Targeting the NPL4 Adaptor of p97/VCP Segregase by Disulfiram as an Emerging Cancer Vulnerability Evokes Replication Stress and DNA Damage while Silencing the ATR Pathway

Cells. 2020 Feb 18;9(2):469. doi: 10.3390/cells9020469.


Research on repurposing the old alcohol-aversion drug disulfiram (DSF) for cancer treatment has identified inhibition of NPL4, an adaptor of the p97/VCP segregase essential for turnover of proteins involved in multiple pathways, as an unsuspected cancer cell vulnerability. While we reported that NPL4 is targeted by the anticancer metabolite of DSF, the bis-diethyldithiocarbamate-copper complex (CuET), the exact, apparently multifaceted mechanism(s) through which the CuET-induced aggregation of NPL4 kills cancer cells remains to be fully elucidated. Given the pronounced sensitivity to CuET in tumor cell lines lacking the genome integrity caretaker proteins BRCA1 and BRCA2, here we investigated the impact of NPL4 targeting by CuET on DNA replication dynamics and DNA damage response pathways in human cancer cell models. Our results show that CuET treatment interferes with DNA replication, slows down replication fork progression and causes accumulation of single-stranded DNA (ssDNA). Such a replication stress (RS) scenario is associated with DNA damage, preferentially in the S phase, and activates the homologous recombination (HR) DNA repair pathway. At the same time, we find that cellular responses to the CuET-triggered RS are seriously impaired due to concomitant malfunction of the ATRIP-ATR-CHK1 signaling pathway that reflects an unorthodox checkpoint silencing mode through ATR (Ataxia telangiectasia and Rad3 related) kinase sequestration within the CuET-evoked NPL4 protein aggregates.

Keywords: ATR pathway; BRCA1; BRCA2; DNA damage; NPL4; disulfiram; replication stress; targeted cancer therapy.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing / metabolism
  • Alcohol Deterrents / pharmacology*
  • Ataxia Telangiectasia Mutated Proteins / antagonists & inhibitors*
  • Ataxia Telangiectasia Mutated Proteins / metabolism
  • Cell Line, Tumor
  • Checkpoint Kinase 1 / metabolism
  • DNA Damage / drug effects*
  • DNA Replication / drug effects*
  • DNA-Binding Proteins / metabolism
  • Disulfiram / pharmacology*
  • Humans
  • Neoplasms / metabolism*
  • Neoplasms / pathology
  • Nuclear Proteins / antagonists & inhibitors*
  • Nuclear Proteins / metabolism
  • Protein Aggregates / drug effects
  • Protein Aggregation, Pathological / chemically induced
  • Signal Transduction / drug effects
  • Valosin Containing Protein / metabolism


  • ATRIP protein, human
  • Adaptor Proteins, Signal Transducing
  • Alcohol Deterrents
  • DNA-Binding Proteins
  • NPLOC4 protein, human
  • Nuclear Proteins
  • Protein Aggregates
  • ATR protein, human
  • Ataxia Telangiectasia Mutated Proteins
  • CHEK1 protein, human
  • Checkpoint Kinase 1
  • VCP protein, human
  • Valosin Containing Protein
  • Disulfiram