Skip to main content
SpringerLink
Log in

Expression of IL-15 in NK cells results in rapid enrichment and selective cytotoxicity of gene-modified effectors that carry a tumor-specific antigen receptor

  • Original article
  • Published:
Cancer Immunology, Immunotherapy Aims and scope Submit manuscript

Abstract

Natural killer (NK) cells hold promise for adoptive cancer immunotherapy but are dependent on cytokines such as interleukin (IL)-2 for growth and cytotoxicity. Here, we investigated the consequences of ectopic expression of IL-15 in human NK cells. IL-2 and IL-15 belong to the common γ chain family of cytokines and have overlapping activities. Transduction of clinically applicable NK-92 cells with lentiviral vectors encoding human IL-15 resulted in predominantly intracellular expression of the cytokine, and STAT5 activation, proliferation and cytotoxicity of the producer cells in the absence of IL-2. Growth of non-transduced bystander cells was not supported, allowing rapid enrichment of gene-modified cells solely by IL-2 withdrawal. This was also the case upon transduction of NK-92 and NKL cells with a bicistronic lentiviral vector encoding IL-15 and a chimeric antigen receptor (CAR) targeting the pancarcinoma antigen EpCAM. Effector cells co-expressing CAR and IL-15 continued to proliferate in the absence of exogenous cytokines and displayed high and selective cell-killing activity against EpCAM-expressing breast carcinoma cells that were resistant to the natural cytotoxicity of unmodified NK cells. This strategy facilitates rapid isolation and continuous expansion of retargeted NK cells and may extend their potential clinical utility.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  1. Gross G, Waks T, Eshhar Z (1989) Expression of immunoglobulin-T-cell receptor chimeric molecules as functional receptors with antibody-type specificity. Proc Natl Acad Sci USA 86:10024–10028

    Article  PubMed  CAS  Google Scholar 

  2. Uherek C, Groner B, Wels W (2001) Chimeric antigen receptors for the retargeting of cytotoxic effector cells. J Hematother Stem Cell Res 10:523–534. doi:10.1089/15258160152509136

    Article  PubMed  CAS  Google Scholar 

  3. Hombach A, Abken H (2007) Costimulation tunes tumor-specific activation of redirected T cells in adoptive immunotherapy. Cancer Immunol Immunother 56:731–737

    Article  PubMed  Google Scholar 

  4. Ngo MC, Rooney CM, Howard JM, Heslop HE (2011) Ex vivo gene transfer for improved adoptive immunotherapy of cancer. Hum Mol Genet 20:R93–R99

    Article  PubMed  CAS  Google Scholar 

  5. Porter DL, Levine BL, Kalos M, Bagg A, June CH (2011) Chimeric antigen receptor-modified T cells in chronic lymphoid leukemia. N Engl J Med 365:725–733. doi:10.1056/NEJMoa1103849

    Article  PubMed  CAS  Google Scholar 

  6. Uherek C, Tonn T, Uherek B, Becker S, Schnierle B, Klingemann HG, Wels W (2002) Retargeting of natural killer-cell cytolytic activity to ErbB2-expressing cancer cells results in efficient and selective tumor cell destruction. Blood 100:1265–1273

    PubMed  CAS  Google Scholar 

  7. Wels W, Biburger M, Müller T, Dälken B, Giesübel U, Tonn T, Uherek C (2004) Recombinant immunotoxins and retargeted killer cells: employing engineered antibody fragments for tumor-specific targeting of cytotoxic effectors. Cancer Immunol Immunother 53:217–226. doi:10.1007/s00262-003-0482-8

    Article  PubMed  CAS  Google Scholar 

  8. Imai C, Iwamoto S, Campana D (2005) Genetic modification of primary natural killer cells overcomes inhibitory signals and induces specific killing of leukemic cells. Blood 106:376–383

    Article  PubMed  CAS  Google Scholar 

  9. Pegram HJ, Kershaw MH, Darcy PK (2009) Genetic modification of natural killer cells for adoptive cellular immunotherapy. Immunotherapy 1:623–630. doi:10.2217/imt.09.36

