scholarly journals Partial maturation and robust ex vivo expansion of Umbilical Cord Blood (UCB) T cells: A step towards adoptive immunotherapy after UCB transplants

2006 ◽  
Vol 12 (2) ◽  
pp. 101
Author(s):  
M.A. Mazur ◽  
Y.A. Lee ◽  
J. Kurtzburg ◽  
P. Szabolcs
Keyword(s):  
T Cells ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Adoptive Immunotherapy ◽  
Ex Vivo ◽  
Ex Vivo Expansion

Ex Vivo Expansion of Umbilical Cord Blood T Cells for Adoptive Immunotherapy.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 35-35
Author(s):  
Melissa A. Mazur ◽  
Young Ah Lee ◽  
Kurtzberg Joanne ◽  
Szabolcs Paul
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Adoptive Immunotherapy ◽  
Ex Vivo ◽  
Cell Activity ◽  
Ex Vivo Expansion ◽  
Hla Dr

Abstract Background: Viral infections cause significant morbidity & mortality in patients undergoing unrelated allogeneic BMT transplantation before immune reconstitution is completed. It poses a greater risk for recipients of unrelated umbilical cord blood (uUCB) transplants as there is no established antiviral immunity in naïve UCB lymphocytes available for adoptive transfer. UCB T cells also lack Th1/Tc1 cytokines, Granzymes & Perforin which are prerequisites to control viral pathogens. Another major limitation of uUCBT is the lack of donor cells available for post-transplant donor leukocyte infusions (DLI) to boost immunity or induce GVL. However, a fraction of the uCB graft could be available for T cell expansion. In this study we evaluated the feasibility of ex vivo expansion of UCB T cells. We postulated that following expansion naïve T cells may mature & acquire a phenotype compatible with effector function as assessed by expression of essential cytokines & de novo expression of members of the granzyme-perforin pathway. Methods: Thawed UBC research samples with a leukocyte content <5% of an average UCB graft are processed. T cells are enriched with “EasySep” (StemCell Tech) to deplete CD14, CD16, CD19, CD56, & glycophorin A + cells. 5–7.5*105 T cells/ml are incubated with “CD3/28 T cell Expander” artificial APC beads (Dynal) in X Vivo-15 (BioWhittacker) + 200u/ml IL2 & 10% human serum in gas permeable bags. The initial purity of the T cells is 77–92%. The starting absolute T cell numbers ranged from 0.75 to 2*106 cells. Media & cytokines are added every other day to maintain a concentration of <2*106 cells/ml. Results: At the end of 14 days UCB T Cells expanded 67 fold +/− 36, n=6. There are significant alterations in phenotype over the 2 weeks (Table 1) with up to 40% of T cells in cell cycle. Compared to the starting resting UCB T cells the majority of expanded cells have acquired the phenotype of activated (HLA-DR+, CD25+ T cells) memory cells, at the expense of naive/recent thymic immigrants (CD45RA+/CD62+). There is an inverted CD4/CD8 ratio due to the higher expansion rate of CD8 T cells (p=0.0035) while there is no difference in apoptosis (p=0.57). However, they all retain expression of CD28 (96% ±8%) along with CD27. Although some T cells have acquired the capacity to secrete granzymes A and B these are still almost a log below normal adult peripheral blood (PB) values & perforin has not been detected. Similarly, while post expansion significantly more T cells secrete cytokines upon PMA + ionomycin stimulation (Table 1) they are below levels of adult PB. Conclusions: From our preliminary results we can demonstrate effective expansion & partial maturation of UCB T Cells. For example, if one starts with 2*106 total T cells & expands them 67 fold this could provide for DLI ~5*106 T cells/kg for an average pediatric patient (25kg). We are further optimizing & characterizing this model for T cell activity & repertoire. In sum, ex vivo expansion with CD3/CD28 co-stimulation may provide clinically relevant numbers T cells available for adoptive immunotherapy that have also undergone partial maturation. Characterization of Expanded T Cells as % of all Lymphocytes Variable Median SD CD3+ 99.8 0.1 CD4+ 35 11 CD4+/CD8+ 2.3 2.8 CD45RA+/RO− 13 11 CD45RO+/RA− 55 22 CD25+ 42 21 CD45RA+/CD62+ 38 20 CD45RA+/CD27+/CD8+ 52 15 CD45RA−/CD27+/CD8+ 46 15 KI67/CD8+ 42 9 Ki67/CD4+ 32 7 HLA DR+ 40 13 Granzyme A/CD8+ 54 18 Granzyme B/CD8+ 2 2 Perforin/CD8+ 0 0

