Single Cord Blood Unit Research Articles

NOTCH-mediated exvivo expansion of human hematopoietic stem and progenitor cells by culture under hypoxia.

SummaryActivation of NOTCH signaling in human hematopoietic stem/progenitor cells (HSPCs) by treatment with an engineered Delta-like ligand (DELTA1ext-IgG [DXI]) has enabled ex vivo expansion of short-term HSPCs, but the effect on long-term repopulating hematopoietic stem cells (LTR-HSCs) remains uncertain. Here, we demonstrate that ex vivo culture of human adult HSPCs with DXI under low oxygen tension limits ER stress in LTR-HSCs and lineage-committed progenitors compared with normoxic cultures. A distinct HSC gene signature was upregulated in cells cultured with DXI in hypoxia and, after 21 days of culture, the frequency of LTR-HSCs increased 4.9-fold relative to uncultured cells and 4.2-fold compared with the normoxia + DXI group. NOTCH and hypoxia pathways intersected to maintain undifferentiated phenotypes in cultured HSPCs. Our work underscores the importance of mitigating ER stress perturbations to preserve functional LTR-HSCs in extended cultures and offers a clinically feasible platform for the expansion of human HSPCs.

Cord blood index predicts engraftment and early non-relapse mortality in adult patients with single-unit cord blood transplantation.

How to select optimal cord blood (CB) remains an important clinical question. We developed and validated an index of CB engraftment, the cord blood index (CBI), which uses three weighted variables representing cell doses and HLA mismatches. We modeled the neutrophil engraftment time with competing events by random survival forests for competing risks as a function of the predictors: total nucleated cells, CD34, colony-forming units for granulocytes/macrophages, and the number of HLA mismatches at the antigen and allele levels. The CBI defined three groups that had different neutrophil engraftment rates at day 30 (High, 83.7% [95% CI, 79.2-88.1%]; Intermediate, 77.0% [95% CI, 73.7-80.2%]; Low, 68.4% [95% CI, 63.6-73.2%]), platelet engraftment rates at day 60 (High, 70.4% [95% CI, 64.9-75.9%]; Intermediate, 62.3% [95% CI, 58.5-66.0%]; Low, 49.3% [95% CI, 44.2-54.5%]), and non-relapse mortality at day 100 (High, 14.1% [95% CI, 9.9-18.3%]; Intermediate, 16.4% [95% CI, 13.5-19.3%]; Low, 21.3% [95% CI, 17.1-25.5%]). This novel approach is clinically beneficial and can be adopted immediately because it uses easily obtained pre-freeze data of CB.

Optimal umbilical cord blood collection, processing and cryopreservation methods for sustained public cord blood banking

Background aimsUmbilical cord blood is an established source of stem cells in patients with hematologic malignancies who do not have HLA-compatible matched related or unrelated donors. The success of an umbilical cord blood transplant depends on the dose of total nucleated and CD34+ cells infused. Therefore, collecting, banking and listing high-quality cord blood units with high total nucleated and CD34+ cell dose are essential. MethodsHere the authors describe their cord blood bank's novel collection technique, which involves both in utero and ex utero collection of a single cord blood unit. The authors also evaluated maternal, neonatal and collection parameters that may impact the cell dose. ResultsMaternal gestational age and race, and neonatal weight and sex correlated with the total nucleated cell dose. ConclusionsThe optimized collection of umbilical cord blood is critical for its use as a source of stem cells for transplantation.

Outcomes of Unrelated Cord Blood Transplantation for 18 Patients with Severe Aplastic Anemia

