Large Ex Vivo Expansion of Human Umbilical Cord Blood CD4+ and CD8+ T Cells

Abstract

The use of human umbilical cord (UC) blood as a source of transplantable hematopoietic stem cells and progenitor cells may present some advantages over the use of BM. For example, it has been suggested that the degree of HLA matching may be less stringent, and the risk of GvHD may be lower. We have been studying the ex vivo expansion of UC blood T lymphocytes with a view to their use in the adoptive immunotherapy of cancer, autoimmunity, and infectious disease. We have developed a new method involving the use of a conditioned medium (XLCM) that consistently results in levels of UC blood T cell expansion not hitherto possible. Primary cultures of unfractionated low-density MNC (LDMNC) derived from UC blood treated with 5% XLCM routinely show expansions greater than 10,000-fold within 4 weeks. By contrast, similar FBS-free cultures treated with IL-2 expand less than 10-fold and not after 1 week, and cultures treated with IL-2 and concanavalin A (ConA) expand to a maximum of only 300-500-fold over 2 weeks and fail to continue to proliferate thereafter. The MAb, OKT3, which, when combined with IL-2 and FBS, is known to stimulate proliferation of adult peripheral blood lymphocytes, permitted only a 17-fold expansion of UC blood lymphocytes under the same conditions. Thus, XLCM, which can also stimulate adult peripheral blood lymphocyte expansion to levels exceeding 100,000-fold in 3-4 weeks, is uniquely able to stimulate proliferation of UC blood lymphocytes to high levels. From initiation of the UC blood or adult peripheral blood LDMNC/XLCM cultures up to approximately 2 weeks, the cultures are dominated by CD4+ T lymphocytes. By 4 weeks, >80% of the cultured cells bear the CD8+ phenotype, whereas UC blood T lymphocytes cultured in the presence of IL-2 are predominantly CD8+. Thus, XLCM not only allows high levels of expansion of UC blood T lymphocytes not heretofore possible but also permits the selective expansion of different T lymphocyte subsets from a single source.

Amplification of T cells from human cord blood in serum-deprived culture stimulated with stem cell factor, interleukin-7 and interleukin-2

“…8,9,11 The increases in T-cell number were associated with a considerable increase in the complexity of the TCR repertoire expressed by the cultured T cells. Although the levels of amplification achieved in this paper were lower than those induced by the mitogens concanavalin A and mezerein, 10 it is debatable whether mitogens will be used for clinical purposes in the near future. Furthermore, in contrast to all the conditions for T-cell amplification established up to now, stimulation of CB-LDC with lymphoid-specific growth factor combinations sustained not only amplification of T cells but also that of progenitor cells.…”

“…This problem, however, could be solved by using as DLI ex vivo expanded T (CD3 + ) cells obtained from a portion of the CB unit 8 and pre-clinical studies are under way to analyze the GVL effect of ex vivo-amplified neonatal T cells in leukemic and cancer patients. 8 Most of the ex vivo manipulation and amplification of T cells realized up to now [9][10][11] are sustained by a combination of IL-2 with either anti-CD3 or anti-CD28, 8,9,11 and the culture conditions include serum, either fetal bovine or human. 9,12 Furthermore, none of these studies has investigated whether the increases in T-cell number are associated with increases in T-cell clonality.…”

Differentiation of naive cord-blood T cells into CD19-specific cytolytic effectors for posttransplantation adoptive immunotherapy

“…While UCB-derived T cells can be propagated ex vivo, [43][44][45][46][47][48][49] the functional immaturity of T cells in circulating umbilical cord blood typically prevents ex vivo identification of endogenous leukemia/ lymphoma-specific T cells. Therefore, we developed a genetic engineering strategy to introduce a chimeric immunoreceptor in order to generate UCB-derived T cells with desired specificity.…”

Ex vivo expansion of CD34+ and T and NK cells from umbilical cord blood for leukemic BALB/C nude mouse transplantation

“…For an adoptive immunotherapy of cancer, Skea et al [14] developed a new method involving the use of a conditioned medium (XLCM) that consistently results in levels of UC blood T cell expansion. It permitted a selective expansion of different T lymphocyte subsets from a single source.…”