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

• Published in: Journal of experimental & clinical cancer research 2017-02, Vol.36 (1), p.22, Article 22
• Main Authors: Bista, Ranjan, Lee, David W, Pepper, Oliver B, Azorsa, David O, Arceci, Robert J, Aleem, Eiman
• Format: Article
• Language: English
• Subjects: Adolescent; Adult; Aldehyde Dehydrogenase - genetics; Aldehyde Dehydrogenase - metabolism; Bortezomib - pharmacology; Cell Line, Tumor; Cell Proliferation - drug effects; Cell Survival - drug effects; Child; Child, Preschool; Cytarabine - administration & dosage; Cytarabine - pharmacology; Disulfiram - pharmacology; Down Syndrome - complications; Drug Resistance, Neoplasm - drug effects; Female; Humans; Infant; Leukemia, Myeloid, Acute - drug therapy; Leukemia, Myeloid, Acute - etiology; Leukemia, Myeloid, Acute - genetics; Leukemia, Myeloid, Acute - metabolism; Male; Middle Aged; Mutation; Proteasome Endopeptidase Complex - genetics; Young Adult
• ISSN: 1756-9966; 0392-9078; 1756-9966
• DOI: 10.1186/s13046-017-0493-5

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. 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/Cu ) 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 ALDELUOR , 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. Ara-C-resistant AML cell lines were sensitive to BTZ and DSF/Cu . The Ara-C-resistant DS-AMKL CMY cells had a high percentage of ALDH "stem-like" populations that may underlie Ara-C resistance. One percent of these cells were still resistant to BTZ but sensitive to DSF/Cu . 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/Cu . In this setting, DSF/Cu induced apoptosis and proteasome inhibition independent of CT-like activity inhibition. We provide evidence that DSF/Cu 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/Cu . Our findings support the clinical development of DSF/Cu as a less toxic efficacious treatment approach in patients with relapsed/refractory DS-AMKL.