Objective To evaluate the outcomes of unrelated cord blood transplantation (UCBT) for severe aplastic anemia (SAA).Methods The clinical data of 18 SAA patients, including 9 very severe aplastic anemia (vSAA), received UCBT from Sep 2009 to Feb 2017 were analyzed retrospectively. Three patients received a single unrelated cord blood unit, fifteen patients received double unrelated cord blood units. One patient was 4/6 HLA match, nine patients were 5/6 HLA match, eight patients were 6/6 HLA match. The conditioning regimens were as follow: 1 patient received modified BU/CY- based regimen, 1 patient received fludarabine (Flu) + antithymocyte globulin (ATG) + cyclophosphamide (CY) + total body irradiation (TBI) regimen, 16 patients received fludarabine (Flu) + antithymocyte globulin (ATG) + cyclophosphamide (CY) regimens. Prophylaxis for graft-versus host disease (GVHD): cyclosporine (CsA) plus short-term methotrexate (MTX).Results There was1 patient early death on day +2, 6 patients experienced primary graft rejection, but 5 of them acquired autologous myeloid recovery, 11 patients were engrafted successfully. The median days of neutrophils(ANC)above 0.5 × 109/L and platelets (PLT) more than 20 × 109/L were 18.5(9~29)days and 26.5(20~48)days, respectively. Six patients (54.5%) developed acute GVHD (aGVHD), 1 for grade I aGVHD, 3 for grade Ⅱ aGVHD, 2 for grade Ⅳ aGVHD. Of 9 patients, 2 occurred chronic GVHD (cGVHD), 1 for mild cGVHD, 1 for severe cGVHD. After a median follow-up of 56(4 ∼ 72)months, 9 patients (50.0%) died, out of which one died of severe aGVHD, the reasons of death were GVHD, severe infection, posttransplant lymphoproliferative disorders (PTLD) and transplant-associated thrombotic microangiopathy (TMA). The 5-year estimated overall survival was 46.9% ± 12.3%. No patient relapsed.Conclusion UCBT is an effective therapy for SAA patients who lack an appropriate matched related donor. DisclosuresNo relevant conflicts of interest to declare.

Kidney Dysfunction Post-Allogeneic Transplant: High Incidence of TMA and Kidney GvHD

Introduction:Kidney dysfunction is an important problem in adult recipients of allogeneic stem cell transplantation (HSCT) and independently predicts overall survival and non-relapse mortality. Kidney dysfunction occurs in ~50% of transplant recipients. More than 20% of all survivors of HSCT in long-term follow up develop chronic kidney disease. However, there is currently limited data on the cause and consequences of kidney dysfunction in adult recipients after allogeneic HSCT.Common causes of kidney dysfunction after HSCT include chemotherapy or radiation toxicity, infections, sepsis, hepatic sinusoidal obstruction, drug toxicities due to immunosuppressive medications, thrombotic microangiopathy, as well as acute or graft versus host disease (GvHD). Kidney biopsies are seldom obtained and hence the tissue diagnosis in these patients often remains speculative. Early identification of the causes of kidney dysfunction may help define preventive and therapeutic strategies and improve outcome.Hence, in August 2016, we designed a prospective study at our center to systematically evaluate kidney dysfunction in adult recipients of allogeneic HSCT. Herein we present our initial report of 8 kidney biopsies.Methods:All adult HSCT recipients with kidney dysfunction were evaluated by Nephrologists to determine the need for kidney biopsy. Kidney dysfunction was defined as either (i) acute kidney failure or (ii) proteinuria. Acute kidney failure was defined by the ‘ Kidney Disease: Improving Global Outcomes (KDIGO)‘ criteria. Proteinuria was defined as (i) spot urine albumin to creatinine ratio of ≥1 (equivalent to ≥1 g in 24 hours). Kidney biopsies were done under ultrasound guidance and the tissue specimens were interrogated by light, immunofluorescence and electron microscopy.Results:A total of 8 HSCT recipients (including one patient who had a biopsy prior to Aug 2016) underwent kidney biopsies. The patient and transplant characteristics are summarized in Table 1.Seven of the 8 pts had AML. One patient had NHL in CR2. Four patients received a MUD transplant. Two received MRD transplants. Two received combined haploidentical and single unit cord blood transplant (haplo-cord). Six of the 8 patients received conditioning with fludarabine and melphalan. One patient received additional 400 Gy TBI. One patient received additional azacytidine.Median time to neutrophil engraftment was 13.5 days (range 9-20). Median time to platelet engraftment was 19 days (range 14-76). There were no cases of graft failure. All patients also demonstrated either acute or chronic GvHD of other organs - most commonly skin.All kidney biopsies were performed without complications. Kidney biopsies were done at median 254 days after transplantation (range 126-680). Baseline serum creatinine (mg/dL) prior to transplant was 0.95 (median, range 0.65-1.1, reference range: 0.64-1.27 mg/dL). Creatinine at time of biopsy: 3.0 (median, range 1.53-5.07). Urine albumin/creatinine ratio: 832 (median, range 55-2864, reference range: <30).The kidney biopsy findings are summarized in Table 2. All patients had evidence of endothelial cell injury, either acute or chronic. Three patients had features suggestive of GvHD of the kidney while one patient had BK virus nephropathy.Four of the 8 recipients died; 80 days (median, range 39-124) from the biopsy, 346 days (median, range 178-804) from transplantation. The clinical outcome is summarized in Table 3.Conclusions:Kidney biopsies post-transplant are an important diagnostic tool that should be considered for prolonged kidney dysfunction or proteinuria, particularly in the setting of other post-transplant complications. Our single-center biopsy results highlight the under-recognition of kidney dysfunction complications and complexities observed post-transplant. All kidney biopsies performed were instrumental in treatment planning. Further investigation is needed to define criteria for diagnosis and to understand the pathophysiology. This requires multi-disciplinary engagement as well as a proactive approach for obtaining kidney biopsies. With appropriate transfusion support, percutaneous kidney biopsies were performed safely without complication. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Favorable Outcomes of Single-Unit Cord Blood Transplantation As Salvaged Treatment for Advanced Hematologic Malignancies

