Allogeneic Umbilical Cord Blood Cells Research Articles

Allogeneic umbilical cord blood cell therapy for children with autism: safety and efficacy of the method

To evaluate effectiveness and safety of umbilical cord blood cells (UCBC) in children with ASD. The study comprised 13 boys and 2 girls, mean age 7.0±0.5 years (test group), and 9 boys and 1 girl, mean age 6.0±1.3 years (control group) diagnosed with autism or autistic syndrome. UCBC were infused intravenously in a single dose of 250±20 million cells, four times at 14±3-day intervals. Dynamics of cognitive functions were assessed with WISC subscales, questionnaire Skvortsov Developmental Profile Survey and brainstem auditory evoked potentials (BAEPs) conducted before therapy and six months after the first injection. The dynamic of autistic symptoms was explored with CASD and ATEC questionnaires, fulfilled at the start of the study, 3 and 6 months later (test group) or at the start of the study, and 6 months later (control group). UCBC was well tolerated and caused no appreciable adverse effects. Observation revealed improvement in cognitive functioning and alleviation of autistic symptoms in patients of the test group six months after the first UCBC injection. Positive dynamics were more noticeable in the test group than in the control group receiving standard therapy. The use of UCBC is safe and might be effective in the complex therapy of autism.

Potentiation of cord blood cell therapy with erythropoietin for children with CP: a 2\u2009\xd7\u20092 factorial randomized placebo-controlled trial

BackgroundConcomitant administration of allogeneic umbilical cord blood (UCB) infusion and erythropoietin (EPO) showed therapeutic efficacy in children with cerebral palsy (CP). However, no clinical studies have investigated the effects of UCB and EPO combination therapy using a 2 × 2 four-arm factorial blinded design with four arms. This randomized placebo-controlled trial aimed to identify the synergistic and individual efficacies of UCB cell and EPO for the treatment of CP.MethodsChildren diagnosed with CP were randomly segregated into four groups: (A) UCB+EPO, (B) UCB+placebo EPO, (C) placebo UCB+EPO, and (D) placebo UCB+placebo EPO. Based on the UCB unit selection criteria of matching for ≥ 4/6 of human leukocyte antigen (HLA)-A, -B, and DRB1 and total nucleated cell (TNC) number of ≥ 3 × 107/kg, allogeneic UCB was intravenously infused and 500 IU/kg human recombinant EPO was administered six times. Functional measurements, brain imaging studies, and electroencephalography were performed from baseline until 12 months post-treatment. Furthermore, adverse events were closely monitored.ResultsEighty-eight of 92 children enrolled (3.05 ± 1.22 years) completed the study. Change in gross motor performance measure (GMPM) was greater in group A than in group D at 1 month (△2.30 vs. △0.71, P = 0.025) and 12 months (△6.85 vs. △2.34, P = 0.018) post-treatment. GMPM change ratios were calculated to adjust motor function at the baseline. Group A showed a larger improvement in the GMPM change ratio at 1 month and 12 months post-treatment than group D. At 12 months post-treatment, the GMPM change ratios were in the order of groups A, B, C, and D. These results indicate synergistic effect of UCB and EPO combination better than each single therapy. In diffusion tensor imaging, the change ratio of fractional anisotropy at spinothalamic radiation was higher in group A than group D in subgroup of age ≥ 3 years. Additionally, higher TNC and more HLA-matched UCB units led to better gross motor outcomes in group A. Adverse events remained unchanged upon UCB or EPO administration.ConclusionsThese results indicate that the efficacy of allogeneic UCB cell could be potentiated by EPO for neurological recovery in children with CP without harmful effects.Trial registrationClinicalTrials.gov, NCT01991145, registered 25 November 2013.

Orphan designation: Allogeneic umbilical cord blood cells treated ex vivo with 16,16-dimethyl prostaglandin E2, Treatment of acute myeloid leukaemia

Orphan designation: Allogeneic umbilical cord blood cells treated ex vivo with 16,16-dimethyl prostaglandin E2, Treatment of acute myeloid leukaemia

Umbilical cord blood versus mesenchymal stem cells for inflammation-induced preterm brain injury in fetal sheep.