    PubMed  CAS  Google Scholar 

  10. Lanier LL (2008) Up on the tightrope: natural killer cell activation and inhibition. Nat Immunol 9:495–502. doi:10.1038/ni1581

    Article  PubMed  CAS  Google Scholar 

  11. Klingemann HG (2005) Natural killer cell-based immunotherapeutic strategies. Cytotherapy 7:16–22

    Article  PubMed  CAS  Google Scholar 

  12. Tonn T, Becker S, Esser R, Schwabe D, Seifried E (2001) Cellular immunotherapy of malignancies using the clonal natural killer cell line NK-92. J Hematother Stem Cell Res 10:535–544. doi:10.1089/15258160152509145

    Article  PubMed  CAS  Google Scholar 

  13. Arai S, Meagher R, Swearingen M, Myint H, Rich E, Martinson J, Klingemann H (2008) Infusion of the allogeneic cell line NK-92 in patients with advanced renal cell cancer or melanoma: a phase I trial. Cytotherapy 10:625–632. doi:10.1080/14653240802301872

    Article  PubMed  CAS  Google Scholar 

  14. Tavri S, Jha P, Meier R, Henning TD, Müller T, Hostetter D, Knopp C, Johansson M, Reinhart V, Boddington S, Sista A, Wels WS, Daldrup-Link HE (2009) Optical imaging of cellular immunotherapy against prostate cancer. Mol Imaging 8:15–26

    PubMed  Google Scholar 

  15. Müller T, Uherek C, Maki G, Chow KU, Schimpf A, Klingemann HG, Tonn T, Wels WS (2008) Expression of a CD20-specific chimeric antigen receptor enhances cytotoxic activity of NK cells and overcomes NK-resistance of lymphoma and leukemia cells. Cancer Immunol Immunother 57:411–423. doi:10.1007/s00262-007-0383-3

    Article  PubMed  Google Scholar 

  16. Esser R, Müller T, Stefes D, Kloess S, Seidel D, Gillies SD, Aperlo-Iffland C, Huston JS, Uherek C, Schönfeld K, Tonn T, Huebener N, Lode HN, Koehl U, Wels WS (2011) NK cells engineered to express a GD(2) -specific antigen receptor display built-in ADCC-like activity against tumor cells of neuroectodermal origin. J Cell Mol Med (in press). doi:10.1111/j.1582-4934.2011.01343.x

  17. Maki G, Klingemann HG, Martinson JA, Tam YK (2001) Factors regulating the cytotoxic activity of the human natural killer cell line, NK-92. J Hematother Stem Cell Res 10:369–383. doi:10.1089/152581601750288975

    Article  PubMed  CAS  Google Scholar 

  18. Overwijk WW, Schluns KS (2009) Functions of gammaC cytokines in immune homeostasis: current and potential clinical applications. Clin Immunol 132:153–165. doi:10.1016/j.clim.2009.03.512

    Article  PubMed  CAS  Google Scholar 

  19. Yamasaki S, Maeda M, Ohshima K, Kikuchi M, Otsuka T, Harada M (2004) Growth and apoptosis of human natural killer cell neoplasms: role of interleukin-2/15 signaling. Leuk Res 28:1023–1031

    Article  PubMed  CAS  Google Scholar 

  20. Zhang J, Sun R, Wei H, Tian Z (2004) Characterization of interleukin-15 gene-modified human natural killer cells: implications for adoptive cellular immunotherapy. Haematologica 89:338–347

    PubMed  CAS  Google Scholar 

  21. Becknell B, Caligiuri MA (2005) Interleukin-2, interleukin-15, and their roles in human natural killer cells. Adv Immunol 86:209–239. doi:10.1016/S0065-2776(04)86006-1

    Article  PubMed  CAS  Google Scholar 

  22. Robertson MJ, Cochran KJ, Cameron C, Le JM, Tantravahi R, Ritz J (1996) Characterization of a cell line, NKL, derived from an aggressive human natural killer cell leukemia. Exp Hematol 24:406–415