Treatments of advanced hematologic malignancies are limited, and the effects are very poor even after allogeneic hematopoietic stem cell transplantation. Recent study showed that cord blood transplantation was benefit for patients with pretransplantation minimal residual disease compared to both HLA-matched and HLA-mismatched unrelated donor transplantation. The relapse rate was lowest in cord blood transplantation group. But the effect of cord blood transplantation in advanced hematologic malignancies as salvaged treatment still remains unknown. We retrospectively analyzed 55 consecutive patients with advanced hematologic malignancies who received single-unit unrelated cord transplantation at our centre between December 2013 and December 2016. Among the patients, 42with acute leukemia, 8 with myelodysplastic syndrome (MDS with refractory anemia with excess blasts [RAEB]), 4 with CML-BC and 1 with plasma cell leukemia. All of them were in non-remission before transplantation. The median age was 10 years (range, 2-43 years), and the median weight was 31 kg (range, 9-82kg).All the patients received intensified conditioning regimen and a combination of CsA and MMF as GVHD prophylaxis. The infused total nucleated cell (TNC) dose was 4.99 (range 2.17-13.9) × 10⁷/kg and CD34⁺ cell dose was 2.53 (range 0.64-7.45) × 10⁵/kg. Neutrophil and platelet engraftment occurred in 49 (89.1%) and 42 (76.4%) patients, respectively. The median times to neutrophil recovery and platelet recovery were 16 days (range, 12-32 days) and 37.5 days (range, 18-83 days), respectively. Forty patients (72.7%) experienced pre-engraftment syndrome (PES). The cumulative incidence rates of II-IV acute GVHD and chronic GVHD were 25.5% (95% CI, 14.8-37.6%) and 18.8% (95% CI, 8.6-32.1%), respectively. The median follow-up period for the surviving patients was 654 days (range, 208 to 720 days). The probability of 2- year OS and LFS were 53.9% and 46.8%, respectively, whereas the cumulative incidence of 2-year NRM and relapse rate were 34.5% (95% CI, 21.6-47.8%) and 20.7% (95% CI, 10.9-32.7%), respectively. To further define the importance of matching at the HLA C locus on our special patient cohort outcomes, we analyzed the effect of HLA C disparity on patient outcomes. Results showed that although OS, LFS and NRM were not significantly different between the two groups, the relapse rate was significantly higher after transplantations with matched at HLA C(n=12) than mismatched at HLA C(n=42)(58.5% VS 27.9%,P=0.0426).Since the favorable outcomes, our results showed that single-unit cord blood transplantation might be a feasible choice as salvaged treatment for advanced hematologic malignancies, even in patients with a high leukemia burden. Choosing the cord blood with mismatch at HLA C locus might reduce the relapse rate without increasing the NRM in our special patient cohort, but this metric still requires further evaluation with a larger cohort. DisclosuresNo relevant conflicts of interest to declare.

