Generation of high-affinity CMV-specific T cells for adoptive immunotherapy using IL-2, IL-15, and IL-21

Clin Immunol. 2020 Aug;217:108456. doi: 10.1016/j.clim.2020.108456. Epub 2020 May 5.

Abstract

Cytomegalovirus (CMV) infection remains a life-threatening condition in individuals with a suppressed immune system. CMV may also represent a clinically relevant target for immune responses in CMV-positive malignancies. We established a protocol to expand CMV-specific T cells (CMV-T) using peripheral blood mononuclear cells (PBMCs). PBMCs from 16 HLA-A*0201 donors were cultured with a cytokine cocktail comprising IL-2/IL-15/IL-21 along with overlapping peptides from CMV-pp65. Ten days later, T cells were stimulated with anti-CD3 (OKT3) and irradiated autologous PBMCs. CMV-T were detected by HLA-A*0201 CMV-pp65NLVPMVATV wild type and q226a mutant tetramers (for high-affinity T cells), intracellular cytokine staining, a CD107a mobilization assays as well as IFN-γ and TNF-α production in cell culture supernatants. We reliably obtained 50.25 ± 27.27% of CD8+ and 22.08 ± 21.83% of CD4+ T cells post-CMV-pp65 stimulation of PBMCs with a Th1-polarized phenotype and decreased Th2/Th17 responses. Most CD3 + CD8 + tetramer+ T cells were effector-memory cells, particularly among high-affinity CMV-T (q226a CMV-tetramer+). High-affinity CMV-T cells, compared to WT-tetramer+ cells, expressed higher IL-21R and lower FasL post-stimulation with CMV-pp65. The IL-2/IL-15/IL-21 cocktail also promoted CCR6 and CXCR3 expression necessary for T-cell migration into tissues. We have optimized methods for generating high-affinity CMV-specific T cells that can be used for adoptive cellular therapy in clinical practice.

Keywords: Adoptive immunotherapy; CMV-specific T cells; Cytokines; IL-15; IL-2; IL-21.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adolescent
  • Adult
  • Aged
  • CD4-Positive T-Lymphocytes / immunology*
  • CD4-Positive T-Lymphocytes / transplantation*
  • Cells, Cultured
  • Cytomegalovirus / immunology*
  • Cytomegalovirus Infections / therapy*
  • HLA-A2 Antigen / immunology
  • Humans
  • Immunotherapy, Adoptive / methods*
  • Interferon-gamma / analysis
  • Interleukin-15 / pharmacology
  • Interleukin-2 / pharmacology
  • Interleukins / pharmacology
  • Middle Aged
  • Tumor Necrosis Factor-alpha / analysis
  • Young Adult

Substances

  • HLA-A*02:01 antigen
  • HLA-A2 Antigen
  • IFNG protein, human
  • IL15 protein, human
  • IL2 protein, human
  • Interleukin-15
  • Interleukin-2
  • Interleukins
  • Tumor Necrosis Factor-alpha
  • Interferon-gamma
  • interleukin-21