Chorioamnionitis and fetal inflammation are principal causes of neuropathology detected after birth, particularly in very preterm infants. Preclinical studies show that umbilical cord blood (UCB) cells are neuroprotective, but it is uncertain if allogeneic UCB cells are a feasible early intervention for preterm infants. In contrast, mesenchymal stem cells (MSCs) are more readily accessible and show strong anti-inflammatory benefits. We aimed to compare the neuroprotective benefits of UCB versus MSCs in a large animal model of inflammation-induced preterm brain injury. We hypothesized that MSCs would afford greater neuroprotection. Chronically instrumented fetal sheep at 0.65 gestation received intravenous lipopolysaccharide (150 ng; 055:B5, n = 8) over 3 consecutive days; or saline for controls (n = 8). Cell-treated animals received 108 UCB mononuclear cells (n = 7) or 107 umbilical cord MSCs (n = 8), intravenously, 6 h after the final lipopolysaccharide dose. Seven days later, cerebrospinal fluid and brain tissue was collected for analysis. Lipopolysaccharide induced neuroinflammation and apoptosis, and reduced the number of mature oligodendrocytes. MSCs reduced astrogliosis, but UCB did not have the same effect. UCB significantly decreased cerebral apoptosis and protected mature myelinating oligodendrocytes, but MSCs did not. UCB appears to better protect white matter development in the preterm brain in response to inflammation-induced brain injury in fetal sheep.

Preterm white matter brain injury is prevented by early administration of umbilical cord blood cells

Infants born very preterm are at high risk for neurological deficits including cerebral palsy. In this study we assessed the neuroprotective effects of umbilical cord blood cells (UCBCs) and optimal administration timing in a fetal sheep model of preterm brain injury. 50 million allogeneic UCBCs were intravenously administered to fetal sheep (0.7 gestation) at 12h or 5d after acute hypoxia-ischemia (HI) induced by umbilical cord occlusion. The fetal brains were collected at 10d after HI. HI (n=7) was associated with reduced number of oligodendrocytes (Olig2+) and myelin density (CNPase+), and increased density of activated microglia (Iba-1+) in cerebral white matter compared to control fetuses (P<0.05). UCBCs administered at 12h, but not 5d after HI, significantly protected white matter structures and suppressed cerebral inflammation. Activated microglial density showed a correlation with decreasing oligodendrocyte number (P<0.001). HI caused cell death (TUNEL+) in the internal capsule and cell proliferation (Ki-67+) in the subventricular zone compared to control (P<0.05), while UCBCs at 12h or 5d ameliorated these effects. Additionally, UCBCs at 12h induced a significant systemic increase in interleukin-10 at 10d, and reduced oxidative stress (malondialdehyde) following HI (P<0.05). UCBC administration at 12h after HI reduces preterm white matter injury, via anti-inflammatory and antioxidant actions.

Clinical safety in using unmatched allogeneic umbilical cord blood mononuclear cells transplantations in non-haematopoietic degenerative conditions.

Evaluation of safety in using unmatched human allogeneic umbilical cord blood cells for therapeutic use in individuals with non-haematopoietic degenerative conditions. The historical data and several recent immunological arguments suggest the therapeutic use of allogeneic Cord Blood Mononuclear Cells (CBMNCs), as these cells do not elicit immune response. Customarily, HLA matched cord blood MNCs are used along with prolonged immunosuppression in treatment of haematological conditions. Lately, unmatched CBMNCs are widely used in case of unavailability of HLA matched cord blood. There have been suggestions for using unmatched allogeneic cord blood MNCs for degenerative conditions without an immunoconditioning regimen. 49 patients with non-haematopoietic degenerative conditions were treated with HLA-unmatched allogeneic hUCB MNCs. Intrathecal/I.V injections (1-2 million cells/kg body weight) were given. Clinical, biochemical and haematological adverse events were evaluated. The haematological and biochemical parameters showed no major deviation from the normal. Clinically, no acute adverse effects or GVHD were observed with the used dosage. This study supports/suggests clinical safety in therapeutic medical use of unmatched allogeneic CBMNCs when used at low dosage in non-haematopoietic degenerative conditions.

