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Disulfiram Overcomes Cisplatin Resistance in Human Embryonal Carcinoma Cells.

ABSTRACT: Cisplatin resistance in testicular germ cell tumors (TGCTs) is a clinical challenge. We investigated the underlying mechanisms associated with cancer stem cell (CSC) markers and modalities circumventing the chemoresistance. Chemoresistant models (designated as CisR) of human embryonal carcinoma cell lines NTERA-2 and NCCIT were derived and characterized using flow cytometry, gene expression, functional and protein arrays. Tumorigenicity was determined on immunodeficient mouse model. Disulfiram was used to examine chemosensitization of resistant cells. ALDH1A3 isoform expression was evaluated by immunohistochemistry in 216 patients' tissue samples. Chemoresistant cells were significantly more resistant to cisplatin, carboplatin and oxaliplatin compared to parental cells. NTERA-2 CisR cells exhibited altered morphology and increased tumorigenicity. High ALDH1A3 expression and increased ALDH activity were detected in both refractory cell lines. Disulfiram in combination with cisplatin showed synergy for NTERA-2 CisR and NCCIT CisR cells and inhibited growth of NTERA-2 CisR xenografts. Significantly higher ALDH1A3 expression was detected in TGCTs patients' tissue samples compared to normal testicular tissue. We characterized novel clinically relevant model of chemoresistant TGCTs, for the first time identified the ALDH1A3 as a therapeutic target in TGCTs and more importantly, showed that disulfiram represents a viable treatment option for refractory TGCTs.

SUBMITTER: Schmidtova S

PROVIDER: S-EPMC6769487 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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Disulfiram Overcomes Cisplatin Resistance in Human Embryonal Carcinoma Cells.

Schmidtova Silvia S Kalavska Katarina K Gercakova Katarina K Cierna Zuzana Z Miklikova Svetlana S Smolkova Bozena B Buocikova Verona V Miskovska Viera V Durinikova Erika E Burikova Monika M Chovanec Michal M Matuskova Miroslava M Mego Michal M Kucerova Lucia L

Cancers 20190822 9

Cisplatin resistance in testicular germ cell tumors (TGCTs) is a clinical challenge. We investigated the underlying mechanisms associated with cancer stem cell (CSC) markers and modalities circumventing the chemoresistance. Chemoresistant models (designated as CisR) of human embryonal carcinoma cell lines NTERA-2 and NCCIT were derived and characterized using flow cytometry, gene expression, functional and protein arrays. Tumorigenicity was determined on immunodeficient mouse model. Disulfiram w ...[more]

PMID: 31443351

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Project description:BackgroundChildren 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.MethodsFourteen 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.ResultsAra-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.ConclusionsWe 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.

Dataset Information

Disulfiram Overcomes Cisplatin Resistance in Human Embryonal Carcinoma Cells.

Publications

Disulfiram Overcomes Cisplatin Resistance in Human Embryonal Carcinoma Cells.

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