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

1999 ◽  
Vol 8 (2) ◽  
pp. 129-139 ◽  
Author(s):  
Danna Skea ◽  
Nan-Hua Chang ◽  
Robin Hedge ◽  
Barbara Dabek ◽  
Truman Wong ◽  
...  
Keyword(s):  
T Cells ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Cd8 T Cells ◽  
Umbilical Cord Blood ◽  
Ex Vivo ◽  
Ex Vivo Expansion ◽  
Human Umbilical Cord Blood ◽  
Human Umbilical Cord

scholarly journals Ex vivo expansion of immature and mature T cells derived from umbilical cord blood (UCB)

2004 ◽  
Vol 10 ◽  
pp. 16 ◽  
Author(s):  
S.L Staba ◽  
K.B Crapnell ◽  
J.G Hall ◽  
M Reese ◽  
J Kurtzberg
Keyword(s):  
T Cells ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Ex Vivo ◽  
Ex Vivo Expansion

Suppressive Characteristics of Umbilical Cord Blood–derived Regulatory T Cells After Ex Vivo Expansion on Autologous and Allogeneic T Effectors and Various Lymphoblastic Cells

2019 ◽  
Vol 42 (4) ◽  
pp. 110-118 ◽  
Author(s):  
Thitinee Vanichapol ◽  
Nutkridta Pongsakul ◽  
Supanart Srisala ◽  
Nopporn Apiwattanakul ◽  
Somchai Chutipongtanate ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Ex Vivo ◽  
Ex Vivo Expansion ◽  
Lymphoblastic Cells

Cord Blood-Derived T Cells Can Be Genetically Engineered to Be B-Lineage All-Specific: Towards Adoptive Immunotherapy after Umbilical Cord Blood Transplantation.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 48-48 ◽  
Author(s):  
Laurence J.N. Cooper ◽  
Lisa Marie Serrano ◽  
Zaid Al-Kadhimi ◽  
Simon Olivares ◽  
Sergio Gonzalez ◽  
...  
Keyword(s):  
T Cells ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Adoptive Immunotherapy ◽  
Ex Vivo ◽  
Non Invasive ◽  
Ganciclovir Treatment ◽  
B Lineage

Abstract Relapse of B-lineage (CD19+) acute lymphoblastic leukemia (ALL) remains a major impediment to the therapeutic success of allogeneic umbilical cord blood transplant (UCBT). The adoptive transfer of donor-derived tumor-specific T-cells is a conceptually attractive means to improve the graft-versus-leukemia-effect at the time of minimal residual disease to improve relapse-rates without exacerbating graft-versus-host-disease. However, adoptive immunotherapy after banked UCBT has been limited by the functional naïveté of neonatal T cells and difficulty obtaining T cells from the unrelated donor. These hurdles can now be overcome by genetically rendering cord blood-derived T cells to be specific for CD19 and expanding the T cells ex vivo from small numbers of cord blood cells, in compliance with current good manufacturing practices for phase I/II trials. To generate T cells that target CD19+ malignant cells, we have used non-viral gene transfer to introduce a DNA plasmid to express a CD19-specific chimeric immunoreceptor, designated CD19R, which binds to cell-surface CD19 via an scFv, independent of MHC, and triggers T-cell activation through CD3- ζ. To safeguard the safety of recipients of adoptive immunotherapy, the DNA plasmid co-expresses the bi-functional hygromycin phosphotransferase/HSV-1 thymidine kinase (HyTK) selection/suicide gene. To assess in vivo the fate of adoptively transferred T cells in mice; a novel tri-functional gene linking firefly luciferase (ffLuc) with HyTK (ffLucHyTK) was generated. The process ex vivo to expand cord blood-derived T cells, which is currently employed at COH in human trials, uses reiterative 14-day additions of OKT3, rhIL-2, cytocidal concentrations of hygromycin, and irradiated peripheral blood mononuclear cells (PBMC) and LCL as feeder cells. The expanded genetically manipulated cord-blood derived T cells express cell-surface markers of differentiated effector cells, similar to the phenotype of CD19-specific T cells derived from PBMC. In vitro the CD19R+HyTK+ cord blood-derived T cells are activated for cytolysis and cytokine production by CD19+ tumor cells. In vivo these genetically modified T cells can be used to eradicate established CD19+ tumors and undergo ganciclovir-mediated ablation, as demonstrated by non-invasive serial imaging of luciferase-mediated bioluminescence (see Figure). These data support a clinical trial to test the safety and feasibility of adoptive transfer of CD19-specific umbilical cord-blood derived T-cells for patients with high risk B-lineage ALL undergoing UCBT. Legend: Non-invasive in vivo biophotonic imaging demonstrates that (A) CD19+ tumor expressing ffLuc gene are eliminated by CD19R+HyTK+ cord-blood derived T cells, and (B) CD19R+ffLuc+HyTK+ cord-blood derived T cells are ablated by ganciclovir. Top row: prior to adoptive immunotherapy or ganciclovir treatment. Bottom row: after adoptive immunotherapy or ganciclovir treatment. Two representative mice are shown. Figure Figure