Omidubicel vs standard myeloablative umbilical cord blood transplantation: results of a phase 3 randomized study

AbstractOmidubicel is an ex vivo expanded hematopoietic progenitor cell and nonexpanded myeloid and lymphoid cell product derived from a single umbilical cord blood unit. We report results of a phase 3 trial to evaluate the efficacy of omidubicel compared with standard umbilical cord blood transplantation (UCBT). Between January 2017 and January 2020, 125 patients age 13 to 65 years with hematologic malignancies were randomly assigned to omidubicel vs standard UCBT. Patients received myeloablative conditioning and prophylaxis with a calcineurin inhibitor and mycophenolate mofetil for graft-versus-host disease (GVHD). The primary end point was time to neutrophil engraftment. The treatment arms were well balanced and racially diverse. Median time to neutrophil engraftment was 12 days (95% confidence interval [CI], 10-14 days) for the omidubicel arm and 22 days (95% CI, 19-25 days) for the control arm (P < .001). The cumulative incidence of neutrophil engraftment was 96% for patients receiving omidubicel and 89% for patients receiving control transplants. The omidubicel arm had faster platelet recovery (55% vs 35% recovery by 42 days; P = .028), had a lower incidence of first grade 2 to 3 bacterial or invasive fungal infection (37% vs 57%; P = .027), and spent more time out of hospital during the first 100 days after transplant (median, 61 vs 48 days; P = .005) than controls. Differences in GVHD and survival between the 2 arms were not statistically significant. Transplantation with omidubicel results in faster hematopoietic recovery and reduces early transplant-related complications compared with standard UCBT. The results suggest that omidubicel may be considered as a new standard of care for adult patients eligible for UCBT. The trial was registered at www.clinicaltrials.gov as #NCT02730299.

Combined impact of HLA-allele matching and the CD34-positive cell dose on optimal unit selection for single-unit cord blood transplantation in adults

The combined effects of HLA-allele matching at six-loci (HLA-A, -B, -C, -DRB1, -DQB1, and -DPB1) and CD34+ cell dose on clinical outcomes were analyzed in 1,226 adult cases with single-unit unrelated cord blood transplantation. In the six-loci analysis, low HLA-allele matches did not significantly increase the overall mortality compared to higher matches, whereas in the five-loci analysis excluding HLA-DPB1, they caused a higher overall mortality (HR 1.42, p = .002), possibly due to the graft-versus-leukemia effect of HLA-DPB1 mismatches. A lower CD34+ cell dose (<.50 × 105/kg) resulted in higher mortality and lower engraftment; these inferior outcomes were offset by high HLA-allele matches (7-10/10 match), while the inferior outcomes of low HLA-allele matches were improved by increasing the CD34+ cell dose. Consideration of the combined effects of the CD34+ cell dose and HLA matching may expand the options for transplantable units when HLA matching or the CD34+ cell dose is inadequate.

Favorable Outcomes after Single Cord Blood Transplantation for Patients with High-Risk Hematologic Diseases: A Single-Institute Retrospective Analysis

The donor selection algorithm for cord blood (CB) with regards to matched related and unrelated donors has not been fully investigated. To assess the potential of CB transplantation (CBT) in patients with hematologic malignancies, especially for high-risk patients, we performed a single-institute retrospective analysis and compared the clinical outcomes of CBT with those of HLA-matched sibling and unrelated donor transplantation. We included 394 patients aged 16 years and older with hematologic diseases who received their first allogeneic hematopoietic cell transplantation between 1990 and 2018 at Kyoto University Hospital. These included 394 recipients of single unrelated cord blood units (UCB, n=108), HLA-matched sibling donors (MSDs, n=143), or HLA-matched unrelated donors (MUDs, n=143). There was no significant difference in relapse-free survival (RFS) between UCB, MSD, and MUD recipients (P=.975). However, we found a significant interaction between transplant year and CBT outcomes (P=.010), with significantly better outcomes observed in the more recent years. Furthermore, we found that CBT showed better RFS than matched donor transplantation (hazard ratio [HR], 0.50; 95% confidence interval [CI], 0.30 to 0.84). This impact was more prominent in high-risk patients (HR, 0.35; 95% CI, 0.16 to 0.77), with lower relapse rates (HR, 0.25; 95% CI, 0.11 to 0.54), and comparable non-relapse mortality (NRM) compared to matched donor transplantation. Extensive chronic graft-versus-host disease was less frequently observed in CBT (HR, 0.58; 95% CI, 0.26 to 1.28). CBT associated with favorable outcomes, particularly in high-risk patients, with good RFS and low relapse rates without an increase in NRM in the single-institute study. Although the findings should be externally validated, CBT might serve as a reasonable donor choice, particularly in high-risk patients.

