A clinically adaptable method to enhance the cytotoxicity of natural killer cells against B-cell malignancies

Cytotherapy. 2012 Aug;14(7):830-40. doi: 10.3109/14653249.2012.671519. Epub 2012 Mar 29.

Abstract

Background aims: Retroviral transduction of anti-CD19 chimeric antigen receptors significantly enhances the cytotoxicity of natural killer (NK) cells against B-cell malignancies. We aimed to validate a more practical, affordable and safe method for this purpose.

Methods: We tested the expression of a receptor containing CD3ζ and 4-1BB signaling molecules (anti-CD19-BB-ζ) in human NK cells after electroporation with the corresponding mRNA using a clinical-grade electroporator. The cytotoxic capacity of the transfected NK cells was tested in vitro and in a mouse model of leukemia.

Results: Median anti-CD19-BB-ζ expression 24 h after electroporation was 40.3% in freshly purified (n =18) and 61.3% in expanded (n = 31) NK cells; median cell viability was 90%. NK cells expressing anti-CD19-BB-ζ secreted interferon (IFN)-γ in response to CD19-positive target cells and had increased cytotoxicity. Receptor expression was detectable 6 h after electroporation, reaching maximum levels at 24-48 h; specific anti-CD19 cytotoxicity was observed at 96 h. Levels of expression and cytotoxicities were comparable with those achieved by retroviral transduction. A large-scale protocol was developed and applied to expanded NK cells (median NK cell number 2.5 × 10(8), n = 12). Median receptor expression after 24 h was 82.0%; NK cells transfected under these conditions exerted considerable cytotoxicity in xenograft models of B-cell leukemia.

Conclusions: The method described here represents a practical way to augment the cytotoxicity of NK cells against B-cell malignancies. It has the potential to be extended to other targets beyond CD19 and should facilitate the clinical use of redirected NK cells for cancer therapy.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antigens, CD19* / genetics
  • Antigens, CD19* / immunology
  • Antigens, CD19* / therapeutic use
  • CD3 Complex / genetics
  • CD3 Complex / therapeutic use
  • Cell Line, Tumor
  • Cell- and Tissue-Based Therapy*
  • Cytotoxicity, Immunologic / genetics
  • Gene Expression Regulation, Leukemic
  • Humans
  • Killer Cells, Natural / cytology*
  • Killer Cells, Natural / metabolism
  • Lymphocyte Activation
  • Lymphoma, Non-Hodgkin* / therapy
  • Mice
  • Precursor Cell Lymphoblastic Leukemia-Lymphoma* / therapy
  • RNA, Messenger / genetics
  • RNA, Messenger / therapeutic use
  • Tumor Necrosis Factor Receptor Superfamily, Member 9 / genetics
  • Tumor Necrosis Factor Receptor Superfamily, Member 9 / therapeutic use

Substances

  • Antigens, CD19
  • CD3 Complex
  • CD3 antigen, zeta chain
  • RNA, Messenger
  • TNFRSF9 protein, human
  • Tumor Necrosis Factor Receptor Superfamily, Member 9