Augmentation of umbilical cord blood (UCB) transplantation with ex vivo–expanded UCB cells: results of a phase 1 trial using the AastromReplicell System

Blood ◽  
2003 ◽  
Vol 101 (12) ◽  
pp. 5061-5067 ◽  
Author(s):  
Jennifer Jaroscak ◽  
Kristin Goltry ◽  
Alan Smith ◽  
Barbara Waters-Pick ◽  
Paul L. Martin ◽  
...  
Keyword(s):  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Ex Vivo ◽  
Phase 1 ◽  
Horse Serum ◽  
Perfusion Culture ◽  
Ex Vivo Expansion ◽  
Hematopoietic Stem ◽  
Phase 1 Trial

AbstractAllogeneic stem cell transplantation with umbilical cord blood (UCB) cells is limited by the cell dose a single unit provides recipients. Ex vivo expansion is one strategy to increase the number of cells available for transplantation. Aastrom Biosciences developed an automated continuous perfusion culture device for expansion of hematopoietic stem cells (HSCs). Cells are expanded in media supplemented with fetal bovine serum, horse serum, PIXY321, flt-3 ligand, and erythropoietin. We performed a phase 1 trial augmenting conventional UCB transplants with ex vivo–expanded cells. The 28 patients were enrolled on the trial between October 8, 1997 and September 30, 1998. UCB cells were expanded in the device, then administered as a boost to the conventional graft on posttransplantation day 12. While expansion of total cells and colony-forming units (CFUs) occurred in all cases, the magnitude of expansion varied considerably. The median fold increase was 2.4 (range, 1.0-8.5) in nucleated cells, 82 (range, 4.6-266.4) in CFU granulocyte-macrophages, and 0.5 (range, 0.09-2.45) in CD34+ lineage negative (lin–) cells. CD3+ cells did not expand under these conditions. Clinical-scale ex vivo expansion of UCB is feasible, and the administration of ex vivo–expanded cells is well tolerated. Augmentation of UCB transplants with ex vivo–expanded cells did not alter the time to myeloid, erythroid, or platelet engraftment in 21 evaluable patients. Recipients of ex vivo–expanded cells continue to have durable engraftment with a median follow-up of 47 months (range, 41-51 months). A randomized phase 2 study will determine whether augmenting UCB transplants with ex vivo–expanded UCB cells is beneficial.

Ex vivo expanded umbilical cord blood T cells maintain naive phenotype and TCR diversity

Cytotherapy ◽  
2006 ◽  
Vol 8 (2) ◽  
pp. 149-157 ◽  
Author(s):  
S. Parmar ◽  
S.N. Robinson ◽  
K. Komanduri ◽  
L. St John ◽  
W. Decker ◽  
...  
Keyword(s):  
T Cells ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Ex Vivo ◽  
Tcr Diversity
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