Patients Undergoing Cord Blood Transplantation Research Articles

Effect of Hematopoietic Stem Cell Transplantation Regimen on Tacrolimus Pharmacokinetics.

Treatment with tacrolimus requires strict control of the whole-blood concentration in patients undergoing allogeneic hematopoietic stem cell transplantation (HSCT). In patients undergoing cord blood transplantation (CBT), there is a negative correlation between volume of distribution of tacrolimus and hemoglobin levels, which reflect the red blood cell (RBC) count. In this study, we evaluated the influence of the conditioning regimen (myeloablative and reduced-intensity conditioning) or donor source (cord blood, bone marrow, and peripheral blood stem cells) on the pharmacokinetics of tacrolimus in patients undergoing HSCT, including those undergoing CBT. We also examined applicability of dosing strategy of tacrolimus considering the RBC count. We retrospectively analyzed clinical data-including whole-blood tacrolimus concentrations-from patients with HSCT. The observation period spanned from first continuous intravenous infusions until switch to oral medication, transfer to another hospital, relapse, or death. Population pharmacokinetic analysis was performed on whole-blood tacrolimus concentrations obtained from therapeutic drug monitoring during the observation period. Patient characteristics and laboratory data were evaluated as covariates. We enrolled 91 patients undergoing HSCT (CBT: n = 56; bone marrow transplantation: n = 22; and peripheral blood stem cell transplantation: n = 13); 58 and 33 patients received myeloablative conditioning and reduced-intensity conditioning, respectively. Whole-blood tacrolimus concentrations were accurately captured (n = 1,658 measurements) using a one-compartment and additive error model. The conditioning regimen and donor source did not have an impact on the pharmacokinetics of tacrolimus. Therefore, these factors were not considered when forming the dosing strategy. Nevertheless, a negative correlation between volume of distribution and hemoglobin level was confirmed, indicating that monitoring the RBC count is useful in assessing the dosing strategy. A tacrolimus dosing strategy that considers the variability in hemoglobin levels applies to all patients undergoing HSCT.

Infusion of Non-HLA-Matched Off-the-Shelf Ex Vivo Expanded Cord Blood Progenitors in Patients Undergoing Cord Blood Transplantation: Result of a Phase II Clinical Trial.

Recipients of myeloablative cord blood transplants (CBT) are known to experience delayed hematopoietic recovery and an increased risk of transplant related mortality (TRM). We developed methods for ex vivo expansion and cryopreservation of CB stem and progenitor cells. 15 patients with hematologic malignancies were enrolled in this single center phase II trial between September 2010 and August 2012 to assess the safety of infusing a non-HLA-matched expanded CB product to bolster a conventional CBT. On the day of transplant, an infusion of the expanded CB product followed the primary graft (1 or 2 unmanipulated CB units). All patients engrafted. Median time to neutrophil and platelet recovery was 19 and 35 days, respectively. Early myelomonocytic recovery was almost entirely due to cells arising from the non-HLA-matched expansion product and were no longer detected at day 14 in all but 2 patients. The probability of 3-years disease free survival was 86%. No TRM was observed throughout the study period, and only 2 patients relapsed. All patients presented with grade II acute graft-versus-host disease (aGVHD) at a median time of 32 days, with no grade III-IV aGVHD observed. At 2 years only 2 patients remain on immunosuppressive therapy for mild chronic GVHD. This phase II safety study demonstrate that infusion of an off-the-shelf non-HLA-matched expanded CB product in addition to a conventional CB graft was safe and led to sustained myeloid recovery. Based on these encouraging results, a prospective multicenter randomized trial utilizing this product has been conducted and results will be soon released. ClinicalTrials.gov Identifier: NCT01175785.

Prognostic factors of children and adolescents with T-cell acute lymphoblastic leukemia after allogeneic transplantation.

