Rapid and Massive Expansion of Cord Blood Derived Cytokine Induced Killer (CIK) Cells: An Innovative Proposal for the Treatment of Leukemia Relapse after Cord Blood Transplantation.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 723-723
Author(s):  
Martino Introna ◽  
Marta Franceschetti ◽  
Alice Ciocca ◽  
Gianmaria Borleri ◽  
Elena Conti ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cord Blood ◽  
Mononuclear Cells ◽  
Cord Blood Transplantation ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone ◽  
Cik Cells ◽  
Blood Transplantation

Abstract Cytokine induced killer cells (CIK) are CD3+/CD56+ T/NK cells with cytotoxic potential against leukemic and other tumor cells but not normal bone marrow in vitro and in vivo. They are expanded in vitro with rhIL-2 after stimulation of peripheral blood mononuclear cells with OKT3 and IFN-γ. We have shown in a recent phase I study that 107/kg allogeneic CIK cells can be safely given to patients relapsing after allogeneic bone marrow transplantation and show evidence of anti-leukemic activity in vivo with very little GVHD. Cord blood (CB) transplantation is progressively becoming an extensively used treatment for patients with malignant disorders. One major limitation of this procedure is the lack of donor derived cells to perform donor lymphocyte infusions in case of relapse. In order to be able therefore to extend the use of CIK cells to the CB transplantation setting, we have standardised a 21 days expansion protocol to produce CIK cells starting from very small amounts of nucleated cells isolated from cord blood. Using this protocol, 15x106 mononuclear cells (MNC) from CB containing a mean 0.3x106 CD3+/CD56+ yielded on average 805 x 106 MNC (50 fold expansion) containing 630 x106 CD3+/CD56+ cells (corresponding to a fold expansion of 1860 for CIK). In order to transfer the method to a clinical setting, we explored the possibility of expanding the residual cells recovered from the empty bags after CB transplantion. Three used CB bags were returned to the laboratory after transplantation and repeatedly washed. An average of 22 x106 nucleated cells could be recovered, yielding a mean 473 x106 CD3/CD56+ cells at the end of the culture period (1485 fold expansion of CIK cells). CIK cells generated from CB showed strong cytotoxic activity against a variety of tumor target cell lines including B and T lymphomas and myeloid leukemias (42–72% killing at a 30:1 E:T ratio). More importantly, they were cytotoxic against AML blasts isolated from 2 patients (41% lysis). During expansion CB derived CIK cells upregulated the NKG2D marker from a mean fluorescence intensity of 49 to 209. Furthermore they expressed perforin and granzyme molecules in >90% of cells. These observations open up the possibility of a future clinical application of this protocol, performed in GMP conditions. Patients relapsing following cord blood transplantation may be treated with CIK cells expanded from the same cord blood unit, where donors would not be anymore available for cell mediated immunotherapy.

scholarly journals Persistence of myeloid progenitor cells expressing BCR-ABL mRNA after allogeneic bone marrow transplantation for chronic myelogenous leukemia

Blood ◽  
1994 ◽  
Vol 84 (7) ◽  
pp. 2109-2114
Author(s):  
G Pichert ◽  
EP Alyea ◽  
RJ Soiffer ◽  
DC Roy ◽  
J Ritz
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Transplantation ◽  
Marrow Transplantation ◽  
Progenitor Cell ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone Marrow ◽  
Myeloid Differentiation ◽  
Allogeneic Bone

