Expansion of mixed immune cells using CD3/CD161 co-stimulation for the treatment of cancer

Abstract

Adoptive cell transfer (ACT) is a type of personalized immunotherapy in which expanded immune cells are administered to patients with cancer. However, single-cell populations, such as killer T cells, dendritic cells, natural killer (NK) cells, and NKT (NKT) cells, have been generally used, and their effectiveness remains limited. Here, we established a novel culture method via CD3/CD161 co-stimulation and successfully expanded CD3⁺/CD4⁺ helper T cells, CD3⁺/CD8⁺ cytotoxic T cells (CTLs), CD3^-/CD56⁺ NK cells, CD3⁺/CD1d⁺ NKT cells, CD3⁺/CD56⁺ NKT cells, CD3⁺/TCRγδ⁺ T cells, and CD3^-/CD11c⁺/HLA-DR⁺ dendritic cells in peripheral blood mononuclear cells from healthy donors; their respective numbers were 155.5, 1132.5, 5.7, 117.0, 659.2, 325.6, and 6.8 times higher than those before expansion. These mixed immune cells showed strong cytotoxicity against cancer cell lines Capan-1 and SW480. Moreover, both CD3⁺/CD8⁺ CTLs and CD3⁺/CD56⁺ NKT cells killed tumor cells in cell contact-dependent and -independent manners via granzyme B and interferon-γ/TNF-α, respectively. Furthermore, the cytotoxicity of the mixed cells was significantly superior to that of CTLs or NKTs alone. A bet-hedging CTL-NKT circuitry is one potential mechanism underlying this cooperative cytotoxicity. Collectively, CD3/CD161 co-stimulation may be a promising culture method to expand multiple, distinct immune cell populations for the treatment of cancer.