The Levels of Pro-Inflammatory Factors Are Significantly Decreased in Cerebral Palsy Patients Following an Allogeneic Umbilical Cord Blood Cell Transplant

The transplantation of human umbilical cord blood cells (hUCBCs) has been shown to attenuate the unregulated activation of microglia in a rat model of cerebral palsy (CP). To investigate whether hUCBCs transplantation is also anti-inflammatory in humans, we performed a clinical trial in patients with CP. Allogeneic or autologous hUCBCs and erythropoietin (EPO) were intravenously injected into human patients with CP (mean age of approximately 38 weeks), and patients were analyzed for their motor function and social behavior. Blood samples were tested for cytokine levels. The most surprising finding in the study was that the cytokine levels were dependent on the donor cell source (allogeneic or autologous). Interestingly, the allogeneic treatment group demonstrated significantly decreased levels of pro-inflammatory factors, such as IL-1α, IL-6, TNF-β, and RANTES, and showed a statistically significant improvement in motor and social behavior compared to the autologous treatment group. Given that inflammation plays a pivotal role in CP, our results suggest that allogeneic hUCBCs therapy may be an appropriate strategy for CP treatment. In addition, prior to transplantation, a detailed analysis of the amount of proinflammatory cytokines in cord blood may be needed to avoid exacerbating inflammatory responses.

Actions of keratinocyte growth factor in leukemic mice allogeneic umbilical cord blood cell transplantation

To explore the actions of keratinocyte growth factor (KGF) in leukemic mice allogeneic umbilical cord blood cell transplantation (UCBT) and elucidate its mechanism. Peripheral blood drawn from the litters of C57BL/6 females was used as umbilical cord blood (UCB) graft. BALB/c mice were randomly divided into 7 groups (n = 12 each). The grouping was as follows. Control group 1, inoculated with leukemia. Control group 2, inoculated with leukemia at -4 d and total body irradiation (TBI) treatment. Control group 3, TBI treatment and reconstituted with 2 × 10(6) UCB-TNCs. Control group 4, injected with PBS subcutaneously, TBI treatment and reconstituted with UCB-TNCs with platelet transfusion. Control group 5, inoculated with leukemia, injected with PBS subcutaneously, TBI treatment and reconstituted with UCB-TNCs with platelet transfusion. Experiment group 1, injected with KGF subcutaneously, TBI treatment and reconstituted with UCB-TNCs with platelet transfusion. Experiment group 2, inoculated with leukemia, injected with KGF subcutaneously, TBI treatment and reconstituted with UCB-TNCs with platelet transfusion. The survival status, pathohistological changes, splenic lymphoid cell subsets and thymic output post-UCBT were compared between groups. The survival time of control group 1 was (11.1 ± 1.5) days and all died of leukemia. The survival time of control group 2 was (11.5 ± 2.5) days and all died of aplasia. Five of 12 mice of control group 3 survived for 100 days and 7 mice died of visceral hemorrhage. Four of 12 mice of control group 5 survived for 100 days and 8 mice died of leukemia with a survival rate of 33.3%. Nine of 12 mice of experiment group 2 survived for 100 days and 3 mice died of leukemia with a survival rate of 75.0%. The survival was prolonged in experiment group 2 mice as compared with that of control group 5 mice (χ² = 4.996, P = 0.0254). The splenic T, NK and B cell counts in control group 4 mice at +35 d were (9.32 ± 0.48) × 10⁶, (1.59 ± 0.11) × 10⁶ and (18.74 ± 2.01) × 10⁶ respectively. While in group 6 mice at +35 d were (13.20 ± 1.14) × 10⁶, (1.75 ± 0.12) × 10⁶ and (20.36 ± 0.86) × 10⁶ respectively. The counts of T cell and NK cell of group 6 were higher than those of group 4 (both P < 0.05). The level of signal joint T-Cell receptor excision circles (sjTRECs) in control group 4 mice was (167 ± 17) copies per 10⁵ cells while that of experiment group 1 mice (228 ± 24) copies per 10⁵ cells. They were higher than that of control mice (P = 0.002). Hematopoietic stem/precursor cells are abundant in full-term murine fetal peripheral blood. The infusion of KGF reduces the post-UCBT relapse of leukemia through the enhancement of thymic output.