Previous studies have shown that tumor-specific bcr-abl mRNA can often be detected by polymerase chain reaction. (PCR) for months to years after allogeneic bone marrow transplantation (BMT) for chronic myelocytic leukemia (CML). Nevertheless, the presence of bcr-abl mRNA by itself does not invariably predict for clinical relapse post-BMT. This has led to the hypothesis that bcr-abl mRNA might be expressed in cells that have lost either proliferative or myeloid differentiation potential. To directly characterize the cells detected by PCR in patients with CML after allogeneic BMT, we first identified five individuals in whom PCR-positive cells could be detected at multiple times post-BMT. Bone marrow samples from these individuals were cultured in vitro and single erythroid, granulocytic, and macrophage colonies, each containing 50 to 100 cells, were examined for the presence of bcr-abl mRNA by PCR. PCR-positive myeloid colonies could be detected in four of five individuals in marrow samples obtained 5 to 56 months post-BMT. Overall, 7 of 135 progenitor cell colonies (5.2%) were found to be PCR-positive. The expression of bcr-abl mRNA appeared to be equally distributed among committed erythroid, macrophage, and granulocyte progenitors. These patients have now been followed-up for an additional 20 to 33 months from the time of progenitor cell PCR analysis but only one of these individuals has been found to have cytogenetic evidence of recurrent Ph+ cells. These results show that long-term persistence of PCR-detectable bcr-abl mRNA after allogeneic BMT can be caused by the persistence of CML-derived clonogenic myeloid precursors that have survived the BMT preparative regimen. These cells continue to have both proliferative and myeloid differentiation capacity in vitro. Nevertheless, these PCR-positive cells do not appear to either expand or differentiate in vivo for prolonged periods, suggesting the presence of mechanisms for suppression of residual clonogenic leukemia cells in vivo.

Cord blood transplantation provides better reconstitution of hematopoietic reservoir compared with bone marrow transplantation

Blood ◽  
2003 ◽  
Vol 102 (3) ◽  
pp. 1138-1141 ◽  
Author(s):  
Francesco Frassoni ◽  
Marina Podestà ◽  
Rita Maccario ◽  
Giovanna Giorgiani ◽  
Gabriele Rossi ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cord Blood ◽  
Mononuclear Cells ◽  
Cord Blood Transplantation ◽  
Hematopoietic Progenitor Cell ◽  
Transplant Recipients ◽  
Platelet Recovery ◽  
Blood Transplantation

Abstract Delayed hematopoietic recovery is the main factor precluding a wider use of cord blood (CB) transplants. We hypothesized that this delayed engraftment might not be related to an insufficient number of stem cells in the graft, but to an intrinsic difficulty of these cells to undergo differentiation. To test our hypothesis, 2 groups of children were compared; 12 received a CB transplant and 12 an adult bone marrow (BM) transplant. We studied neutrophil and platelet recovery and, at a median time of approximately 1 year after transplantation, the frequency of colony-forming cells (CFCs) and long-term culture initiating cells (LTC-ICs) in the BM of the 2 groups. Recipients of BM transplants received 1-log more cells and had significantly faster neutrophil and platelet recovery. Conversely, the frequency of committed and early progenitors was significantly higher in the BM of children given CB cells compared with BM transplant recipients (median count of CFC/2 × 104 BM mononuclear cells, 20 versus 11, P = .007; median count of LTC-IC/106 BM mononuclear cells, 8.2 versus 0.2 P = .001). CB, but not adult BM stem cells, can better restore the host hematopoietic progenitor cell reservoir; the delayed engraftment after CB transplantation may reflect the difficulty of CB progenitors to reprogram themselves toward differentiation.

Human Mesenchymal Stem Cells Attenuate Graft-Versus-Host Disease and Maintain Graft-Versus-Leukemia in Murine Allogeneic Bone Marrow Transplantation

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1907-1907 ◽  
Author(s):  
Jeffery J Auletta ◽  
Saada Eid ◽  
Matthew Keller ◽  
Leland Metheny ◽  
Rocio Guardia-Wolff ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Transplantation ◽  
Graft Versus Host Disease ◽  
Marrow Transplantation ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone ◽  
Graft Versus Host ◽  
Graft Versus Leukemia