Acute lymphoblastic leukemia (ALL) is the most common cancer during childhood, and some high-risk patients with ALL require hematopoietic stem cell transplantation (HSCT). Mainly due to small patient numbers, studies focusing specifically on children and adolescents with T-cell ALL (T-ALL) are limited. Using a nationwide registry, we retrospectively analyzed data from patients under 20years old who underwent their first HSCT for T-ALL between 2000 and 2018. As a result, total 484 patients were included, and their median follow-up period was 6.9years after HSCT for survivors. While patients receiving HSCT at first complete remission (CR) showed relatively good 5-year leukemia free survival (5yLFS, 73.5%), once relapse occurred, their prognosis was much worse (44.4%) even if they attained second remission again (p<0.001). Among patients receiving HSCT at CR1, grade II-IV acute graft versus host disease was associated with worse overall and LFS than grade 0-I (5yLFS 69.5% vs. 82.1%, p=0.026) mainly due to high non-relapse mortality. Among those patients, patients receiving related bone marrow transplantation, unrelated bone marrow transplantation, or unrelated cord blood transplantation showed similar survival (5yLFS, 73.2%, 76.3%, and 77.0%, respectively). For patients undergoing cord blood transplantation at CR1, total-body irradiation-based myeloablative conditioning was associated with better 5yLFS than other conditioning regimens (85.4% vs. 62.2%, p=0.044), as it reduced the risk of relapse. These results indicate that relapsed patients have much less chance of cure, and that identifying patients who require HSCT for cure and offering them HSCT with optimal settings during CR1 are crucial for children and adolescents with T-ALL.

Lower vancomycin trough levels in adults undergoing unrelated cord blood transplantation

Vancomycin (VCM) is frequently used for neutropenic patients undergoing cord blood transplantation (CBT). We retrospectively examined the relationship between VCM trough levels and the efficacy and toxicity in 122 adult patients undergoing CBT in our institute. The median initial dose of VCM based on body weight was 9.1 mg/kg/dose (range, 6.0–22.6 mg/kg/dose). The median initial trough level of VCM for all patients was 4.50 µg/mL (range, 1.20–24.05 µg/mL), at a median of 3 days (range, 2–6 days) after VCM administration. The cumulative incidence of acute kidney injury (AKI) was 19% at 30 days after VCM administration. A higher median trough level of VCM during the first 7 days was significantly associated with the development of AKI in the multivariate analysis (Hazard ratio: 1.28, p = .026). These data suggest that a lower VCM trough level may be safe in adult patients undergoing CBT under therapy with nephrotoxic drugs.

Akraba Dışı Kordon Kanı Kaynaklı Kök Hücre Transplantasyonu Retrospektif Analizi

Objective: Cord blood stem cells are used for hematopoietic stem cell transplantation (HSCT) in the treatment of hematological malignant and non-malignant diseases, especially in pediatric patients. In this present study, we aimed to evaluate the retrospective analysis of HLA match and survival between donor and patients who underwent cord blood transplantation through the Istanbul University Faculty of Medicine Bone Marrow Bank. Materials and Methods: In order to research the related factors of umbilical cord blood transplantation in Turkey, 102 cases of umbilical cord blood transplantation performed between 2008 and 2014 were analyzed retrospectively. Cord blood unit search applications were performed through Istanbul University, Istanbul Medical Faculty, Bone Marrow Bank. Results: It was, on average, 45 days between the search request and transplantation (range: 11-113 days). Average age of the patients was 42.48±2.8 (1-192 months), male-to-female ratio was 64/38. The most frequent diseases resulting in transplantation were SCID (n=23), AML (n=13), osteopetrosis (n=10), ALL (n=10) and other diseases. There were 38 (37%) cord blood transplantations with matched HLA-A,-B,-DR; 54 (52%) mismatched for HLA-A or -B antigens; 9 (9%) mismatched for HLA-DR only and 1 (1%) mismatched for HLA-B and -DR antigens. The overall survival rate after HSCT with cord blood was 81.5% (n=31/39). The overall survival rate of the patients with HLA class-I antigens mismatch is 48.1% (26/54), the patients with HLA classII antigens mismatch was 22.2% (2/9) after the HSCT. Conclusion: In conclusion, we found a significant effect in patients undergoing cord blood transplantation with 6/6 HLA matching on overall survival.

