRAS G13D
amplification.

20. Drug repurposing in neuroblastoma to overcome chemoresistance

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Madhu Kollareddy , Alice Sherard , Martin Michaelis , Jindrich Cintal Jr , Abderrahmane Kaidi ,
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Karim Malik 1
1 Cancer Epigenetics Laboratory, School of Cellular and Molecular Medicine, University of Bristol, Bristol, UK
2 Nuclear Dynamics Laboratory, School of Cellular and Molecular Medicine, University of Bristol, Bristol, UK
3 Centre for Molecular Processing and School of Biosciences, University of Kent, Canterbury, UK
4 Institut für Medizinische Virologie, Klinikum der Goethe-Universität, Frankfurt am Main, Germany