Abstract Abstract 1907 Defining in vivo effects and biodistribution of human bone marrow-derived mesenchymal stem cell (hMSCs) following allogeneic bone marrow transplantation (alloBMT) could impact the clinical utility of MSC therapy for the prevention and treatment of graft-versus-host disease (GvHD). Using an established model of murine alloBMT, we defined hMSC effects on GvHD and graft-versus-leukemia (GvL) activity. We first studied whether hMSC could modulate in vitro murine T-cell (TC) alloreactivity in mixed leukocyte cultures (MLCs). Specifically, hMSCs added to MLCs significantly reduced TC proliferation in a concentration-dependent manner distinct from human fibroblasts. In contrast to MLC cultures alone, MLCs containing hMSCs had significant reduction in TNFα, IFNγ, and IL-10 levels and higher levels of PGE2 and TGFβ1. Modulation in the inflammatory milieu was associated with changes in TC phenotypes, including more naïve and less activated TC surface marker expression (CD62L+CD69−) and the induction of CD4+CD25+FoxP3+ T-regulatory cells. To determine whether hMSCs could modulate in vivo mTC alloreactivity, irradiated recipient B6D2F1 (H-2bxd) mice were transplanted with allogeneic C57BL/6 (H-2b) BM and purified splenic TCs (B6→B6D2F1) and then were tail-vein injected with hMSC infusions (1 million per injection) on days one and four post-transplant. Syngeneic transplant recipients (B6D2F1→B6D2F1) were used as controls. hMSC-treated alloBMT mice had significantly prolonged survival and improved clinical GvHD scores, reduced splenic TC expansion and TNFα and IFNγ-producing TCs, and lower circulating TNFα and IFNγ levels versus untreated alloBMT mice. Bioluminescence imaging showed redistribution of labeled hMSCs from the lungs to abdominal organs within 72 hours following infusion. Importantly, GvHD target tissues (small and large bowel and liver) harvested from hMSC-treated alloBMT mice had significantly lower GvHD pathology scores than untreated alloBMT mice. We next determined the effects of hMSCs on GvL activity using the murine mastocytoma cell line, P815 (H-2d). TCs co-cultured with hMSCs maintained potent in vitro cytotoxic T-lymphocyte (CTL) activity comparable to untreated control CTLs. After challenge with P815 tumor cells, hMSCs-treated alloBMT mice had less severe GvHD, eradication of tumor burden, and improved leukemia-free survival compared to alloBMT control mice. Lastly, indomethacin (IM) added to MLC-hMSC co-cultures significantly reversed attenuation in both murine TC alloreactivity and surface activation expression. In addition, IM administered to hMSC-treated alloBMT mice reversed hMSC-associated survival advantage, suggesting that PGE2 in part mediates hMSC immunomodulatory effects. Together, our results show that hMSC infusions effectively attenuate GvHD and maintain GvL potency in alloBMT mice and reveal potential biomarkers and mechanisms of action underlying hMSC effects. Disclosures: Solchaga: Bimemetic Therapeutics: Employment. Cooke:Amgen: Provides experimental drug and central pharmacy support for 2 trials for which I am Co-PI.

Differences in the Kinetics of Hematopoietic and Stem Cell (SC) Engraftment Following Murine Newborn/Cord Blood (NB) and Bone Marrow (BM) Transplantation.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4949-4949
Author(s):  
Tao Du ◽  
George F. Atweh ◽  
Yelena Galperin ◽  
Rona Singer Weinberg
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Cord Blood ◽  
Mononuclear Cells ◽  
Cord Blood Transplantation ◽  
Platelet Recovery ◽  
Platelet Counts ◽  
Blood Transplantation ◽  
Cell Counts ◽  
Kinetics Of

