Disulfiram overcomes bortezomib and cytarabine resistance in Down-syndrome-associated acute myeloid leukemia cells

Ranjan Bista, David W. Lee, Oliver B. Pepper, David O. Azorsa, Robert J. Arceci, Eiman Aleem

Research output: Contribution to journalArticlepeer-review

Abstract

Background: Children with Down syndrome (DS) have increased risk for developing AML (DS-AMKL), and they usually experience severe therapy-related toxicities compared to non DS-AMKL. Refractory/relapsed disease has very poor outcome, and patients would benefit from novel, less toxic, therapeutic strategies that overcome resistance. Relapse/resistance are linked to cancer stem cells with high aldehyde dehydrogenase (ALDH) activity. The purpose of the present work was to study less toxic alternative therapeutic agents for relapsed/refractory DS-AMKL. Methods: Fourteen AML cell lines including the DS-AMKL CMY and CMK from relapsed/refractory AML were used. Cytarabine (Ara-C), bortezomib (BTZ), disulfiram/copper (DSF/Cu2+) were evaluated for cytotoxicity, depletion of ALDH-positive cells, and resistance. BTZ-resistant CMY and CMK variants were generated by continuous BTZ treatment. Cell viability was assessed using CellTiter-Glo®, ALDH activity by ALDELUORTM, and proteasome inhibition by western blot of ubiquitinated proteins and the Proteasome-Glo™ Chymotrypsin-Like (CT-like) assay, apoptosis by Annexin V Fluos/Propidium iodide staining, and mutations were detected using PCR, cloning and sequencing. Results: Ara-C-resistant AML cell lines were sensitive to BTZ and DSF/Cu2+. The Ara-C-resistant DS-AMKL CMY cells had a high percentage of ALDHbright “stem-like” populations that may underlie Ara-C resistance. One percent of these cells were still resistant to BTZ but sensitive to DSF/Cu2+. To understand the mechanism of BTZ resistance, BTZ resistant (CMY-BR) and (CMK-BR) were generated. A novel mutation PSMB5 Q62P underlied BTZ resistance, and was associated with an overexpression of the β5 proteasome subunit. BTZ-resistance conferred increased resistance to Ara-C due to G1 arrest in the CMY-BR cells, which protected the cells from S-phase damage by Ara-C. CMY-BR and CMK-BR cells were cross-resistant to CFZ and MG-132 but sensitive to DSF/Cu2+. In this setting, DSF/Cu2+ induced apoptosis and proteasome inhibition independent of CT-like activity inhibition. Conclusions: We provide evidence that DSF/Cu2+ overcomes Ara-C and BTZ resistance in cell lines from DS-AMKL patients. A novel mutation underlying BTZ resistance was detected that may identify BTZ-resistant patients, who may not benefit from treatment with CFZ or Ara-C, but may be responsive to DSF/Cu2+. Our findings support the clinical development of DSF/Cu2+ as a less toxic efficacious treatment approach in patients with relapsed/refractory DS-AMKL.

Original languageEnglish (US)
Pages (from-to)1-14
Number of pages14
JournalJournal of Experimental and Clinical Cancer Research
Volume36
Issue number1
DOIs
StatePublished - Feb 1 2017

Keywords

  • ALDH
  • Bortezomib
  • Chemoresistance
  • Cytarabine
  • Disulfiram
  • Down syndrome-associated AML
  • Relapsed acute myeloid leukemia

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

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