Keratinocyte growth factor enhanced immune reconstitution in murine allogeneic umbilical cord blood cell transplant

Umbilical cord blood (UCB) is used increasingly as a source of hematopoietic cells because of a lower risk of graft-versus-host disease (GVHD). Myeloablative conditioning before allogeneic umbilical cord blood transplant (allo-UCBT) results in thymic epithelial cell injury and T-cell immune deficiency. Full-term fetal blood cells were used as hematopoietic cells in a previous murine allo-UCBT model with a limited number of mice surviving the myeloablative conditioning. We designed a viable murine allo-UCBT protocol with platelet concentrate support. Keratinocyte growth factor (KGF) is a mitogen of thymic epithelial cells that promotes recovery of thymic epithelium when given before total body irradiation (TBI)-containing conditioning in experimental murine models. We hypothesized that KGF pre-administration would improve post-allo-UCBT thymopoiesis. To test this hypothesis, allo-UCBT recipient mice were given KGF or control saline prior to UCBT. Platelet concentrate support significantly improved the survival rate of murine allo-UCBT recipients. KGF administration significantly increased donor-derived T and natural killer T (NKT) cells at day +35 in spleens of allo-UCBT recipients. KGF administration also improved thymic function after allo-UCBT, resulting in higher copies of signal joint T-cell receptor rearrangement excision circles (sjTRECs) in splenocytes. Finally, we found that KGF pre-administration could enhance the graft-versus-leukemia effect. In conclusion, KGF can be administered safely to recipients of allo-UCBT to enhance T-cell immune reconstitution.

Characteristics of Immune Reconstitution by Mixed Umbilical Cord Blood Transplantation In Mice: Predominant Engraftment of Major Histocompatibility Complex (MHC)-Matched Umbilical Cord Blood Stem Cell In Recipients' Bone Marrow.

AbstractAbstract 3721 Background:Currently, umbilical cord blood cells (UCBCs) are used as the primary source of hematopoietic stem cells (HSCs) for transplantation instead of bone marrow cells (BMCs). UCBC transplantation has several advantages over BMC transplantation, including the much larger size of the available donor pool, the rich proportion of hematopoietic progenitor cells, and the low content of mature T cells that might cause a graft-versus-host reaction. However, the limited quantity of cord blood samples that can be obtained from a pregnant woman is considered to be a disadvantage of UCBC transplantation. To overcome quantity limitations, the use of mixed cord blood might be necessary. Due to the lack of appropriate animal models, the analyses of the differentiation capacity of mixed UCBCs in recipients have been limited to in vivo xenogeneic experiments and clinical observations. In this study, we evaluated the reconstitution and functioning of immune systems induced by mixed UCBC transplantation in mice. Method:To deplete NK cells, female C57BL/6 (B6) [H-2b] recipient mice were intraperitoneally administered rabbit anti-asialo GM1 antibody 1 day before transplantation. On the following day, the B6 recipients irradiated with a radiation dose of 9 Gray were transplanted with 3 different combinations of mixed UCBC {[1] GFP-Tg B6 (H-2b) and C3H (H-2k); [2] GFP-Tg B6 (H-2b) and BALB/c (H-2d); and [3] C3H (H-2k) and BALB/c (H-2d)}, with each combination containing equal number of cells. Engraftment of cells in the recipients' peripheral blood was detected by flow cytometric analysis using a specific antibody against lineage markers such as CD3e (T cells), CD45R/B220 (B cells), CD11b (macrophages), or Ly-6G (granulocytes) at intervals of 4 weeks for 16 weeks. In addition, the donor origin of each lineage population was determined by H-2Kk and/or H-2Kd antibody staining. GFP+ lineage cells were found to have B6 donor origin. The dermis samples harvested from the B6, C3H, and BALB/c mice were simultaneously skingrafted on the shaved backs of the recipients at over 16 weeks after transplantation. After the recipients rejected the skin graft, alloreactive T cell and antibody responses were examined to evaluate the functional maturity of the reconstituted immune system in the recipients. Result:The survival rates of the recipient mice at 16 weeks after transplantation were as follows: (1), 73%, 8/11; (2), 92%, 12/13; and (3), 100%, 1/1. Flow cytometric analysis showed that cells of all lineages were reconstituted by only GFP+ cells in almost all the B6 recipients transplanted with mixed UCBCs: (1), 50%, 4/8 and (2), 100%, 12/12. This findings indicated predominant bone marrow engraftment of UCBC-HSCs from MHC-matched B6 donors. Furthermore, the allogeneic UCBCs in the recipients transplanted with combination [3] were eliminated from the recipients, and their lineage cells (GFP− H-2Kk− H-2Kd−) in peripheral blood were derived from the B6 recipient's own X-ray-resistant HSC in the bone marrow. The recipient mice in which the immune system was reconstituted by UCBC-HSCs from the B6 donors accepted the skin graft from the B6 donors, but completely rejected the skin grafts from the C3H and BALB/c donors. This finding indicates that both CD8+ killer and CD4+ helper T cells were functionally mature in the recipients. Furthermore, the functional competence of both cellular and humoral immunity in the recipients that rejected the skin grafts from the C3H and BALB/c donors was determined by evaluating alloreactive T cell and antibody responses against H-2k and H-2d in an in vitro experiment. Conclusion:This study showed the promising potential of transplantation of mixed UCBCs for achieving high survival rates and functional immune reconstitution. We found that only MHC-matched HSCs were involved in hematopoiesis in the recipients' bone marrow. This finding suggests the presence of a surveillance system in bone marrow with the ability to distinguish self from non-self with different MHC antigens. Disclosures:No relevant conflicts of interest to declare.