Abstract Cord blood is being used at an increasing frequency as a source of stem and progenitor cells in human hematopoietic transplantation. However, very little is known about the kinetics of engraftment of cord blood relative to bone marrow derived stem cells. We have used a murine model of newborn/cord blood transplantation to address this question. Our studies demonstrated that murine NB can provide long-term engraftment in primary, secondary, and tertiary transplant recipients. Although NB engrafts more slowly than BM, at one year, engraftment was similar in both types of recipients. We compared the rate of recovery of blood counts, hematopoietic progenitor counts and putative stem cell [SC] counts (i.e. Sca-1+, c-kit+, Lin- cells) in recipients of NB and BM transplantation. Interestingly, the SC ratio in mononuclear cells from donor NB/donor BM was 3.25 ± 0.8 (range = 2.45 – 4.75, n=15). Similarly, donor NB mononuclear cells contained approximately 30% of the number of megakaryocytic progenitors, 12% of the myeloid progenitors and 5% of the erythroid progenitors of donor BM mononuclear cells. The repopulation kinetics in recipients of donor BM and NB transplantation were analyzed at regular intervals, up to 8 months after transplantation. During the first two weeks, NB recipients had lower hemoglobin, WBC and platelet counts than BM recipients. However, by 1 month, the hemoglobin and WBC counts were at similar levels in NB and BM recipients. In contrast, NB transplantation recipients required 2 to 3 months to achieve platelet counts similar to those in BM recipients. These results are reminiscent of the well-known delayed platelet recovery following human cord blood transplantation. Progenitor cell counts in the bone marrow of recipients paralleled the hematological recovery described above. At 2 weeks post-transplantation, progenitor counts of all lineages in NB recipients were 25 to 35% of those in BM recipients. By 1 month, erythroid and myeloid progenitor numbers were similar in NB and BM recipients. In contrast, the appearance of megakaryocytic progenitors was delayed following NB transplantation and did not reach the same level as BM recipients until about 2 months after transplantation. During recovery, the number of SC in the bone marrow of both types of recipients increased gradually over time. At one month after transplantation, the number of SC in BM recipients was significantly greater than that in NB recipients, with a ratio of 4.2 ± 0.2. This SC ratio decreased gradually during the next several months. At 2, 4, and 8 months following transplantation, the ratios of SC in BM recipients/NB recipients were 3.5 ± 0.4, 2.6 ± 0.5, and 2.2 ± 0.3, respectively (n=5). This gradual decrease in the ratio of BM/NB SC suggests that NB SC increase more rapidly than BM SC. In conclusion, these data demonstrate that the kinetics of hematopoietic and stem cell recovery following NB and BM transplantation are significantly different. Although hematopoietic recovery after NB transplantation is slow at first, final engraftment is similar following NB and BM transplantation. Furthermore, the number of SC in NB recipients increases at a faster rate than the number of SC in BM recipients. These differences in SC recovery may be a reflection of differences either in the homing capacity or in the functional maturity of NB relative to BM SC. Further investigation is required to distinguish between these two possibilities.

scholarly journals Persistence of myeloid progenitor cells expressing BCR-ABL mRNA after allogeneic bone marrow transplantation for chronic myelogenous leukemia

Blood ◽  
1994 ◽  
Vol 84 (7) ◽  
pp. 2109-2114 ◽  
Author(s):  
G Pichert ◽  
EP Alyea ◽  
RJ Soiffer ◽  
DC Roy ◽  
J Ritz
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Transplantation ◽  
Marrow Transplantation ◽  
Progenitor Cell ◽  
Allogeneic Bone Marrow Transplantation ◽  
Allogeneic Bone Marrow ◽  
Myeloid Differentiation ◽  
Allogeneic Bone

