TY - JOUR
T1 - R-Loop–Mediated ssDNA Breaks Accumulate following Short-Term Exposure to the HDAC Inhibitor Romidepsin
AU - Safari, Maryam
AU - Litman, Thomas
AU - Robey, Robert W.
AU - Aguilera, Andres
AU - Chakraborty, Arup R.
AU - Reinhold, William C.
AU - Basseville, Agnes
AU - Petrukhin, Lubov
AU - Scotto, Luigi
AU - O’Connor, Owen A.
AU - Pommier, Yves
AU - Fojo, Antonio T.
AU - Bates, Susan E.
N1 - Funding Information:
T. Litman reports personal fees from LEO Pharma A/S outside the submitted work. A. Basseville reports grants from the European Commission during the conduct of the study. O.A. O’Connor reports grants from Celgene outside the submitted work. S.E. Bates reports grants from Celgene during the conduct of the study; as well as a patent for Depsipeptide for therapy of kidney cancer issued and Deacetylase inhibitor therapy issued. No disclosures were reported by the other authors.
Funding Information:
The authors acknowledge the important and vital contributions of Victoria Luchenko, who died before this work was completed; her efforts were critical in establishing the dominant effect of DNA damage and importance of cell context in the response to HDAC inhibitors. The authors would like to thank Sudhir Varma for his help in analyzing the CellMiner data. The authors would like to acknowledge the support of the research and imaging facility at the James J. Peters VA Medical Center. Part of this work was supported by the Center for Cancer Research, Intramural Research Program of the National Cancer Institute; (1ZIABC010621–12; to S.E. Bates) and (BC-006161 and BC-006150; to W.C. Reinhold and Y. Pommier), and the European Research Council (ERC2014 AdG669898 TARLOOP; to A. Aguilera).
Publisher Copyright:
©2021 The Authors; Published by the American Association for Cancer Research
PY - 2021
Y1 - 2021
N2 - Histone deacetylase inhibitors (HDACi) induce hyperacetylation of histones by blocking HDAC catalytic sites. Despite regulatory approvals in hematological malignancies, limited solid tumor clinical activity has constrained their potential, arguing for better understanding of mechanisms of action (MOA). Multiple activities of HDACis have been demonstrated, dependent on cell context, beyond the canonical induction of gene expression. Here, using a clinically relevant exposure duration, we established DNA damage as the dominant signature using the NCI-60 cell line database and then focused on the mechanism by which hyperacetylation induces DNA damage. We identified accumulation of DNA–RNA hybrids (R-loops) following romidepsin-induced histone hyperacetylation, with single-stranded DNA (ssDNA) breaks detected by single-cell electrophoresis. Our data suggest that transcription-coupled base excision repair (BER) is involved in resolving ssDNA breaks that, when overwhelmed, evolve to lethal dsDNA breaks. We show that inhibition of BER proteins such as PARP will increase dsDNA breaks in this context. These studies establish accumulation of R-loops as a consequence of romidepsin-mediated histone hyperacetylation. We believe that the insights provided will inform design of more effective combination therapy with HDACis for treatment of solid tumors. Implications: Key HDAC inhibitor mechanisms of action remain unknown; we identify accumulation of DNA–RNA hybrids (R-loops) due to chromatin hyperacetylation that provokes single-stranded DNA damage as a first step toward cell death.
AB - Histone deacetylase inhibitors (HDACi) induce hyperacetylation of histones by blocking HDAC catalytic sites. Despite regulatory approvals in hematological malignancies, limited solid tumor clinical activity has constrained their potential, arguing for better understanding of mechanisms of action (MOA). Multiple activities of HDACis have been demonstrated, dependent on cell context, beyond the canonical induction of gene expression. Here, using a clinically relevant exposure duration, we established DNA damage as the dominant signature using the NCI-60 cell line database and then focused on the mechanism by which hyperacetylation induces DNA damage. We identified accumulation of DNA–RNA hybrids (R-loops) following romidepsin-induced histone hyperacetylation, with single-stranded DNA (ssDNA) breaks detected by single-cell electrophoresis. Our data suggest that transcription-coupled base excision repair (BER) is involved in resolving ssDNA breaks that, when overwhelmed, evolve to lethal dsDNA breaks. We show that inhibition of BER proteins such as PARP will increase dsDNA breaks in this context. These studies establish accumulation of R-loops as a consequence of romidepsin-mediated histone hyperacetylation. We believe that the insights provided will inform design of more effective combination therapy with HDACis for treatment of solid tumors. Implications: Key HDAC inhibitor mechanisms of action remain unknown; we identify accumulation of DNA–RNA hybrids (R-loops) due to chromatin hyperacetylation that provokes single-stranded DNA damage as a first step toward cell death.
U2 - 10.1158/1541-7786.MCR-20-0833
DO - 10.1158/1541-7786.MCR-20-0833
M3 - Journal article
C2 - 34050002
AN - SCOPUS:85111044165
VL - 19
SP - 1361
EP - 1374
JO - Molecular Cancer Research
JF - Molecular Cancer Research
SN - 1541-7786
IS - 8
ER -