Retrospective analysis of the correlation between tacrolimus concentrations measured in whole blood and variations of blood cell counts in patients undergoing allogeneic haematopoietic stem cell transplantation.

Tacrolimus is administered to patients undergoing haematopoietic stem cell transplantation (HSCT) as prophylaxis for graft-versus-host disease. As a high blood tacrolimus concentration within a narrow therapeutic range must be maintained after HSCT, therapeutic drug monitoring (TDM) is necessary. We investigated the correlation between blood tacrolimus concentration and blood cell count in HSCT patients to assess how changes in blood cell count affect tacrolimus TDM. A retrospective analysis was performed for 24 patients who underwent allogeneic HSCT and received tacrolimus. The correlation between variations in blood tacrolimus concentration and blood cell count was evaluated for three consecutive weeks, starting 1 week after HSCT. Variations in blood tacrolimus concentration were significantly correlated with variations in red blood cell (RBC) count, haemoglobin level and haematocrit value, but not with variations in white blood cell or platelet counts. Further, the above variations were significantly correlated in patients undergoing cord blood transplantation and peripheral blood stem cell transplantation, but not in those undergoing bone marrow transplantation. These findings demonstrate that RBC count is associated with variations in blood tacrolimus concentration, with the relevance of this association depending on the source of transfused stem cells. Thus, variations in RBC count might be useful for tacrolimus TDM.

Pharmacokinetic and Pharmacodynamic Markers of Mycophenolic Acid Associated with Effective Prophylaxis for Acute Graft-Versus-Host Disease and Neutrophil Engraftment in Cord Blood Transplant Patients

Mycophenolate mofetil (MMF) is a prodrug of mycophenolic acid (MPA) and is frequently used to prevent acute graft-versus-host disease (aGVHD) in patients receiving hematopoietic stem cell transplantation (HSCT). However, optimal doses of MMF and target MPA concentrations in HSCT patients have not been defined. In this study, relationships between pharmacokinetic or pharmacodynamic markers of MPA and successful aGVHD prevention and neutrophil engraftment were evaluated to inform individualized MPA treatments in HSCT patients. We recruited 35 patients undergoing cord blood transplantation (CBT) who were treated with MMF. Area under the concentration–time curves from 0 to 24 hours (AUC0-24) for free MPA and MPA acyl glucuronide (AcMPAG) at 1 week after the start of MMF treatments were significantly higher in patients with gastrointestinal (GI) aGVHD at stage ≥I than those at stage 0. Patients with faster neutrophil engraftment had higher free MPA AUC0-24 at 1 week after the start of MMF treatments compared with those with slower neutrophil engraftment. Inosine-5′-monophosphate dehydrogenase activity in peripheral blood mononuclear cells and single nucleotide polymorphisms in genes that were previously associated with MPA pharmacokinetics and pharmacodynamics were not an independent predictor for the clinical outcomes. Receiver-operating characteristic model analyses showed that cutoff values of AUC0-24 for successful GI aGVHD prevention were .689 and 15.6 µg⋅hour⋅mL−1 for free MPA and AcMPAG, respectively. In addition, the cut-off value of free MPA AUC0-24 for neutrophil engraftment by day 25 was .405 µg⋅hour⋅mL−1. In conclusion, free MPA AUC0-24 may be a better predictor of the prevention of GI aGVHD and neutrophil engraftment compared with that of total MPA in patients receiving CBT. Hence, monitoring of the free MPA AUC0-24 between .405 and .689 µg⋅hour⋅mL−1 could be considered informative of individualized MPA treatments in CBT patients.