    PubMed  CAS  Google Scholar 

  23. Ball RK, Friis RR, Schoenenberger CA, Doppler W, Groner B (1988) Prolactin regulation of beta-casein gene expression and of a cytosolic 120-kd protein in a cloned mouse mammary epithelial cell line. EMBO J 7:2089–2095

    PubMed  CAS  Google Scholar 

  24. Demaison C, Parsley K, Brouns G, Scherr M, Battmer K, Kinnon C, Grez M, Thrasher AJ (2002) High-level transduction and gene expression in hematopoietic repopulating cells using a human immunodeficiency virus type 1-based lentiviral vector containing an internal spleen focus forming virus promoter. Hum Gene Ther 13:803–813. doi:10.1089/10430340252898984

    Article  PubMed  CAS  Google Scholar 

  25. Willuda J, Honegger A, Waibel R, Schubiger PA, Stahel R, Zangemeister-Wittke U, Plückthun A (1999) High thermal stability is essential for tumor targeting of antibody fragments: engineering of a humanized anti-epithelial glycoprotein-2 (epithelial cell adhesion molecule) single-chain Fv fragment. Cancer Res 59:5758–5767

    PubMed  CAS  Google Scholar 

  26. Zufferey R, Nagy D, Mandel RJ, Naldini L, Trono D (1997) Multiply attenuated lentiviral vector achieves efficient gene delivery in vivo. Nat Biotechnol 15:871–875. doi:10.1038/nbt0997-871

    Article  PubMed  CAS  Google Scholar 

  27. Zimmermann S, Wels W, Froesch BA, Gerstmayer B, Stahel RA, Zangemeister-Wittke U (1997) A novel immunotoxin recognising the epithelial glycoprotein-2 has potent antitumoural activity on chemotherapy-resistant lung cancer. Cancer Immunol Immunother 44:1–9

    Article  PubMed  CAS  Google Scholar 

  28. Stonier SW, Schluns KS (2010) Trans-presentation: a novel mechanism regulating IL-15 delivery and responses. Immunol Lett 127:85–92

    Article  PubMed  CAS  Google Scholar 

  29. Boissel L, Betancur M, Wels WS, Tuncer H, Klingemann H (2009) Transfection with mRNA for CD19 specific chimeric antigen receptor restores NK cell mediated killing of CLL cells. Leuk Res 33:1255–1259. doi:10.1016/j.leukres.2008.11.024

    Article  PubMed  CAS  Google Scholar 

  30. Hoyos V, Savoldo B, Quintarelli C, Mahendravada A, Zhang M, Vera J, Heslop HE, Rooney CM, Brenner MK, Dotti G (2010) Engineering CD19-specific T lymphocytes with interleukin-15 and a suicide gene to enhance their anti-lymphoma/leukemia effects and safety. Leukemia 24:1160–1170. doi:10.1038/leu.2010.75

    Article  PubMed  CAS  Google Scholar 

  31. Chmielewski M, Kopecky C, Hombach AA, Abken H (2011) IL-12 release by engineered T cells expressing chimeric antigen receptors can effectively muster an antigen-independent macrophage response on tumor cells that have shut down tumor antigen expression. Cancer Res 71:5697–5706. doi:10.1158/0008-5472.CAN-11-0103

    Article  PubMed  CAS  Google Scholar 

  32. Jiang W, Zhang J, Tian Z (2008) Functional characterization of interleukin-15 gene transduction into the human natural killer cell line NKL. Cytotherapy 10:265–274. doi:10.1080/14653240801965156

    Article  PubMed  CAS  Google Scholar 

  33. Kurys G, Tagaya Y, Bamford R, Hanover JA, Waldmann TA (2000) The long signal peptide isoform and its alternative processing direct the intracellular trafficking of interleukin-15. J Biol Chem 275:30653–30659. doi:10.1074/jbc.M002373200

