The immunobiology of cord blood transplantation

Abstract

Despite significant recent advances in the applicability and outcome following unrelated cord blood transplantation (UCBT), infections remain a major cause of mortality associated with poor immune recovery in the first 6 months after UCBT. Enhanced immune reconstitution not only could improve survival by reduced transplant related mortality, but may also favorably impact on relapse incidence by improved graft-versus-leukemia effects. This review will summarize our current understanding of the biology of immune recovery post-UCBT with an emphasis on adaptive T cell dependent immunity. New efforts to boost immunity will be also highlighted including our own laboratory, where ex vivo T cell expansion is pursued towards adoptive immunotherapy.

Keywords: Adoptive immunotherapy; Cord blood stem cell transplantation; Graft vs leukemia effect; Immune reconstitution; Opportunistic infection.

Fig. 1

Kaplan-Meier curve of survival (months) after UCBT in 330 consecutive patients. Death related to opportunistic infections (OI ) is the major cause of failure, most occurring by 6 months. Reproduced with permission from Cytotherapy © 2007, Informa Healthcare Journals.

Fig. 2

(A) Time to death from all causes in the "Day 50" cohort by opportunistic infection status. (B) Time to death from OI by presence or absence of severe GVHD. Reproduced with permission from Cytotherapy © 2007, Informa Healthcare Journals.

Fig. 3

(A) CD3⁺ T cell expansion is superior in the presence of IL-7. Frozen/thawed cord blood T cells were enriched by negative selection then split equally into two under identical culture conditions except for the presence of IL-7 as indicated. Cells were cultured for 12-14 days with ClinExVivo™ Dynabeads®, while medium and cytokines were replenished ×3/week. A 50 µL aliquot was removed from the bags at indicated time points and absolute T cells number was enumerated in Trucount® tubes. (B) Irrespective of IL-7 in the culture medium, expansion leads to dilution and near complete loss of sjTREC in day 14 progeny. The signal joint TCR excision circles (sjTREC) were measured before and after expansion (N=4). For each sample total nucleated cell count and absolute T cells content was enumerated by Trucount FACS method. TREC content was expressed after adjustment for 10⁵ T cells/sample. Reproduced with permission from Cancer Research © 2010.

Fig. 4

(A) Kinetic analysis of surface CD25 and CD127 expression. Simultaneous monitoring of IL-2Rα (CD25) and IL-7Rα (CD127) was performed after FACS surface staining and acquisition as described [30, 64] on serial aliquots obtained before (Day 0) and during expansion on the indicated days. (B) Cell death after 24 h of rest in cytokine free medium was assayed and scored by positive staining for Annexin and 7-AAD in parallel after freeze and thaw of expanded day 14 T cells, representative of 4 experiments. Reproduced with permission from Cancer Research © 2010.

Fig. 5

Flow cytometry profile of the expanded T cell progeny±IL-7. Surface and intracellular (ic) FACS characterization was performed as shown previously [30, 64, 67]. The relative size of T cell subsets in each quadrant is expressed as the percentage of total viable T cells, see Table 1 for P-values. (A) CD45-PERCP/SSC defines an unambiguous region of viable cells. All other CD45 dim cells (recently apoptotic) stain also dim for CD3, data not shown. (B) icKI-67 staining (upper quads) identifies more proliferating T cells when expanded with IL-7 than without. (C) When expanded without IL-7 more T cells undergo apoptosis and stain with ic ActiveCasp-3⁺ even though gated from the viable region of Fig. 2A. (D) More T cells display the phenotype of 'naïve/CD45RA⁺/RO^- T cells when expanded with IL-7. Representative of 10 experiments. Reproduced with permission from Cancer Research © 2010.

Fig. 6

Absent cytotoxicity of the expanded CB T cells against allogeneic targets irrespective of±IL-7. Effector T cells were obtained from PBL of healthy volunteers as positive controls and compared with CD3/28 co-stimulated CB T cells±IL-7. First, effectors were primed/sensitized against a highly immunogeneic (HLA-DR+, CD40+, CD80+, CD86+) IM9 cell line for 7-9 days at 1:1 to 1:3 responder:stimulator ratio, then re-exposed to fresh BATDA®-loaded IM9 targets at the indicated E:T ratios for 2 & 3 h. Europium release was measured by the Delfia® EuTDA cytotoxicity assay [67] and the calculated percent specific cytotoxicity is presented on the Y-axis. Representative of 7 experiments.

Fig. 7

Leukemia-specific CTL can be in vitro primed starting with the CD3/28-expanded CB T cells. T cells were first CD3/28-expanded in the presence of IL2+ IL-7 over 14 days as described and thereafter were primed/sensitized against 2 killed leukemia cell lines in parallel cultures for 7-9 days at 10:1 responder:stimulator ratio in the presence of IL-12, IL-7, and IL-15. (A) CTL primed in vitro with Mitomycin C-treated IM9 cells. (B) CTL primed in vitro with IFN-treated and Mitomycin C-treated U937 cells. Each CTL culture was re-stimulated 2 more times (1^st IL7+ IL-15, thereafter 2^nd in IL15 alone) for a total of 3 weeks with their respective killed leukemia cells. Cytotoxicity of washed effectors after 3 weeks in CTL culture was tested against fresh, unmodified, BATDA®-loaded IM9 (▴), U937 (▪) cells, and recipient PHA blasts (♦) at the indicated E:T ratios for 3 h, as indicated. Europium release was measured by the Delfia® EuTDA cytotoxicity assay, and the calculated percent specific cytotoxicity is presented on the Y-axis. Reproduced with permission from Cancer Research © 2010.