Article Text

Download PDFPDF

1160 Phosphonate-antibody-drug conjugates: a novel immunostimulatory class of ADCs driving inside-out activation of Vγ9Vδ2 T cells leading to selective tumor cell killing
  1. Mary J van Helden,
  2. Ronald C Elgersma,
  3. Daphne WJ van Kuppeveld,
  4. Dennis Waalboer,
  5. Désirée Damming,
  6. Ellen WH Santegoeds-Lenssen,
  7. Inge Leenders,
  8. Karin de Laat-Arts,
  9. Lilian Driessen-Engels,
  10. Marc CBC Paradé,
  11. Menno Winkel,
  12. Anja Scholzen,
  13. Daniëlle EJW van Wijk,
  14. Diels van den Dobbelsteen,
  15. Genny Filiciotto,
  16. Laura Assink,
  17. Myrthe Rouwette,
  18. Panagiota I Spantidea,
  19. Roel J Arends,
  20. Timo K van den Berg,
  21. Wendela A Kappers,
  22. Gijs Verheijden,
  23. Wim HA Dokter and
  24. Miranda MC van der Lee
  1. Byondis BV, Nijmegen, Gelderland, Netherlands
  • Journal for ImmunoTherapy of Cancer (JITC) preprint. The copyright holder for this preprint are the authors/funders, who have granted JITC permission to display the preprint. All rights reserved. No reuse allowed without permission.


Background Gamma delta (γδ) T cells are cytotoxic effector cells that can recognize and kill tumor cells in a major histocompatibility complex (MHC)-independent fashion. Their infiltration into malignant tissue correlates with a favorable prognosis in a broad range of cancers. Vγ9Vδ2 T cells are the predominant γδ T cell population in human peripheral blood and can sense accumulated phosphoantigens (pAgs) bound intracellularly to the BTN3A/BTN2A complex. Early clinical studies using aminobisphosphonates or synthetic pAgs, alone or in combination with a low dose of IL-2, have demonstrated expansion and activation of peripheral Vγ9Vδ2 T cells. Clinical efficacy, however, was variable, possibly due to the absence of tumor targeting and a very short half-life of these compounds.

Methods To circumvent these problems, we generated antibody-drug conjugates (ADCs) that deliver synthetic phosphonates selectively to tumor cells and drive inside-out activation of Vγ9Vδ2 T cells. The antibody moiety binds to a tumor-associated antigen (TAA), triggering internalization. The linker conjugating the phosphonate to the antibody is cleaved in lysosomes by proteases releasing the phosphonate payload which is then free to activate BTN3A/BTN2A complexes. The payload was conjugated to various TAA-targeting antibodies, including anti-CD123, anti-CD20, and anti-HER2 mAbs.

Results Multiple phosphonate-ADC-pretreated TAA-positive tumor cell lines were able to activate Vγ9Vδ2 T cells in a BTN3A-dependent fashion, leading to cytokine production, degranulation and killing of the tumor cells. The phosphonate-ADCs specifically activated Vγ9Vδ2 T cells in all tested healthy donors (N>40), but not other T cells. Stimulation of FcγR-positive immune cells through Fc-tail interactions were preserved.

Conclusions Overall, these immunostimulatory phosphonate-ADCs represent a promising novel approach to targeted therapy in the field of cancer.

This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See

Statistics from

Request Permissions

If you wish to reuse any or all of this article please use the link below which will take you to the Copyright Clearance Center’s RightsLink service. You will be able to get a quick price and instant permission to reuse the content in many different ways.