Characterization of Cord Blood Natural Killer and Lymphokine Activated Killer Lymphocytes Following Ex Vivo Cellular Engineering

Abstract

Cord blood (CB) natural killer (NK) and lymphokine-activated killer (LAK) cytotoxic cells are poorly characterized but might be used to treat minimal residual and/or recurrent malignant disease. Currently, there is no mechanism to use CB for adoptive cancer cellular immunotherapy after CB transplantation (CBT). Recognizing this as a deficiency, we hypothesized that CB aliquots could be engineered ex vivo for potential donor lymphocyte infusion after CBT. Cryopreserved CB aliquots were thawed, depleted of monocytes, and cultured in serum-free medium alone or serum-free medium with anti-CD3 and interleukins 2, 7, and 12 combined with antibody/cytokines for 48 hours. Immunophenotyping, cytotoxicity, and proliferation were evaluated. A significant expansion of CD3 + was seen, in addition to increases in lymphocyte subsets of CD8 +, CD8 +/CD25 +, and CD3 +/45RO + versus medium alone. A significant enhancement of CD3 proliferation ( P < .001), NK cytotoxicity, NK subset expansion, LAK cytotoxicity, and T-helper 1 subset expansion was also demonstrated. Significant enrichment was seen in NK CD16 +/CD56 +bright, CD16 +/CD56 +dim, CD56 +bright and CD56 +dim/KIR3DL1 +, CD56 +bright and CD56 +dim/KIR2DL1 +, CD56 +bright and CD56 +dim/KIR2DL2 + and CD94 +/NKG2a + subsets. These increases in CB NK subsets were in part secondary to augmentation of cell survival. Further, survival of NOD-SCID mice xenografted with human K562 cells and treated with CB cells expanded with antibody/cytokines was significantly higher than that in animals that received no treatment (phosphate buffered saline) and those that were treated with CB ex vivo expanded in medium alone ( P < .005, respectively). These data suggest that cryopreserved CB cells could be ex vivo engineered for potential use as adoptive cancer cellular immunotherapy for donor lymphocyte infusion after CBT.