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

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. © 2017 the author(s).