Single cord blood transplantation for acute myeloid leukemia patients aged 60 years or older: a retrospective study in Japan

The availability of alternative donor sources could allow elderly patients to receive allogeneic hematopoietic cell transplantation (HCT). We retrospectively evaluated the outcomes of single-unit cord blood transplantation (CBT) in 1577 patients aged ≥60 years with acute myeloid leukemia (AML) in Japan between 2002 and 2017. In total, 990 (63%) patients were not in complete remission (CR) at the time of CBT. A myeloablative conditioning regimen (52%) and calcineurin inhibitor (CI) + mycophenolate mofetil (MMF)-based graft-versus-host disease (GVHD) prophylaxis (45%) were more commonly used. With a median follow-up for survivors of 31 months, the probability of overall survival and the cumulative incidence of leukemia-related mortality at 3 years was 31% and 29%, respectively. The cumulative incidence of non-relapse mortality (NRM) at 100 days and 3 years were 24% and 41%, respectively. The cumulative incidences of grade II-IV and grade III-IV acute GVHD at 100 days and extensive chronic GVHD at 2 years were 44%, 16%, and 14%, respectively. The cumulative incidence of neutrophil engraftment was 80% at 42 days. Results of multivariate analysis indicated that the following factors were significantly associated with higher overall mortality: performance status ≥1, hematopoietic cell transplantation-specific comorbidity index ≥3, adverse cytogenetics, extramedullary disease at diagnosis, and non-CR status at CBT. By contrast, female sex, HLA disparities ≥2, mycophenolate mofetil-based GVHD prophylaxis, and recent CBT were significantly associated with lower overall mortality. In conclusion, single CBT offers a curative option for AML patients aged ≥60 years with careful patient selection.

RXR Negatively Regulates Ex Vivo Expansion of Human Cord Blood Hematopoietic Stem and Progenitor Cells.

Ex vivo expansion of human cord blood (CB) hematopoietic stem cells (HSCs) is one approach to overcome limited numbers of HSCs in single CB units. However, there is still no worldwide acceptable HSC ex vivo expansion system. A main reason is that we still have very limited knowldege regarding mechanisms underlying maintenance and expansion of CB HSCs. Here we report that retinoid X receptor (RXR) activity is of significance for CB HSC ex vivo expansion. RXR antagonist HX531 significantly promoted ex vivo expansion of CB HSCs and progenitor cells (HPCs). RXR agonist Bexarotene notably suppressed ex vivo expansion of CB HSCs. Activation of RXR by Bexarotene significantly blocked expansion of phenotypic HSCs and HPCs and expressed increased functional HPCs as assessed by colony formation induced by UM171 and SR1. In vivo transplantation experiments in immune-deficient mice demonstrated that HX531 expanded CB HSCs possess long-term reconstituting capacities, and Bexarotene treatment inhibited expansion of functional CB HSCs. RNA-seq analysis revealed that RXR regulates expression of FBP1 (a negative regulator of glucose metabolism) and many genes involved in differentation. ECAR analysis showed that HX531 significantly promoted glycolytic activity of CB CD34+ HSCs and HPCs. Our studies suggest that RXR is a negative regulator of ex vivo expansion of CB HSCs and HPCs.

Impact of a prior history of cancer on prognosis after myeloablative single-unit cord blood transplantation.

