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Immunotherapy

Blockade of the PD-1/PD-L1 axis augments lysis of AML cells by the CD33/CD3 BiTE antibody construct AMG 330: reversing a T-cell-induced immune escape mechanism

Leukemia volume 30, pages 484–491 (2016)Cite this article

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Abstract

Bispecific T-cell engagers (BiTEs) are very effective in recruiting and activating T cells. We tested the cytotoxicity of the CD33/CD3 BiTE antibody construct AMG 330 on primary acute myeloid leukemia (AML) cells ex vivo and characterized parameters contributing to antileukemic cytolytic activity. The E:T ratio and the CD33 expression level significantly influenced lysis kinetics in long-term cultures of primary AML cells (n=38). AMG 330 induced T-cell-mediated proinflammatory conditions, favoring the upregulation of immune checkpoints on target and effector cells. Although not constitutively expressed at the time of primary diagnosis (n=123), PD-L1 was strongly upregulated on primary AML cells upon AMG 330 addition to ex vivo cultures (n=27, P<0.0001). This phenomenon was cytokine-driven as the sole addition of interferon (IFN)-γ and tumor necrosis factor-α also induced expression. Through blockade of the PD-1/PD-L1 interaction, AMG 330-mediated lysis (n=9, P=0.03), T-cell proliferation (n=9, P=0.01) and IFN-γ secretion (n=8, P=0.008) were significantly enhanced. The combinatorial approach was most beneficial in settings of protracted AML cell lysis. Taken together, we have characterized a critical resistance mechanism employed by primary AML cells under AMG 330-mediated proinflammatory conditions. Our results support the evaluation of checkpoint molecules in upcoming clinical trials with AMG 330 to enhance BiTE antibody construct-mediated cytotoxicity.

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Acknowledgements

We thank Kerstin Lämmermann and Elke Luxenburger (Clinical Co-operation Group Immunotherapy, Helmholtz Zentrum München), Elke Habben, Sabine Reinkunz, Ewelina Zientara and Tanja Kröll (Laboratory for Leukemia Diagnostics, University Hospital Munich) for their excellent technical support, and Maike Fritschle (Helmholtz Zentrum München) for her support in in vivo engraftment experiments in mice, Maja Rothenberg-Thurley for providing experiments on clonal stability and AMGEN Research (Munich) GmbH for providing AMG 330 and control antibody. The study was supported by funding from AMGEN Research (Munich) GmbH and Gert Riethmüller (Emeritus from the Institute for Immunology, Ludwig-Maximilians-University, Munich, Germany).

Author contributions

CK was involved in research design and performed the experiments, collected, analyzed and interpreted the data and wrote the manuscript. FSL generated the data of PD-L1 expression at primary diagnosis, was involved in data interpretation and contributed to the writing of the manuscript. TK was involved in statistical analysis and helped in preparation of figures and tables. BV and IJ provided the AML NSG mouse model and performed the animal part of the in vivo engraftment experiment. KHM performed targeted multiplexed amplicon resequencing experiments. GR, PK, RK and GZ were involved in research design and data interpretation. GR, PK, RK, GZ and PB provided AMG 330 and the control antibody. WH, SS, KHM, KS, MF and TA provided patient characteristics including molecular and cytogenetic data. MS designed the research, interpreted the data and supervised the project.

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Authors and Affiliations

  1. Department of Internal Medicine III, LMU—Klinikum der Universität München, Munich, Germany

    C Krupka, F S Lichtenegger, T Köhnke, K H Metzeler, T Altmann, S Schneider, M Fiegl, K Spiekermann, W Hiddemann & M Subklewe

  2. Clinical Co-operation Group Immunotherapy at the Helmholtz Zentrum München, Munich, Germany

    C Krupka, F S Lichtenegger, T Köhnke, T Altmann & M Subklewe

  3. AMGEN Research (Munich) GmbH, Munich, Germany

    P Kufer, R Kischel, G Zugmaier & P A Bauerle

  4. Department of Gene Vectors, Helmholtz Zentrum München, German Research Center for Environmental Health, Munich, Germany

    B Vick & I Jeremias

  5. German Cancer Consortium (DKTK), Heidelberg, Germany

    B Vick, I Jeremias, K Spiekermann, W Hiddemann & M Subklewe

  6. German Cancer Research Center (DKFZ), Heidelberg, Germany

    B Vick, I Jeremias, K Spiekermann, W Hiddemann & M Subklewe

  7. Department of Oncology, Dr von Haunersches Kinderspital, Ludwig Maximilians-Universität (LMU), Munich, Germany

    I Jeremias

  8. Institute for Immunology, Ludwig-Maximilians-Universität, Munich, Germany

    G Riethmüller

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  1. C Krupka
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Correspondence to M Subklewe.

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Competing interests

PK, RK and GZ are employed by AMGEN Research (Munich) GmbH. PK, RK, GZ, PAB and GR were involved in invention and development of the antibody and are shareholders of Amgen Research GmbH. PAB was employed by AMGEN Research at the time the data were generated. The study was supported by funding from AMGEN Research GmbH and Gert Riethmüller (Emeritus from the Institute for Immunology, Ludwig-Maximilians-University, Munich, Germany). The remaining authors declare no conflict of interest.

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Krupka, C., Kufer, P., Kischel, R. et al. Blockade of the PD-1/PD-L1 axis augments lysis of AML cells by the CD33/CD3 BiTE antibody construct AMG 330: reversing a T-cell-induced immune escape mechanism. Leukemia 30, 484–491 (2016). https://doi.org/10.1038/leu.2015.214

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