Reconstitution of B-1a Cells in Murine Allogeneic Transplantation of Umbilical Cord Blood Cells: A New Advantage of the Umbilical Cord Blood-Derived Hematopoietic System.

Abstract 2427Poster Board II-404 Background:Umbilical cord blood cells (UCBCs) have been used for the treatment of various hematological diseases. In clinical situations, perfect major histocompatibility complex (MHC) matching cannot be expected in UCBC transplantation because UCBC transplantation essentially involves non-related donor-recipient combinations. Although even MHC-mismatched UCBCs can be used because of the low content of mature T cells, the extent of recovery of immune function in lymphocytes derived from MHC-mismatched UCBC transplantation has remained unclear. To resolve this issue, we examined the functional maturation of lymphocytes derived from a fully MHC-mismatched UCBC transplantation in a murine model system. For comparison, bone marrow cell (BMC) transplantation was also performed. In this study, we first found that B-1a cells, which produce natural IgM antibodies against pathogens such as Streptococcus pneumoniae and the influenza virus, can be generated from UCBCs but not from BMCs. Methods:RAG2(-/-)BALB/c [H-2d] mice exposed to 8-Gray irradiation were transplanted with 3 different doses (1×105, 5×105, or 1×106 cells) of UCBCs or BMCs, which had been depleted of mature T cells, obtained from green fluorescent protein-transgenic C57BL/6 (B6) [H-2b] mice. More than 16 weeks after transplantation, we observed successful reconstitution of immune cells in the recipients, as shown by flow cytometric analysis. In addition, the function of T and B cells developed from UCBCs and BMCs was evaluated by examining the rejection of skin grafts from third-party donors and the production of antibodies against intraperitoneally injected 2,4,6-trinitrophenyl-keyhole limpet hemocyanin (TNP-KLH), respectively. Furthermore, the ability of UCBCs and BMCs to repopulate natural IgM antibody-producing B-1a cells was examined using flow cytometry and enzyme-linked immunosorbent assay. Results:At 16 weeks after transplantation, a similar survival rate (40-100%), which was dose-dependent, was observed between recipient mice with UCBC and BMC transplants. Phenotypically mature fluorescent cells of donor origin, including T cells, B cells, monocytes, and granulocytes, were detected in the recipients' peripheral blood, even after MHC-mismatched UCBC and BMC transplantation. Functional analysis showed that mice with allogeneic UCBC and BMC transplants accepted skin grafts from both BALB/c and C57BL/6 mice. However, these chimeric mice completely rejected skin grafts from third-party C3H/HeJ [H-2k] mice, indicating that both CD8+ killer and CD4+ helper T cells were functionally mature in the recipients. In addition, TNP-KLH-immunized mice with UCBC and BMC transplants produced both TNP-specific IgM and IgG antibodies. These findings indicate the ability of recipient mice to develop antibody responses with Ig class switching to T-dependent antigens, thereby confirming that both B cells and CD4+ helper T cells derived from allogeneic UCBC were immunologically competent. In contrast, surprisingly, flow cytometric analysis revealed the presence of peritoneal B-1a cells only in UCBC recipients but not in BMC recipients; the presence of these cells appeared to correlate with the bacterial polysaccharide-reactive natural IgM level in UCBC recipients. Conclusion:Although UCBCs and BMCs essentially have similar ability for reconstituting the hematopoietic system, this study showed the promising potential of UCBCs in reconstituting B-1a cells even in an allogeneic environment, thereby replenishing polysaccharide-reactive IgM as a front-line defensive factor against invading pathogens. On the basis of our findings, we strongly recommend the use of UCBCs in hematopoietic stem cell transplantation. Disclosures:No relevant conflicts of interest to declare.