Human monocyte-targeted non-viral siRNA delivery for the suppression of inflammatory macrophages

Mycophenolate mofetil (MMF) is a prodrug of mycophenolic acid (MPA) and is frequently used to prevent acute graft-versus-host disease (aGVHD) in patients receiving hematopoietic stem cell transplantation (HSCT). However, optimal doses of MMF and target MPA concentrations in HSCT patients have not been defined. In this study, relationships between pharmacokinetic or pharmacodynamic markers of MPA and successful aGVHD prevention and neutrophil engraftment were evaluated to inform individualized MPA treatments in HSCT patients. We recruited 35 patients undergoing cord blood transplantation (CBT) who were treated with MMF. Area under the concentration–time curves from 0 to 24 hours (AUC0-24) for free MPA and MPA acyl glucuronide (AcMPAG) at 1 week after the start of MMF treatments were significantly higher in patients with gastrointestinal (GI) aGVHD at stage ≥I than those at stage 0. Patients with faster neutrophil engraftment had higher free MPA AUC0-24 at 1 week after the start of MMF treatments compared with those with slower neutrophil engraftment. Inosine-5′-monophosphate dehydrogenase activity in peripheral blood mononuclear cells and single nucleotide polymorphisms in genes that were previously associated with MPA pharmacokinetics and pharmacodynamics were not an independent predictor for the clinical outcomes. Receiver-operating characteristic model analyses showed that cutoff values of AUC0-24 for successful GI aGVHD prevention were .689 and 15.6 µg⋅hour⋅mL−1 for free MPA and AcMPAG, respectively. In addition, the cut-off value of free MPA AUC0-24 for neutrophil engraftment by day 25 was .405 µg⋅hour⋅mL−1. In conclusion, free MPA AUC0-24 may be a better predictor of the prevention of GI aGVHD and neutrophil engraftment compared with that of total MPA in patients receiving CBT. Hence, monitoring of the free MPA AUC0-24 between .405 and .689 µg⋅hour⋅mL−1 could be considered informative of individualized MPA treatments in CBT patients.

40 - Allogeneic Hematopoietic Stem Cell Transplantation for Patients Aged 60 Years or Older with Myelodysplastic Syndrome in Japan

Mycophenolate mofetil (MMF) is a prodrug of mycophenolic acid (MPA) and is frequently used to prevent acute graft-versus-host disease (aGVHD) in patients receiving hematopoietic stem cell transplantation (HSCT). However, optimal doses of MMF and target MPA concentrations in HSCT patients have not been defined. In this study, relationships between pharmacokinetic or pharmacodynamic markers of MPA and successful aGVHD prevention and neutrophil engraftment were evaluated to inform individualized MPA treatments in HSCT patients. We recruited 35 patients undergoing cord blood transplantation (CBT) who were treated with MMF. Area under the concentration–time curves from 0 to 24 hours (AUC0-24) for free MPA and MPA acyl glucuronide (AcMPAG) at 1 week after the start of MMF treatments were significantly higher in patients with gastrointestinal (GI) aGVHD at stage ≥I than those at stage 0. Patients with faster neutrophil engraftment had higher free MPA AUC0-24 at 1 week after the start of MMF treatments compared with those with slower neutrophil engraftment. Inosine-5′-monophosphate dehydrogenase activity in peripheral blood mononuclear cells and single nucleotide polymorphisms in genes that were previously associated with MPA pharmacokinetics and pharmacodynamics were not an independent predictor for the clinical outcomes. Receiver-operating characteristic model analyses showed that cutoff values of AUC0-24 for successful GI aGVHD prevention were .689 and 15.6 µg⋅hour⋅mL−1 for free MPA and AcMPAG, respectively. In addition, the cut-off value of free MPA AUC0-24 for neutrophil engraftment by day 25 was .405 µg⋅hour⋅mL−1. In conclusion, free MPA AUC0-24 may be a better predictor of the prevention of GI aGVHD and neutrophil engraftment compared with that of total MPA in patients receiving CBT. Hence, monitoring of the free MPA AUC0-24 between .405 and .689 µg⋅hour⋅mL−1 could be considered informative of individualized MPA treatments in CBT patients.

