Inhibition of RNA polymerase I transcription initiation by CX-5461 activates non-canonical ATM/ATR signaling

// Jaclyn Quin 1, 2, 11 , Keefe T. Chan 1 , Jennifer R. Devlin 1, 12 , Donald P. Cameron 1, 3 , Jeannine Diesch 1, 13 , Carleen Cullinane 1 , Jessica Ahern 1 , Amit Khot 1 , Nadine Hein 4 , Amee J. George 4, 5, 6 , Katherine M Hannan 2, 4 , Gretchen Poortinga 1, 7 , Karen E. Sheppard 1, 2, 3 , Kum Kum Khanna 8 , Ricky W. Johnstone 1, 3, 5 , Denis Drygin 9 , Grant A. McArthur 1, 3, 5, 7 , Richard B. Pearson 1, 2, 3, 10 , Elaine Sanij 1, 5, * , Ross D. Hannan 1, 2, 3, 4, 6, 10, * 1 Research Division, Peter MacCallum Cancer Centre, St. Andrews Place, East Melbourne, Victoria, Australia 2 Department of Biochemistry and Molecular Biology, University of Melbourne, Parkville, Victoria, Australia 3 Sir Peter MacCallum Department of Oncology, University of Melbourne, Parkville, Victoria, Australia 4 The John Curtin School of Medical Research, Australian National University, Acton, ACT, Australia 5 Department of Pathology, University of Melbourne, Parkville, Victoria, Australia 6 School of Biomedical Sciences, University of Queensland, Brisbane, Queensland, Australia 7 Department of Medicine, St Vincent’s Hospital, University of Melbourne, Fitzroy, Victoria, Australia 8 QIMR Berghofer Medical Research Institute, Brisbane City, Qld, Australia 9 Pimera Inc, San Diego, CA, USA 10 Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria, Australia 11 Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, Stockholm, Sweden 12 Institute for Molecular Medicine Finland, Biomedicum 2, Helsinki, Finland 13 Josep Carreras Institute for Leukaemia Research (IJC), Campus ICO-HGTP, Badalona, Barcelona, Spain * These authors contributed equally to this work Correspondence to: Elaine Sanij, email: elaine.sanij@petermac.org Ross D. Hannan, email: ross.hannan@anu.edu.au Keywords: RNA polymerase I, rDNA, CX-5461, nucleolar stress response, DNA damage signaling Received: March 17, 2016      Accepted: June 13, 2016      Published: July 06, 2016 ABSTRACT RNA polymerase I (Pol I)-mediated transcription of the ribosomal RNA genes (rDNA) is confined to the nucleolus and is a rate-limiting step for cell growth and proliferation. Inhibition of Pol I by CX-5461 can selectively induce p53-mediated apoptosis of tumour cells in vivo . Currently, CX-5461 is in clinical trial for patients with advanced haematological malignancies (Peter Mac, Melbourne). Here we demonstrate that CX-5461 also induces p53-independent cell cycle checkpoints mediated by ATM/ATR signaling in the absence of DNA damage. Further, our data demonstrate that the combination of drugs targeting ATM/ATR signaling and CX-5461 leads to enhanced therapeutic benefit in treating p53-null tumours in vivo , which are normally refractory to each drug alone. Mechanistically, we show that CX-5461 induces an unusual chromatin structure in which transcriptionally competent relaxed rDNA repeats are devoid of transcribing Pol I leading to activation of ATM signaling within the nucleoli. Thus, we propose that acute inhibition of Pol transcription initiation by CX-5461 induces a novel nucleolar stress response that can be targeted to improve therapeutic efficacy.