Functional evaluation of ex vivo expanded cord blood lymphocytes

Abstract

To examine the possibility of adoptive cellular immunotherapy such as donor lymphocyte infusion using ex vivo expanded cord blood (CBL) lymphocytes, the potential expansion ability of CBL lymphocytes and the function of expanded CBL lymphocytes were evaluated.Mononuclear cell fractions derived from CBL or peripheral blood (PBL) were placed in anti-CD3 monoclonal antibody-coated flasks and cultured in the presence of recombinant human interleukin-2 for 4 days. Cells then were transferred to noncoated flasks and cultured for another 2 weeks. On day 14, polyclonality, cell surface markers, killer activity, and intracellular cytokine profiles were evaluated.Cells were polyclonally expanded. The differences in cumulative fold expansion on day 14 between CBL [1174 +/- 637 (292-1939), n = 6] and PBL [1247 +/- 568 (517-2328), n = 9] were not significant (p = 0.95). Phenotypic patterns of both expanded CBL and PBL were similar. CD4/CD8 ratio of expanded CBL appeared to remain greater than 1 on day 8. In contrast, that of expanded PBL became less than 1. In both cases, approximately 20% of cells had the CD3(+)CD8(+)CD56(+) phenotype. At an effector to target ratio (E/T) of 40:1, the natural killer activity of expanded CBL (64.5% +/- 10.8%, n = 9) was significantly higher than that of expanded PBL (48.3% +/- 16.8%, n = 9) (p < 0.01, Mann-Whitney U-test). However, there was no significant difference in lymphokine-activated killer activity between expanded CBL (45.3% +/- 25.2%, n = 7) and expanded PBL (67.2% +/- 12.3%, n = 7). Interferon-gamma-producing cells were dominant in both cases.It was feasible to achieve approximately 1000-fold expansion of CBL, and the phenotype and function of expanded CBLs were essentially equivalent to those of expanded PBL. This suggested that ex vivo expanded CBL may be applied to adoptive cellular immunotherapy such as donor lymphocyte infusion.