Abstract

The cytostatic deoxycytidine analog cytarabine (ara-C) is the most active agent available against acute myelogenous leukemia (AML). Together with anthracyclines, ara-C forms the backbone of AML treatment for children and adults1. In AML, both the cytotoxicity of ara-C in vitro and the clinical response to ara-C therapy are correlated with the ability of AML blasts to accumulate the active metabolite ara-C triphosphate (ara-CTP)2,3,4,5, which causes DNA damage through perturbation of DNA synthesis6. Differences in expression levels of known transporters or metabolic enzymes relevant to ara-C only partially account for patient-specific differential ara-CTP accumulation in AML blasts and response to ara-C treatment7,8,9. Here we demonstrate that the deoxynucleoside triphosphate (dNTP) triphosphohydrolase SAM domain and HD domain 1 (SAMHD1) promotes the detoxification of intracellular ara-CTP pools. Recombinant SAMHD1 exhibited ara-CTPase activity in vitro, and cells in which SAMHD1 expression was transiently reduced by treatment with the simian immunodeficiency virus (SIV) protein Vpx were dramatically more sensitive to ara-C-induced cytotoxicity. CRISPR–Cas9-mediated disruption of the gene encoding SAMHD1 sensitized cells to ara-C, and this sensitivity could be abrogated by ectopic expression of wild-type (WT), but not dNTPase-deficient, SAMHD1. Mouse models of AML lacking SAMHD1 were hypersensitive to ara-C, and treatment ex vivo with Vpx sensitized primary patient-derived AML blasts to ara-C. Finally, we identified SAMHD1 as a risk factor in cohorts of both pediatric and adult patients with de novo AML who received ara-C treatment. Thus, SAMHD1 expression levels dictate patient sensitivity to ara-C, providing proof-of-concept that the targeting of SAMHD1 by Vpx could be an attractive therapeutic strategy for potentiating ara-C efficacy in hematological malignancies.

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Acknowledgements

We are grateful to M. Wickström, N. Eissler and L. Elfman for consulting and assistance with xenotransplantation. We thank S. Olsson and M. Gustafsson for assistance with animal work. We would like to thank I. Bodin for help with paraffin-embedding and processing of tumor tissue and A. Björklund for assistance with primary AML samples. We acknowledge M. Nordenskjöld and S. Eriksson for laboratory assistance, D. Gavhed and K. Edfeldt for administrative assistance. We thank D. Trono for the gift of the pMD.G plasmid. We would like to thank J. Cinatl for initial discussions. We would like to express our gratitude to NCI's Office of Cancer Genomics (OCG), The Cancer Genome Atlas (TCGA) and Therapeutically Applicable Research To Generate Effective Treatments initiative (TARGET) for granting public access to their AML databases, and to the Broad Institute for access to the Cancer Therapeutics Response Portal (CTRP). This work was supported by grants from the Swedish Children's Cancer Foundation (TJ2016-0040 (to N.H.); 2015-0005 (to J.-I.H.); PR2015-0009 (to D.G.); and PR2013-0002 and PR2014-0048 (both to T.H.)), the Swedish Cancer Society (CAN 2016/837 to J.W.; CAN 2014/814 to S.L.; CAN 2015/768 to D.G.; CAN 2013/396 to J.-I.H.; and CAN 2012/770 and CAN 2015/255 to T.H.), the Swedish Research Council (2014-1839 to M.U.; 2015-02498 to S.L.; 2012-2037 to D.G.; and 2012-5935 and 2013-3791 to T.H.), Radiumhemmet's Research Foundations (154242 to G.R. and 144063 to D.G.), the Knut and Alice Wallenberg Foundation (KAW2014.0273 to T.H.), the Swedish Pain Relief Foundation (SSF/01-05 to T.H.), the Torsten and Ragnar Söderberg Foundation (to T.H.), the David and Astrid Hagelén Foundation (C24702193 to B.D.G.P.) and the Stockholm County Council (ALF project) (20150353 to S.L. and 20150016 to J.-I.H.). This work was supported by the German Research Foundation (DFG) (SCHA1950/1-1 to T.S.) and partially through the Federal Ministry of Education and Research of Germany (BMBF)–supported Immunoquant project (0316170 C to T.S.) and HIVERA: EURECA project (01KI1307B to T.S.). S.G.R. is supported by an EMBO Long-Term Fellowship (ALTF-605-2014). Chemical Biology Consortium Sweden is funded by the Swedish Research Council, Science for Life Laboratories and Karolinska Institutet (829-2009-6241 to H.A. and T.L.).

