Expansion of human Valpha24+ NKT cells by repeated stimulation with KRN7000

J Immunol Methods. 2004 Feb 15;285(2):197-214. doi: 10.1016/j.jim.2003.12.003.


Changes in Valpha24+Vbeta11+ NKT cell number and function are associated with human autoimmune diseases and cancer. Restoration of this corresponding NKT cell population in mice or in vivo activation with alpha-galactosylceramide (KRN7000) can prevent or reduce tumor growth and autoimmunity. Although the therapeutic value of these natural killer T (NKT) cells in man remains to be determined, large numbers of functional antigen-specific NKT cells can be expanded in vitro. We show that Valpha24+Vbeta11+ human NKT cells are expanded by repeated stimulation with KRN7000, unfractionated donor peripheral blood mononuclear cells (PBMC), and recombinant human interleukin-2 (rhIL-2). NKT cells were expanded continuously for more than 2 months with a potential yield of >10(12) cells. The expanded NKT cells retained their CD4+ or CD4- phenotype after restimulation and were functional as shown by cytokine secretion, killing of antigen-pulsed target cells, and activation of NK cell cytotoxicity. This expansion method may be useful for proof-of-concept studies involving adoptive transfer of ex vivo-expanded NKT cells as a new therapeutic option for cancer and autoimmune diseases.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adjuvants, Immunologic / pharmacology*
  • CD4 Antigens / metabolism
  • CD8 Antigens / metabolism
  • Cells, Cultured
  • Cytotoxicity, Immunologic
  • Enzyme-Linked Immunosorbent Assay
  • Flow Cytometry
  • Galactosylceramides / pharmacology*
  • Growth Substances / analysis
  • Growth Substances / pharmacology
  • Humans
  • Interleukin-2 / analysis
  • Interleukin-2 / pharmacology
  • Jurkat Cells
  • Killer Cells, Natural / drug effects*
  • Killer Cells, Natural / immunology*
  • Lymphocyte Activation / immunology*
  • Receptors, Antigen, T-Cell / immunology
  • Recombinant Proteins / pharmacology


  • Adjuvants, Immunologic
  • CD4 Antigens
  • CD8 Antigens
  • Galactosylceramides
  • Growth Substances
  • Interleukin-2
  • Receptors, Antigen, T-Cell
  • Recombinant Proteins
  • KRN 7000