Abstract Previous studies have shown that tumor-specific bcr-abl mRNA can often be detected by polymerase chain reaction. (PCR) for months to years after allogeneic bone marrow transplantation (BMT) for chronic myelocytic leukemia (CML). Nevertheless, the presence of bcr-abl mRNA by itself does not invariably predict for clinical relapse post-BMT. This has led to the hypothesis that bcr-abl mRNA might be expressed in cells that have lost either proliferative or myeloid differentiation potential. To directly characterize the cells detected by PCR in patients with CML after allogeneic BMT, we first identified five individuals in whom PCR-positive cells could be detected at multiple times post-BMT. Bone marrow samples from these individuals were cultured in vitro and single erythroid, granulocytic, and macrophage colonies, each containing 50 to 100 cells, were examined for the presence of bcr-abl mRNA by PCR. PCR-positive myeloid colonies could be detected in four of five individuals in marrow samples obtained 5 to 56 months post-BMT. Overall, 7 of 135 progenitor cell colonies (5.2%) were found to be PCR-positive. The expression of bcr-abl mRNA appeared to be equally distributed among committed erythroid, macrophage, and granulocyte progenitors. These patients have now been followed-up for an additional 20 to 33 months from the time of progenitor cell PCR analysis but only one of these individuals has been found to have cytogenetic evidence of recurrent Ph+ cells. These results show that long-term persistence of PCR-detectable bcr-abl mRNA after allogeneic BMT can be caused by the persistence of CML-derived clonogenic myeloid precursors that have survived the BMT preparative regimen. These cells continue to have both proliferative and myeloid differentiation capacity in vitro. Nevertheless, these PCR-positive cells do not appear to either expand or differentiate in vivo for prolonged periods, suggesting the presence of mechanisms for suppression of residual clonogenic leukemia cells in vivo.

scholarly journals An in vivo model of double-unit cord blood transplantation that correlates with clinical engraftment

Blood ◽  
2010 ◽  
Vol 116 (19) ◽  
pp. 3999-4006 ◽  
Author(s):  
Lamis K. Eldjerou ◽  
Sonali Chaudhury ◽  
Ada Baisre-de Leon ◽  
Mai He ◽  
Maria E. Arcila ◽  
...  
Keyword(s):  
Cord Blood ◽  
Single Unit ◽  
Mononuclear Cells ◽  
Cord Blood Transplantation ◽  
In Vivo Model ◽  
Cell Potential ◽  
Blood Transplantation ◽  
Cd34 Cells ◽  
Colony Forming Cell

Abstract Double-unit cord blood transplantation (DCBT) appears to enhance engraftment despite sustained hematopoiesis usually being derived from a single unit. To investigate DCBT biology, in vitro and murine models were established using cells from 39 patient grafts. Mononuclear cells (MNCs) and CD34+ cells from each unit alone and in DCB combination were assessed for colony-forming cell and cobblestone area-forming cell potential, and multilineage engraftment in NOD/SCID/IL2R-γnull mice. In DCB assays, the contribution of each unit was measured by quantitative short tandem repeat region analysis. There was no correlation between colony-forming cell (n = 10) or cobblestone area-forming cell (n = 9) numbers and clinical engraftment, and both units contributed to DCB cocultures. In MNC transplantations in NOD/SCID/IL2R-γnull mice, each unit engrafted alone, but MNC DCBT demonstrated single-unit dominance that correlated with clinical engraftment in 18 of 21 cases (86%, P < .001). In contrast, unit dominance and clinical correlation were lost with CD34+ DCBT (n = 11). However, add-back of CD34− to CD34+ cells (n = 20) restored single-unit dominance with the dominant unit correlating not with clinical engraftment but also with the origin of the CD34− cells in all experiments. Thus, unit dominance is an in vivo phenomenon probably associated with a graft-versus-graft immune interaction mediated by CD34− cells.

scholarly journals Allogeneic cell therapy with donor peripheral blood cells and recombinant human interleukin-2 to treat leukemia relapse after allogeneic bone marrow transplantation

Blood ◽  
1996 ◽  
Vol 87 (6) ◽  
pp. 2195-2204 ◽  
Author(s):  
S Slavin ◽  
E Naparstek ◽  
A Nagler ◽  
A Ackerstein ◽  
S Samuel ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Therapy ◽  
Allogeneic Bone Marrow Transplantation ◽  
Interleukin 2 ◽  
Hematologic Malignancies ◽  
Remission Induction ◽  
Allogeneic Bone ◽  
Allogeneic Cell Therapy