Author information

Author notes

    • José M Calderón-Montaño
    •  & Thomas Lundbäck

    Present address: Andalusian Molecular Biology and Regenerative Medicine Centre CABIMER, University of Seville-CSIC, Seville, Spain (J.M.C.-M.) and AstraZeneca R&D, Mölndal, Sweden (T.L.).

    • Nikolas Herold
    •  & Sean G Rudd

    These authors contributed equally to this work.

Affiliations

  1. Childhood Cancer Research Unit, Department of Women's and Children's Health, Karolinska Institutet, and Karolinska University Hospital, Stockholm, Sweden.

    • Nikolas Herold
    • , Linda Ljungblad
    • , Ida Hed Myrberg
    • , Bianca Tesi
    • , Julia Bladh
    • , Mats Heyman
    • , Per Kogner
    •  & Jan-Inge Henter
  2. Science for Life Laboratory, Division of Translational Medicine and Chemical Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.

    • Sean G Rudd
    • , Kumar Sanjiv
    • , Cynthia B J Paulin
    • , Anna Hagenkort
    • , Brent D G Page
    • , José M Calderón-Montaño
    • , Olga Loseva
    • , Ann-Sofie Jemth
    • , Hanna Axelsson
    • , Nicholas C K Valerie
    • , Andreas Höglund
    • , Elisée Wiita
    • , Ulrika Warpman-Berglund
    • , Thomas Lundbäck
    •  & Thomas Helleday
  3. Department of Medicine, Center of Hematology and Regenerative Medicine, Karolinska Hospital and Karolinska Institutet, Stockholm, Sweden.

    • Yaser Heshmati
    • , Julian Walfridsson
    •  & Sören Lehmann
  4. Department of Infectious Diseases, Virology, University Hospital Heidelberg, Heidelberg, Germany.

    • Juliane Kutzner
    • , Lorenzo Bulli
    •  & Torsten Schaller
  5. Chemical Biology Consortium, Stockholm, Sweden.

    • Hanna Axelsson
    •  & Thomas Lundbäck
  6. Division of Pediatrics, Department of Clinical Science, Intervention and Technology, Karolinska Institutet, Stockholm, Sweden.

    • Mikael Sundin
  7. Paediatric Blood Disorders, Immunodeficiency and Stem Cell Transplantation, Astrid Lindgren Children's Hospital, Karolinska University Hospital, Stockholm, Sweden.

    • Mikael Sundin
  8. Department of Oncology-Pathology, Karolinska Institutet, Stockholm, Sweden.

    • Michael Uhlin
    • , Georgios Rassidakis
    • , Katja Pokrovskaja Tamm
    •  & Dan Grandér
  9. Department of Clinical Immunology and Transfusion Medicine, Karolinska University Hospital, Stockholm, Sweden.

    • Michael Uhlin
  10. Department of Medical Sciences, Uppsala University, Uppsala, Sweden.

    • Sören Lehmann

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Contributions

N.H. and T.S. conceived the study. N.H., S.G.R., J.-I.H., T.H. and T.S. wrote the manuscript, which was revised by all authors. N.H., J.K., L.B. and T.S. created CRISPR–Cas9 knockout cell lines and Vpx-VLPs. N.H., S.G.R., C.B.J.P, E.W., J.K., J.B. and T.S. designed and performed the proliferation inhibition assays for THP-1, HuT-78 and HL-60 cells. Experiments with primary AML blasts and hematopoietic stem cells were planned by N.H., J.W., S.L., M.U., M.S., S.G.R., N.C.K.V., G.R., K.P.T., M.H. and D.G., and proliferation inhibition and apoptosis assays for blasts treated with Vpx-VLPs were performed by N.H., Y.H. and H.A. N.H., L.L., P.K. and T.S. designed the animal experiments. A.H. and U.W.-B. established the orthotopic AML animal model. N.H., L.L. and K.S. performed the animal experiments. N.H., I.H.M., B.T. and J.I.H. analysed TCGA and TARGET data. A.S.J. produced the expression construct, and O.L. purified recombinant SAMHD1. A.-S.J., T.L., S.G.R. and C.B.J.P. established the in vitro SAMHD1 activity assay, and subsequent experiments were performed by S.G.R. and C.B.J.P. N.H., S.G.R., B.D.G.P. and A.H. conceived the ara-CTP pool and ara-C DNA incorporation assays, and the respective experiments were performed by A.H. and B.D.G.P. S.G.R. and J.M.C.-M. performed DNA damage-response experiments. S.G.R. performed DNA content analysis. N.H., J.K., L.B. and T.S. performed kinetic analysis on Vpx-mediated SAMHD1 degradation.

Competing interests

T.L. is presently employed with AstraZeneca.

Corresponding authors

Correspondence to Nikolas Herold or Torsten Schaller.

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DOI

https://doi.org/10.1038/nm.4265

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