Study On the Mechanism of Enhancing Immune Reconstitution of Keratinocyte Growth Factor in Murine Allogeneic Umbilical Cord Blood Cell Transplantation.

Abstract 3544Poster Board III-481 ObjectiveTo explore the impact and mechanism of keratinocyte growth factor (KGF) infusion on immune reconstitution post murine allogeneic umbilical cord blood cell transplantation (UCBT). MethodsMix cord blood was obtained at E19.5 from several litters of B6 females and was used as umbilical cord blood (UCB) graft. Thirty-two BALB/c mice were randomly assigned to 4 groups with 8 mice per group in the first cohort UCBT, mice were reconstituted with PBS or 1×106, 2×106, 3×106 UCB mononuclear cells (MNCs). Twenty-four BALB/c mice were randomly assigned to 3 groups with 8 mice per group in the second cohort UCBT, mice were injected with 1×106, 2×106 or 3×106 MNCs. All mice received platelet transfusion on +8d. Sixteen BALB/c mice were randomly assigned to 2 groups with 8 mice per group in the third cohort UCBT, mice were injected s.c. with KGF or PBS before TBI. All mice were injected with 2×106 MNCs and were supported with platelet transfusion on +8d. We studied survival, splenic lymphoid cell subsets, sjTREC assay after UCBT. ResultsThe survival rates at +100d of mice resconstituted with allogeneic 1×106, 2×106, 3×106 UCB MNCs were 25%, 37.5% and 37.5% in the first cohort UCBT,respectively. The survival rates at +100d of mice injected with 1×106, 2×106, 3×106 MNCs and with platelet support were 87.5%, 100.0% and 100.0%, respectively. The splenic T, NKT, NK and B cell counts in PBS treated control mice on +35d were (9.57±0.74)×106, (0.64±0.06)×106, (1.43±0.10)×106 and (19.13±1.50)×106, respectively. While in KGF treated mice on +35d were (13.47±0.74)×106, (0.89±0.03)×106, (1.79±0.04)×106 and (20.50±0.91)×106, respectively. The level of sjTREC in control mice was 182.2±10.7copies per 105 cells Gwhile that of KGF treated mice was 224.2±9.6 copied per 105 cells and was higher than that of control mice (P=0.019). ConclusionPeripheral blood abtained from E19.5d fetus is rich in hematopoietic stem cells. We established a murine allogeneic UCBT model with platelet support on +8d. KGF treatment could enhance immune reconstitution after UCBT. Disclosures:No relevant conflicts of interest to declare.

Umbilical cord blood transplantation using non-myeloablative conditioning: The Mexican experience

Twenty-eight children and adults underwent transplantation with allogeneic umbilical cord blood cells (UCB) using a non-ablative conditioning regimen; there were 15 males and 13 females. Seven patients were grafted because a non-malignant condition and 21 for a malignant disease. The median age was 8 years (range 4 months–72 years). Ten UCB were obtained from Mexican cord bloods banks, five cords were from compatible siblings and the remaining 13 cords were obtained from abroad. Median time to recover > 0.5 × 109/1 granulocytes was 24 days (range 8–32 days), whereas median time to recover > 20 × 109/1 platelets was 26 days (range 12–50 days). Twelve recipients never engrafted and recovered subsequently endogenous hematopoiesis. The non-engraftment rate was significantly higher in patients allografted for benign conditions (71 %) than in those allografted for malignant conditions (28%). The median overall post-transplant survival (SV) was 33 months and the 73-months overall SV was 39%. The cumulative incidence of grade II–IV acute GVHD and grade III–IV GVHD for the entire cohort of patients were 14 and 7%, respectively. Additional studies are needed to define if non-myeloablative conditioning is preferable over conventional conditioning in the case of UCB allografting.