Establishing and characterizing a novel doxorubicin-resistant acute myeloid leukaemia cell line

Abstract

Drug resistance is a major setback in cancer treatment, thus models to study its mechanisms are needed. Our work aimed to establish and characterize a resistant cell line from a sensitive acute myeloid leukaemia (AML) cell line – HL60 – by treating the sensitive cells with increasing concentrations of doxorubicin. We confirmed (cell viability assays) that the established subline, HL60-CDR, was resistant to doxorubicin for at least 30 days without drug treatment. The HL60-CDR cells were also resistant to three other drugs (cisplatin, etoposide and daunorubicin), exhibiting a multidrug resistant (MDR) profile. We verified (Western Blotting) that the MDR cells do not express drug efflux pumps, nor present altered expression of apoptotic proteins, when compared with the parental cell line. HL60-CDR cells presented alterations in the cell cycle profile, and in the expression levels of proteins involved in DNA repair mechanisms and drug metabolism, when compared with their drug sensitive counterpart. Proteomic analysis revealed that HL60-CDR cells presented an upregulation of proteins involved in oncogenic pathways, such as TSC2, PDPK1, Annexin A2, among others. Overall, we established an AML MDR subline – HL60-CDR – which presents several resistance mechanisms, providing an in vitro model to test new compounds to circumvent MDR in AML.

Keywords:

• Acute myeloid leukaemia
• cancer drug resistant cells
• chemotherapy
• doxorubicin
• drug treatment
• multidrug resistance

Acknowledgements

The M.H.V. group is supported by FEDER – Fundo Europeu de Desenvolvimento Regional through COMPETE 2020 and by FCT – Foundation for Science and Technology, in the framework of project POCI-01-0145-FEDER-030457. The mass spectrometry technique was performed at the i3S Proteomics Scientific Platform with the assistance of Dr. Hugo Osório. This work was supported by the Portuguese mass spectrometry network, integrated in the National Roadmap of Research Infrastructures of Strategic Relevance (ROTEIRO/0028/2013; LISBOA-01-0145-FEDER-022125).

Disclosure statement

No potential conflict of interest was reported by the authors.

Author contributions

The contribution of the authors are as follows: Conceptualization, C.P.R.X. and M.H.V.; Methodology, I.C., H.B., V.L.-R., C.P.R.X.; Formal analysis, I.C., C.P.R.X.; Resources, M.H.V.; Writing – original draft preparation, I.C., C.P.R.X.; Writing – review and editing, C.P.R.X. and M.H.V.; Funding acquisition, M.H.V.; Supervision: M.H.V. All authors have read and agreed to the published version of the manuscript.

Data availability statement

All data generated or analyzed during this study are included in this published article.

Additional information

Funding

Cristina P. R. Xavier is supported by the Fundação para a Ciência e Tecnologia (FCT) and Fundo Social Europeu (FSE), Portugal, through the post-doc grant SFRH/BPD/122871/2016.