    Article  PubMed  CAS  Google Scholar 

  34. Bergamaschi C, Jalah R, Kulkarni V, Rosati M, Zhang GM, Alicea C, Zolotukhin AS, Felber BK, Pavlakis GN (2009) Secretion and biological activity of short signal peptide IL-15 is chaperoned by IL-15 receptor alpha in vivo. J Immunol 183:3064–3072

    Article  PubMed  CAS  Google Scholar 

  35. Browder TM, Abrams JS, Wong PM, Nienhuis AW (1989) Mechanism of autocrine stimulation in hematopoietic cells producing interleukin-3 after retrovirus-mediated gene transfer. Mol Cell Biol 9:204–213

    PubMed  CAS  Google Scholar 

  36. Konstantinidis KV, Alici E, Aints A, Christensson B, Ljunggren HG, Dilber MS (2005) Targeting IL-2 to the endoplasmic reticulum confines autocrine growth stimulation to NK-92 cells. Exp Hematol 33:159–164. doi:10.1016/j.exphem.2004.11.003

    Article  PubMed  CAS  Google Scholar 

  37. Hombach A, Wieczarkowiecz A, Marquardt T, Heuser C, Usai L, Pohl C, Seliger B, Abken H (2001) Tumor-specific T cell activation by recombinant immunoreceptors: CD3 zeta signaling and CD28 costimulation are simultaneously required for efficient IL-2 secretion and can be integrated into one combined CD28/CD3 zeta signaling receptor molecule. J Immunol 167:6123–6131

    PubMed  CAS  Google Scholar 

  38. Munz M, Baeuerle PA, Gires O (2010) The emerging role of EpCAM in cancer and stem cell signaling. Cancer Res 69:5627–5629. doi:10.1158/0008-5472.CAN-09-0654

    Article  Google Scholar 

  39. van der Gun BT, Melchers LJ, Ruiters MH, de Leij LF, McLaughlin PM, Rots MG (2010) EpCAM in carcinogenesis: the good, the bad or the ugly. Carcinogenesis 31:1913–1921. doi:10.1093/carcin/bgq187

    Article  PubMed  Google Scholar 

  40. Tam YK, Miyagawa B, Ho VC, Klingemann HG (1999) Immunotherapy of malignant melanoma in a SCID mouse model using the highly cytotoxic natural killer cell line NK-92. J Hematother 8:281–290

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

This work was supported in part by grants from the Bundesministerium für Bildung und Forschung (BMBF) FKZ 01GU0805, the Deutsche Forschungsgemeinschaft (DFG) GRK1172, and the LOEWE Center for Cell and Gene Therapy Frankfurt (CGT). We thank Dr. Uwe Zangemeister-Wittke, University of Bern, for MOC31 antibody, Dr. Manuel Grez, Georg-Speyer-Haus, for helpful discussions and for providing lentiviral vectors and packaging constructs, Dr. Stefan Stein and Mr. Tevik Merovci, Georg-Speyer-Haus for help with flow cytometric cell sorting, and Mrs. Annemarie Schimpf, Georg-Speyer-Haus, for excellent technical assistance.

Conflict of interest

The authors declare that they have no conflict of interest.

Author information

Authors and Affiliations

  1. Chemotherapeutisches Forschungsinstitut Georg-Speyer-Haus, Paul-Ehrlich-Straße 42-44, 60596, Frankfurt am Main, Germany

    Christiane Sahm, Kurt Schönfeld & Winfried S. Wels

Authors
  1. Christiane Sahm
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Kurt Schönfeld
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Winfried S. Wels
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Winfried S. Wels.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (PDF 264 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Sahm, C., Schönfeld, K. & Wels, W.S. Expression of IL-15 in NK cells results in rapid enrichment and selective cytotoxicity of gene-modified effectors that carry a tumor-specific antigen receptor. Cancer Immunol Immunother 61, 1451–1461 (2012). https://doi.org/10.1007/s00262-012-1212-x

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00262-012-1212-x

Keywords

Search

Navigation