Myeloablative Cord Blood Transplantation Yields Excellent Disease Free Survival in Patients with Acute Lymphoblastic Leukemia

Background: Allogeneic hematopoietic cell transplantation (HCT) is a potential curative treatment for children and adults with acute lymphoblastic leukemia (ALL). However, relapse occurs in approximately 30% of patients with ALL after HCT, with even worse outcomes in patients with minimal residual disease (MRD) pre-transplant [1]. Lower relapse rates have been reported for patients undergoing cord blood transplantation (CBT) compared to unrelated donor transplant. In the present study we sought not only to evaluate outcomes for ALL patients undergoing a myeloablative CBT, but also to see if the outcomes varied according to MRD status.Methods: Using prospectively collected data, we reviewed 67 consecutive patients with ALL who received a CBT between 2006 and 2016. The conditioning regimens consisted of either TBI (1320 cGy), Cytoxan and Fludarabine (FLU), (n=49) or Treosulfan (Treo), FLU and 200 cGY TBI (n=18). GVHD prophylaxis consisted of cyclosporine and mycophenolate mofetil (MMF). Donor units were selected to be at least a 4 out of 6 match to recipient at HLA-A and HLA-B antigens and HLA-DRB allele. Ten-color multiparameter flow cytometry studies on bone marrow aspirates were performed before HCT to determine the presence of MRD. Disease free-survival (DFS) was evaluated using the method of Kaplan and Meier. Probabilities of non-relapse mortality (NRM) and relapse were summarized using cumulative incidence estimates using the appropriate competing risks.Results: Patient characteristics are shown in Table 1. The median age and weight was 22 years (range, 0.6-58 years) and 57 kg (range 8-99 kg), respectively. Thirty-nine patients (58%) were non-Caucasian. At the time of transplant 32 patients (47%) were in first complete remission (CR1), 28 (42%) in CR2 and 7 (11%) in CR≥3. Any level of residual disease was considered MRD-positive (n=22). Fifteen patients (22%) had Ph+ ALL. Nine patients (13%) had a prior allogeneic transplant. The majority of patients (n=48; 72%) received a double-CB graft; the rest received a single-CB graft. In addition, 14 patients (21%) received an ex-vivo-expanded CB unit as part of their graft, combined with an unmanipulated CB unit. Median time to engraftment was 31 days (range, 21-80 days). The overall DFS at 5 years was 74% (CI 95%: 60-83) with no difference between MRD-negative 78% (CI 95%: 61-88) and MRD-positive patients 67% (CI 95%:43-81) (p=0.40) [Figure 1A and 1B]. The overall cumulative incidence of relapse and NRM at 5 years post-transplant was 14% (CI 95%: 5-31) and 12% (95%: 3-26), respectively. The risk of relapse was not significantly higher in MRD-positive compared to MRD-negative (HR 1.80 (95 CI: 0.50-6.51) p=0.36), when adjusting for year of transplant, CMV serostatus, age, and disease risk. The cumulative incidence of acute GVHD grade II-IV and III-IV at day 100 was 86% (CI 95%: 75-92) and 16% (CI 95%: 8-26), respectively.Conclusions: Our data suggest that outcomes in patients with ALL following myeloablative CBT are remarkable with 74% DFS at 5 years and very low rate of relapse. The implication of these results is particularly important for patients from racial and ethnic minorities. Indeed, because mismatches in CB units can be tolerated, it is possible to find suitable donors for nearly all patients, regardless of their race/ethnicity. Furthermore, the risk of relapse was similar between MRD-negative and positive patients, although this requires further study in larger populations given the relatively low number of MRD-positive patients.