Allogeneic bone marrow transplantation (BMT) is the only effective treatment for hematologic malignancies resistant to conventional chemotherapy. Until recently, no cure existed for patients who relapsed post-BMT. We present our long-term observations on remission induction, after relapse post-BMT, by allogeneic cell therapy (allo-CT) and the feasibility of remission induction in allo-CT-resistant patients by activation of antileukemia effector cells with recombinant human interleukin-2 (rhIL-2) in vitro and in vivo. The longest observation of successful allo-CT (event-free survival, greater than 8 years) was made in a patient with resistant pre-B lymphoblastic leukemia who received infusions with graded increments of donor (female) peripheral blood lymphocytes (PBL) as soon as bulky hematologic and extramedullary relapse was noticed early post-BMT. The patient is currently without evidence of residual host (male) cells as determined by polymerase chain reaction (PCR). Of 17 patients with acute and chronic leukemia in relapse after BMT, 10 were reinduced into complete remission. Four patients with cytogenetic relapse responded to allo-CT alone, while five of six patients with overt hematologic relapse responded only after additional activation of donor with rhIL-2. Allo-CT can, therefore, successfully reverse chemoradiotherapy-resistant relapse of both acute and chronic leukemia. Moreover, in patients resistant to donor lymphocyte infusion, remission can be accomplished by additionally activating donor PBL in vitro and/or in vivo with rhIL-2. Based on our observations, after BMT, allo-CT should be considered the treatment of choice for patients with hematologic malignancies resistant to conventional anticancer modalities. Allogeneic activated cell therapy (allo ACT) should be considered for patients with tumor cells resistant to allo-CT. Although allo-CT, followed if indicated by allo-ACT, can be effective for patients with overt hematologic relapse, reversal of persistent minimal residual disease or documented molecular/cytogenetic relapse early after BMT may also be considered as a possible indication for allo-CT.

scholarly journals Primitive Human Hematopoietic Cells Are Enriched in Cord Blood Compared With Adult Bone Marrow or Mobilized Peripheral Blood as Measured by the Quantitative In Vivo SCID-Repopulating Cell Assay

Blood ◽  
1997 ◽  
Vol 89 (11) ◽  
pp. 3919-3924 ◽  
Author(s):  
Jean C.Y. Wang ◽  
Monica Doedens ◽  
John E. Dick
Keyword(s):  
Bone Marrow ◽  
Cord Blood ◽  
Peripheral Blood ◽  
Hematopoietic Cells ◽  
Allogeneic Transplantation ◽  
Functional Assay ◽  
Hematopoietic Stem ◽  
Mobilized Peripheral Blood

Abstract We have previously reported the development of in vivo functional assays for primitive human hematopoietic cells based on their ability to repopulate the bone marrow (BM) of severe combined immunodeficient (SCID) and nonobese diabetic/SCID (NOD/SCID) mice following intravenous transplantation. Accumulated data from gene marking and cell purification experiments indicate that the engrafting cells (defined as SCID-repopulating cells or SRC) are biologically distinct from and more primitive than most cells that can be assayed in vitro. Here we demonstrate through limiting dilution analysis that the NOD/SCID xenotransplant model provides a quantitative assay for SRC. Using this assay, the frequency of SRC in cord blood (CB) was found to be 1 in 9.3 × 105 cells. This was significantly higher than the frequency of 1 SRC in 3.0 × 106 adult BM cells or 1 in 6.0 × 106 mobilized peripheral blood (PB) cells from normal donors. Mice transplanted with limiting numbers of SRC were engrafted with both lymphoid and multilineage myeloid human cells. This functional assay is currently the only available method for quantitative analysis of human hematopoietic cells with repopulating capacity. Both CB and mobilized PB are increasingly being used as alternative sources of hematopoietic stem cells in allogeneic transplantation. Thus, the findings reported here will have important clinical as well as biologic implications.

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