p97/VCP inhibition causes excessive MRE11-dependent DNA end resection promoting cell killing after ionizing radiation

Susan Kilgas; Abhay Narayan Singh; Salome Paillas; Chee Kin Then; Ignacio Torrecilla; Judith Nicholson; Lisa Browning; Iolanda Vendrell; Rebecca Konietzny; Benedikt M. Kessler; Anne E. Kiltie; Kristijan Ramadan

doi:10.1016/j.celrep.2021.109153

p97/VCP inhibition causes excessive MRE11-dependent DNA end resection promoting cell killing after ionizing radiation

Susan Kilgas, Abhay Narayan Singh, Salome Paillas, Chee Kin Then, Ignacio Torrecilla, Judith Nicholson, Lisa Browning, Iolanda Vendrell, Rebecca Konietzny, Benedikt M. Kessler, Anne E. Kiltie^*, Kristijan Ramadan

^*Corresponding author for this work

The Rowett Institute of Nutrition and Health

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Abstract

The ATPase p97 is a central component of the ubiquitin-proteasome degradation system. p97 uses its ATPase activity and co-factors to extract ubiquitinated substrates from different cellular locations, including DNA lesions, thereby regulating DNA repair pathway choice. Here, we find that p97 physically and functionally interacts with the MRE11-RAD50-NBS1 (MRN) complex on chromatin and that inactivation of p97 blocks the disassembly of the MRN complex from the sites of DNA damage upon ionizing radiation (IR). The inhibition of p97 function results in excessive 5′-DNA end resection mediated by MRE11 that leads to defective DNA repair and radiosensitivity. In addition, p97 inhibition by the specific small-molecule inhibitor CB-5083 increases tumor cell killing following IR both in vitro and in vivo. Mechanistically, this is mediated via increased MRE11 nuclease accumulation. This suggests that p97 inhibitors might be exploited to improve outcomes for radiotherapy patients.

Original language	English
Article number	109153
Number of pages	25
Journal	Cell Reports
Volume	35
Issue number	8
DOIs	https://doi.org/10.1016/j.celrep.2021.109153
Publication status	Published - 25 May 2021

Keywords

bladder cancer
CB-5083
DNA damage
DNA double-strand break repair
homologous recombination
ionizing radiation
IR
MRE11
p97
single-strand annealing

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Kilgas, S., Singh, A. N., Paillas, S., Then, C. K., Torrecilla, I., Nicholson, J., Browning, L., Vendrell, I., Konietzny, R., Kessler, B. M., Kiltie, A. E., & Ramadan, K. (2021). p97/VCP inhibition causes excessive MRE11-dependent DNA end resection promoting cell killing after ionizing radiation. Cell Reports, 35(8), [109153]. https://doi.org/10.1016/j.celrep.2021.109153

@article{3e1606f8efea4a74a7a9bc8094121f2d,

title = "p97/VCP inhibition causes excessive MRE11-dependent DNA end resection promoting cell killing after ionizing radiation",

abstract = "The ATPase p97 is a central component of the ubiquitin-proteasome degradation system. p97 uses its ATPase activity and co-factors to extract ubiquitinated substrates from different cellular locations, including DNA lesions, thereby regulating DNA repair pathway choice. Here, we find that p97 physically and functionally interacts with the MRE11-RAD50-NBS1 (MRN) complex on chromatin and that inactivation of p97 blocks the disassembly of the MRN complex from the sites of DNA damage upon ionizing radiation (IR). The inhibition of p97 function results in excessive 5′-DNA end resection mediated by MRE11 that leads to defective DNA repair and radiosensitivity. In addition, p97 inhibition by the specific small-molecule inhibitor CB-5083 increases tumor cell killing following IR both in vitro and in vivo. Mechanistically, this is mediated via increased MRE11 nuclease accumulation. This suggests that p97 inhibitors might be exploited to improve outcomes for radiotherapy patients.",

keywords = "bladder cancer, CB-5083, DNA damage, DNA double-strand break repair, homologous recombination, ionizing radiation, IR, MRE11, p97, single-strand annealing",

author = "Susan Kilgas and Singh, {Abhay Narayan} and Salome Paillas and Then, {Chee Kin} and Ignacio Torrecilla and Judith Nicholson and Lisa Browning and Iolanda Vendrell and Rebecca Konietzny and Kessler, {Benedikt M.} and Kiltie, {Anne E.} and Kristijan Ramadan",

