Cancer Immunology, Immunotherapy

, Volume 65, Issue 9, pp 1047–1059 | Cite as

Human NK cells maintain licensing status and are subject to killer immunoglobulin-like receptor (KIR) and KIR-ligand inhibition following ex vivo expansion

  • Wei Wang
  • Amy K. Erbe
  • Kory A. Alderson
  • Emily Phillips
  • Mikayla Gallenberger
  • Jacek Gan
  • Dario Campana
  • Jacquelyn A. Hank
  • Paul M. SondelEmail author
Original Article


Infusion of allogeneic NK cells is a potential immunotherapy for both hematopoietic malignancies and solid tumors. Interactions between killer immunoglobulin-like receptors (KIR) on human NK cells and KIR-ligands on tumor cells influence the magnitude of NK function. To obtain sufficient numbers of activated NK cells for infusion, one potent method uses cells from the K562 human erythroleukemia line that have been transfected to express activating 41BB ligand (41BBL) and membrane-bound interleukin 15 (mbIL15). The functional importance of KIRs on ex vivo expanded NK cells has not been studied in detail. We found that after a 12-day co-culture with K562-mbIL15-41BBL cells, expanded NK cells maintained inhibition specificity and prior in vivo licensing status determined by KIR/KIR-ligand interactions. Addition of an anti-CD20 antibody (rituximab) induced NK-mediated antibody-dependent cellular cytotoxicity and augmented killing of CD20+ target cells. However, partial inhibition induced by KIR/KIR-ligand interactions persisted. Finally, we found that extended co-cultures of NK cells with stimulatory cells transduced to express various KIR-ligands modified both the inhibitory and activating KIR repertoires of the expanded NK cell product. These studies demonstrate that the licensing interactions known to occur during NK ontogeny also influence NK cell function following NK expansion ex vivo with HLA-null stimulatory cells.


Ex vivo expansion NK cells Killer immunoglobulin-like receptors HLA ADCC 



Antibody-dependent cellular cytotoxicity


Activating killer immunoglobulin-like receptor








Inhibitory killer immunoglobulin-like receptor


Killer immunoglobulin-like receptor


Membrane-bound interleukin 15


Major histocompatibility complex


Natural cytotoxicity receptor




41BB ligand



The authors thank all healthy donors participating in this study. Thanks also to Dr. Robert DeMars, who provided 721.221 cells, and Dr. Deric Wheeler who shared the empty vector. This research was supported by a Hyundai Hope on Wheels Grant; Midwest Athletes Against Childhood Cancer; NIH Grants R01 CA166105 and R35 CA197078; and a grant from The Stand Up To Cancer Foundation and St. Baldrick’s Foundation.

Compliance with ethical standards

Conflict of interest

The authors have no financial conflicts of interest.

Supplementary material

262_2016_1864_MOESM1_ESM.pdf (428 kb)
Supplementary material 1 (PDF 428 kb)


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Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Wei Wang
    • 1
  • Amy K. Erbe
    • 1
  • Kory A. Alderson
    • 1
  • Emily Phillips
    • 1
  • Mikayla Gallenberger
    • 1
  • Jacek Gan
    • 1
  • Dario Campana
    • 2
  • Jacquelyn A. Hank
    • 1
  • Paul M. Sondel
    • 1
    • 3
    Email author
  1. 1.Department of Human OncologyUniversity of WisconsinMadisonUSA
  2. 2.Department of PediatricsNational University of SingaporeSingaporeSingapore
  3. 3.Department of PediatricsUniversity of WisconsinMadisonUSA

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