A prior history of cancer was associated with higher non-relapse mortality or overall mortality in patients undergoing allogeneic haematopoietic cell transplantation. Because it is unclear whether the outcomes after cord blood transplantation are influenced by a prior history of cancer, we retrospectively assessed the prevalence and prognostic impact of a prior history of cancer in adult patients undergoing myeloablative single-unit cord blood transplantation in our institute between 2004 and 2020. The univariate analysis showed that a prior history of cancer did not affect the probability of overall survival; the cumulative incidence of relapse; or non-relapse mortality. In the multivariate analysis, prior history of cancer was not associated with overall mortality, relapse or non-relapse mortality. No patients with a prior history of cancer had experienced prior cancer relapse. A prior history of cancer was not associated with non-relapse mortality or overall mortality following single-unit cord blood transplantation.

Umbilical cord blood transplantation: a maturing technology.

The use of umbilical cord blood (CB) as a source of hematopoietic progenitor cells for patients with high-risk hematologic disorders receiving allogeneic hematopoietic cell transplantations (HCTs) has increased significantly. Single-institution and registry studies have shown a decreased relapse rate and an increased transplantation-related mortality rate with similar overall survival rates after allogeneic HCT with CB compared with other donor sources. The transplantation of double CB units has overcome the dose limitation inherent in a single CB unit and thus has markedly extended the use of CB to larger children and adults. Similarly, the use of reduced intensity conditioning in the CB transplantation setting has allowed the treatment of older patients who would be unable to tolerate the myeloablative regimens used in the original CB transplantation protocols.

Use of the HLA-B leader to optimize cord blood transplantation

Cord blood transplantation (CBT) can cure life-threatening blood disorders. The HLA-B leader affects the success of unrelated donor transplantation but its role in CBT is unknown. We tested the hypothesis that the HLA-B leader influences CBT outcomes in unrelated single-unit cord blood transplants performed by Eurocord/European Blood and Marrow Transplant (EBMT) centers between 1990 and 2018 with data reported to Eurocord. Among 4,822 transplants, 2,178 had one HLA-B mismatch of which 1,013 were HLAA and HLA-A and -DRB1 matched. The leader (methionine [M] or threonine [T]) was determined for each HLA-B allele in patients and units to define the genotype. Among single HLA-B-mismatched transplants, the patient/unit mismatched alleles were defined as leader-matched if they encoded the same leader, or leader-mismatched if they encoded different leaders; the leader encoded by the matched (shared) allele was determined. The risks of graft-versus-host disease, relapse, non-relapse mortality and overall mortality were estimated for various leader-defined groups using multi-variable regression models. Among the 1,013 HLA-A and -DRB1-matched transplants with one HLA-B mismatch, increasing numbers of cord blood unit M-leader alleles was associated with increased risk of relapse (hazard ratio [HR] for each increase in one Mleader allele 1.30, 95% Confidence Interval [CI]: 1.05-1.60, P=0.02). Furthermore, leader mismatching together with an M-leader of the shared HLA-B allele lowered non-relapse mortality (HR 0.44, 95% CI: 0.23-0.81; P=0.009) relative to leader matching and a shared T-leader allele. The HLA-B leader may inform relapse and non-relapse mortality risk after CBT. Future patients might benefit from the appropriate selection of units that consider the leader.

TNFSF15 facilitates human umbilical cord blood haematopoietic stem cell expansion by activating Notch signal pathway.

The lack of efficient ex vivo expansion methods restricts clinical use of haematopoietic stem cells (HSC) for the treatment of haematological malignancies and degenerative diseases. Umbilical cord blood (UCB) serves as an alternative haematopoietic stem cell source. However, currently what limits the use of UCB‐derived HSC is the very low numbers of haematopoietic stem and progenitor cells available for transplantation in a single umbilical cord blood unit. Here, we report that TNFSF15, a member of the tumour necrosis factor superfamily, promotes the expansion of human umbilical cord blood (UCB)‐derived HSC. TNFSF15‐treated UCB‐HSC is capable of bone marrow engraftment as demonstrated with NOD/SCID or NOD/Shi‐SCID/IL2Rgnull (NOG) mice in both primary and secondary transplantation. The frequency of repopulating cells occurring in the injected tibiae is markedly higher than that in vehicle‐treated group. Additionally, signal proteins of the Notch pathway are highly up‐regulated in TNFSF15‐treated UCB‐HSC. These findings indicate that TNFSF15 is useful for in vitro expansion of UCB‐HSC for clinical applications. Furthermore, TNFSF15 may be a hopeful selection for further UCB‐HSC application or study.