note = "Funding Information: This work was funded by Cancer Research UK (CRUK) program grant C5255/A23755 to A.E.K. Medical Research Council UK (MRC) program grant MC_PC 12001/1 (MC_UU_00001/1) and Breast Cancer Now (Grant No. 2019DecPR1406) to K.R. S.K. was supported by the MRC Oxford Institute of Radiation Oncology (OIRO) CRUK studentship. We thank Dr. Sovan Sarkar (Department of Oncology, University of Oxford) for generously providing DR-GFP U2OS cells. We thank Diogo Dias (Ludwig Cancer Research Institute, University of Oxford) for his technical advice on HR and SSA assays and assistance with the analysis. We thank Dr. Lisa Folkes and Alix Hampson for the high-performance liquid chromatography (HPLC) analysis of CB-5083 concentration in tissue extracts from CD-1 nude mice bearing subcutaneous RT112 tumors. We also thank the Oxford Radcliffe Biobank for providing us with human tissue sections. ",

year = "2021",

month = may,

day = "25",

doi = "10.1016/j.celrep.2021.109153",

language = "English",

volume = "35",

journal = "Cell Reports",

issn = "2211-1247",

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T1 - p97/VCP inhibition causes excessive MRE11-dependent DNA end resection promoting cell killing after ionizing radiation

AU - Kilgas, Susan

AU - Singh, Abhay Narayan

AU - Paillas, Salome

AU - Then, Chee Kin

AU - Torrecilla, Ignacio

AU - Nicholson, Judith

AU - Browning, Lisa

AU - Vendrell, Iolanda

AU - Konietzny, Rebecca

AU - Kessler, Benedikt M.

AU - Kiltie, Anne E.

AU - Ramadan, Kristijan

N1 - Funding Information: This work was funded by Cancer Research UK (CRUK) program grant C5255/A23755 to A.E.K. Medical Research Council UK (MRC) program grant MC_PC 12001/1 (MC_UU_00001/1) and Breast Cancer Now (Grant No. 2019DecPR1406) to K.R. S.K. was supported by the MRC Oxford Institute of Radiation Oncology (OIRO) CRUK studentship. We thank Dr. Sovan Sarkar (Department of Oncology, University of Oxford) for generously providing DR-GFP U2OS cells. We thank Diogo Dias (Ludwig Cancer Research Institute, University of Oxford) for his technical advice on HR and SSA assays and assistance with the analysis. We thank Dr. Lisa Folkes and Alix Hampson for the high-performance liquid chromatography (HPLC) analysis of CB-5083 concentration in tissue extracts from CD-1 nude mice bearing subcutaneous RT112 tumors. We also thank the Oxford Radcliffe Biobank for providing us with human tissue sections.

PY - 2021/5/25

Y1 - 2021/5/25

N2 - The ATPase p97 is a central component of the ubiquitin-proteasome degradation system. p97 uses its ATPase activity and co-factors to extract ubiquitinated substrates from different cellular locations, including DNA lesions, thereby regulating DNA repair pathway choice. Here, we find that p97 physically and functionally interacts with the MRE11-RAD50-NBS1 (MRN) complex on chromatin and that inactivation of p97 blocks the disassembly of the MRN complex from the sites of DNA damage upon ionizing radiation (IR). The inhibition of p97 function results in excessive 5′-DNA end resection mediated by MRE11 that leads to defective DNA repair and radiosensitivity. In addition, p97 inhibition by the specific small-molecule inhibitor CB-5083 increases tumor cell killing following IR both in vitro and in vivo. Mechanistically, this is mediated via increased MRE11 nuclease accumulation. This suggests that p97 inhibitors might be exploited to improve outcomes for radiotherapy patients.

AB - The ATPase p97 is a central component of the ubiquitin-proteasome degradation system. p97 uses its ATPase activity and co-factors to extract ubiquitinated substrates from different cellular locations, including DNA lesions, thereby regulating DNA repair pathway choice. Here, we find that p97 physically and functionally interacts with the MRE11-RAD50-NBS1 (MRN) complex on chromatin and that inactivation of p97 blocks the disassembly of the MRN complex from the sites of DNA damage upon ionizing radiation (IR). The inhibition of p97 function results in excessive 5′-DNA end resection mediated by MRE11 that leads to defective DNA repair and radiosensitivity. In addition, p97 inhibition by the specific small-molecule inhibitor CB-5083 increases tumor cell killing following IR both in vitro and in vivo. Mechanistically, this is mediated via increased MRE11 nuclease accumulation. This suggests that p97 inhibitors might be exploited to improve outcomes for radiotherapy patients.

KW - bladder cancer

KW - CB-5083

KW - DNA damage

KW - DNA double-strand break repair

KW - homologous recombination

KW - ionizing radiation

KW - IR

KW - MRE11

KW - p97

KW - single-strand annealing

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U2 - 10.1016/j.celrep.2021.109153

DO - 10.1016/j.celrep.2021.109153

M3 - Article

C2 - 34038735

AN - SCOPUS:85106522028

VL - 35

JO - Cell Reports

JF - Cell Reports

SN - 2211-1247

IS - 8

M1 - 109153

ER -

p97/VCP inhibition causes excessive MRE11-dependent DNA end resection promoting cell killing after ionizing radiation

Abstract

Keywords

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