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Journal of Experimental & Clinical Cancer Research

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Open Access 01-12-2021 | Acute Myeloid Leukemia | Research

Differences between intrinsic and acquired nucleoside analogue resistance in acute myeloid leukaemia cells

Authors: Tamara Rothenburger, Dominique Thomas, Yannick Schreiber, Paul R. Wratil, Tamara Pflantz, Kirsten Knecht, Katie Digianantonio, Joshua Temple, Constanze Schneider, Hanna-Mari Baldauf, Katie-May McLaughlin, Florian Rothweiler, Berna Bilen, Samira Farmand, Denisa Bojkova, Rui Costa, Nerea Ferreirós, Gerd Geisslinger, Thomas Oellerich, Yong Xiong, Oliver T. Keppler, Mark N. Wass, Martin Michaelis, Jindrich Cinatl Jr

Published in: Journal of Experimental & Clinical Cancer Research | Issue 1/2021

Abstract

Background

SAMHD1 mediates resistance to anti-cancer nucleoside analogues, including cytarabine, decitabine, and nelarabine that are commonly used for the treatment of leukaemia, through cleavage of their triphosphorylated forms. Hence, SAMHD1 inhibitors are promising candidates for the sensitisation of leukaemia cells to nucleoside analogue-based therapy. Here, we investigated the effects of the cytosine analogue CNDAC, which has been proposed to be a SAMHD1 inhibitor, in the context of SAMHD1.

Methods

CNDAC was tested in 13 acute myeloid leukaemia (AML) cell lines, in 26 acute lymphoblastic leukaemia (ALL) cell lines, ten AML sublines adapted to various antileukaemic drugs, 24 single cell-derived clonal AML sublines, and primary leukaemic blasts from 24 AML patients. Moreover, 24 CNDAC-resistant sublines of the AML cell lines HL-60 and PL-21 were established. The SAMHD1 gene was disrupted using CRISPR/Cas9 and SAMHD1 depleted using RNAi, and the viral Vpx protein. Forced DCK expression was achieved by lentiviral transduction. SAMHD1 promoter methylation was determined by PCR after treatment of genomic DNA with the methylation-sensitive HpaII endonuclease. Nucleoside (analogue) triphosphate levels were determined by LC-MS/MS. CNDAC interaction with SAMHD1 was analysed by an enzymatic assay and by crystallisation.

Results

Although the cytosine analogue CNDAC was anticipated to inhibit SAMHD1, SAMHD1 mediated intrinsic CNDAC resistance in leukaemia cells. Accordingly, SAMHD1 depletion increased CNDAC triphosphate (CNDAC-TP) levels and CNDAC toxicity. Enzymatic assays and crystallisation studies confirmed CNDAC-TP to be a SAMHD1 substrate. In 24 CNDAC-adapted acute myeloid leukaemia (AML) sublines, resistance was driven by DCK (catalyses initial nucleoside phosphorylation) loss. CNDAC-adapted sublines displayed cross-resistance only to other DCK substrates (e.g. cytarabine, decitabine). Cell lines adapted to drugs not affected by DCK or SAMHD1 remained CNDAC sensitive. In cytarabine-adapted AML cells, increased SAMHD1 and reduced DCK levels contributed to cytarabine and CNDAC resistance.

Conclusion

Intrinsic and acquired resistance to CNDAC and related nucleoside analogues are driven by different mechanisms. The lack of cross-resistance between SAMHD1/ DCK substrates and non-substrates provides scope for next-line therapies after treatment failure.

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Title: Differences between intrinsic and acquired nucleoside analogue resistance in acute myeloid leukaemia cells
Authors: Tamara Rothenburger
Dominique Thomas
Yannick Schreiber
Paul R. Wratil
Tamara Pflantz
Kirsten Knecht
Katie Digianantonio
Joshua Temple
Constanze Schneider
Hanna-Mari Baldauf
Katie-May McLaughlin
Florian Rothweiler
Berna Bilen
Samira Farmand
Denisa Bojkova
Rui Costa
Nerea Ferreirós
Gerd Geisslinger
Thomas Oellerich
Yong Xiong
Oliver T. Keppler
Mark N. Wass
Martin Michaelis
Jindrich Cinatl Jr
Publication date: 01-12-2021
Publisher: BioMed Central
Keywords: Acute Myeloid Leukemia
Acute Lymphoblastic Leukemia
Leukemia
Published in: Journal of Experimental & Clinical Cancer Research / Issue 1/2021
Electronic ISSN: 1756-9966
DOI: https://doi.org/10.1186/s13046-021-02093-4

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