Recombinant human thrombopoietin promotes platelet engraftment after umbilical cord blood transplantation

Delayed platelet engraftment is a common complication after umbilical cord blood transplantation (UCBT) accompanied by increased transplant-related complications or death. This study was designed to determine the safety and efficacy of recombinant human thrombopoietin (rhTPO) in promoting platelet engraftment after UCBT. A total of 120 patients scheduled to receive UCBT were randomly assigned to the rhTPO group (300 U/kg once daily from days 14 to 28 after UCBT, n = 60) or the control group (n = 60). The primary outcome was the 60-day cumulative incidence of platelet engraftment after single-unit cord blood transplantation. The 60-day cumulative incidence of platelet engraftment (platelet count ≥20 × 109/L) and the 120-day cumulative incidence of platelet recovery (platelet count ≥50 × 109/L) were both significantly higher in the rhTPO group than in the control group (83.1% vs 66.7%, P = .020; and 81.4% vs 65.0%, P = .032, respectively). In addition, the number of required platelet infusions was significantly lower in the rhTPO group than in the control group (6 vs 8 units, respectively; P = .026). The cumulative incidence of neutrophil engraftment and the probability of 2-year overall survival, disease-free survival, and graft-versus-host disease-free relapse-free survival did not differ between the 2 groups. Other transplant-related outcomes and complications did not differ between the 2 groups, and no severe adverse effects were observed in patients receiving rhTPO. This study demonstrated that rhTPO is well tolerated in patients and could effectively promote platelet engraftment after UCBT. This study was registered on the Chinese Clinical Trial Registry (http://www.chictr.org.cn/index.aspx) as ChiCTR-IPR-16009357.

Impact of Intestinal Microbiota on Reconstitution of Circulating Monocyte, Dendritic Cell, and Natural Killer Cell Subsets in Adults Undergoing Single-Unit Cord Blood Transplantation.

The intestinal microbiota plays a fundamental role in the development of host innate immune cells, such as monocytes, dendritic cells (DCs), and natural killer (NK) cells. We examined the association between intestinal microbiota and subsequent immune reconstitution of circulating monocyte, DC, and NK cell subsets in 38 adult patients undergoing single-unit cord blood transplantation (CBT). A higher diversity of intestinal microbiota at 1 month was significantly associated with higher counts of plasmacytoid DCs at 7 months after CBT, as measured by the Chao1 index. Principal coordinate analysis of unweighted UniFrac distances showed significant differences between higher and lower classical monocyte reconstitution at 7 months post-CBT. The families Neisseriaceae, Burkholderiaceae, Propionibacteriaceae, and Coriobacteriaceae were increased in higher classical monocyte reconstitution at 7 months post-CBT, whereas the family Bacteroidaceae was increased in lower classical monocyte reconstitution at 7 months post-CBT. These data show that intestinal microbiota composition affects immune reconstitution of classical monocyte and plasmacytoid DCs following single-unit CBT.

Cord blood transplants supported by unrelated donor CD34+ progenitor cells.

Alternative donor transplantation with the haplo-cord platform allows the use of a lower-dose single umbilical cord blood unit (CBU) by co-infusion of third-party CD34+-selected cells from a haploidentical relative, which provides early transient engraftment while awaiting durable CBU engraftment. In our experience, ~15% of patients lack a suitable haploidentical donor. Here we report 26 patients who underwent haplo-cord transplant using CD34+-selected partially matched unrelated donor grafts. Twenty-four were conditioned with fludarabine/melphalan +/- low-dose TBI (n = 16). Twenty-five received ATG and all received posttransplant tacrolimus and mycophenolate mofetil. Median time to neutrophil and platelet recovery was 11 and 18 days. CBU engraftment, with CD33 and CD3 >5% cord chimerism in the myeloid/lymphoid compartment by day +60, occurred in 20 of 24 patients (83%). Incidence of grade 2-4 acute graft-versus-host disease (GVHD) was 27% at day +100, and chronic GVHD was 4% at 1 year. Overall survival at 1 year was 54%. For patients in need of an alternative transplant who lack a haploidentical donor, haplo-cord transplantation using CD34+-selected partially matched unrelated donor grafts results in rapid engraftment with no increased rate of cord graft failure or GVHD.

Mechanistic Basis of ex Vivo Umbilical Cord Blood Stem Progenitor Cell Expansion.

Umbilical cord blood (CB) transplantation has been used successfully in humans for three decades due to its rapid availability for patients lacking a suitable allogeneic donor, less stringent HLA matching requirements, and low rates of relapse and chronic graft-versus-host disease (GVHD). However, CB transplantation is associated with complications, such as delayed hematopoietic engraftment, graft failure, which increases infection and bleeding and causes longer hospital stays, and transplant-related mortality. The majority of these biological limitations are due to the unforeseeable functional potency of multipotent hematopoietic stem cells (HSCs), which reduce the predictability of successful transplantation; however, several strategies have been developed to increase the number of hematopoietic stem progenitor cells (HSPCs) infused during CB transplantation. This review primarily addresses the methods that promote ex vivo CB expansion within the context of symmetrical and asymmetrical HSC division and those that rely on epigenetic mechanisms, along with the reportedly most successful cytokine combinations. We also review recent clinical research on small molecules (StemRegenin-1, UM171, and nicotinamide) in ex vivo expanded CB and discuss yet unvalidated preclinical strategies. Expanding and transplanting CB graft enriched in HSPCs in a single CB unit is a particularly exciting prospect with the potential to improve the use and availability of CB grafts. Greater knowledge of optimal ex vivo expansion strategies, cell longevity, and graft potency will expand the scope of cellular therapies. Also the development of adequate ex vivo HSPC expansion strategies could bring expanded cord blood grafts to the forefront of transplant therapy and regenerative medicine.

An off the shelf, GMP compliant,fully closed and semi-automated large-scale production system for allogeneic NK cells

Background & Aim Natural killer cells (NK) have been on the mainstay in cancer treatment for over two decades. Most of the clinical trials till date, have either used autologous or allogeneic NK cells that were often produced in limited quantities to tailor the needs of a single patient. With the highly unmet need to treat large cohort of patients and with multiple doses, a true off the shelf approach is the only way. Glycostem began to envision this, by setting up a world's first completely closed manufacturing platform for allogeneic NK cells (oNKord®) from fresh umbilical cord blood stem cells. A closed system is a process where the cells are never exposed to the open environment to avoid a potential risk for contamination. Methods, Results & Conclusion Glycostem manufactures oNKord® at its own state of the art Class C/D GMP clean room, using PVC Sterile Connection Device and Sealer (Terumo), C-Flex Biowelder and Sealer (Sartorius), to connect and seal two tubes aseptically and keeping the system closed. These equipments are key elements in the routine functioning of the closed system, while cells are transferred, or medium is added. In our process, fresh cord blood is used as a starting material, using fully automated CliniMACS Prodigy to select CD34+ cells, which are then expanded using static cultures in incubators and further differentiated into mature NK cells in Xuri bioreactors. After 40 days of culture, NK cells are harvested, washed and aliquoted into cryobags and cryopreserved in controlled rate freezing device. Optimised freezing and thawing conditions, which result in high recovery and product functionality, enabled cell banking of multiple infusion doses produced from one manufacturing batch. Frozen doses are shipped to clinical centres in temperature controlled dry shippers and thawed using waterbath at the time for infusion. The NK cell product is scalable resulting in a 2000 - 3000-fold expansion, averaging up to 11 billion NK cells from a single cord blood unit (n=3). Validated QC testing methods, in process controls using flow cytometry assays and standardised potency assays are in place to address product safety, identity and functionality for release. Setting up of a completely closed system faces many challenges, and during this journey over the last two years, Glycostem has successfully resolved several bottlenecks and worked closely with the regulatory authorities in achieving an “universal off the shelf” NK cell therapy product.