Bloodstream Infections in Adult Patients Undergoing Cord Blood Transplantation from Unrelated Donors after Myeloablative Conditioning Regimen

The incidence, epidemiology, and risk factors of bloodstream infection (BSI) and their impact on transplant outcomes after umbilical cord blood transplantation (UCBT) are not well defined. Between May 1997 and December 2012, 202 isolates in 189 episodes of BSI were registered in 134 of 241 patients who underwent single-unit myeloablative UCBT. Cumulative incidence (CI) of developing at least 1 episode of BSI was 21%, 29%, 34%, 42%, and 52% at days +7, +14, +30, +100, and +365, respectively. The median time of onset for the first BSI episode was day +10 (range, day –7 to +1217). Early BSI before day 7 was associated with increased nonrelapse mortality (relative risk [RR], 1.5; 95% confidence interval [CI], 1.1 to 2.3; P = .04), whereas BSI before day 14 was an independent adverse risk factor for neutrophil recovery (RR, .6; 95% CI, .5 to .9; P = .002). A higher CD8+ cell dose of the graft was the only variable independently associated with reduced risk of BSI (RR, .1; 95% CI, .02 to .7; P = .02). The gram-negative rod (GNR) to gram-positive bacteria ratio was .9 before day +30 and 1.6 thereafter (P = .03). Escherichia coli (31%) and Pseudomonas sp. (28%) were the most frequently isolated among GNR. The overall crude mortality rate was 12% at day 7 and was higher for GNR (18%) compared with gram-positive bacteria (7%) (P = .03). These findings emphasize the importance of preventing bacterial infections during conditioning and the very early post-UCBT period.

Real-Time Immunophenotyping of Peripheral Blood Early Post Myeloablative Cord Blood Transplant Can Identify Patients at Risk for Primary Graft Failure

▪Introduction: Primary graft failure (PGF) is a potentially life threatening complication following hematopoietic cell transplantation. Patients undergoing cord blood transplantation (CBT) are at higher risk for PGF and also experience delayed hematopoietic recovery. The ability to distinguish between PGF and delayed engraftment is critical for correct clinical management. Limited data exists analyzing the kinetics of engraftment of specific circulating cell lineages within 14 days after double CBT (dCBT). Herein, we investigate the potential for real time immunophenotyping (RTIP) of day 7 and day 14 post-transplant peripheral blood (PB) samples as an effective and economically feasible tool to distinguish PGF versus delayed engraftment.Methods: Between Feb 2013 and Jun 2014, 26 patients underwent a myeloablative dCBT at our institute. Heparinized PB samples (30 ml) were obtained from patients on days 7 and 14 post-transplant. PB mononuclear cells (PBMCs) were isolated by density gradient separation and total cell number was counted. RTIP with 9-color flow cytometry was performed at each time point using isolated fresh PBMC.Results: Median time to engraftment was 19 days (range 12-51) in 23 evaluable patients. The remaining 3 patients had no documented hematopoietic recovery by routine daily CBCs. Two patients were declared PGFs when day 21 PB chimerism results demonstrated 100% host T cells followed by confirmation of 0% donor engraftment on day 28. The third patient never had sufficient quantity of circulating cells to obtain RTIP results and died on day 28 with no hematopoietic recovery. Excluding this patient, we were able to quantify the absolute number of PBMC at day 7 (median: 3.1/µl; 95% CI: 2.6-4.6) and day 14 (median: 26/µl; 95% CI: 33-103) in all other patients. There was an increase in the absolute number of PBMC from day 7 to 14 in all patients except the two who experienced PGF. Furthermore, RTIP of day 7 PBMC revealed a predominance of T cells that were donor-derived, while day 14 RTIP of PBMC demonstrated a decreased frequency of T cells and increased frequency of predominantly donor-derived monocytes and NK cells. In contrast to the engrafting patients, the two PGF patients displayed a markedly different pattern in RTIP with minimal evidence of circulating monocytes in the day 14 samples (Fig 1A). Importantly, RTIP demonstrated that day 7 PBMC contained a higher frequency of CD14+CD16- monocytes and CD56brightCD16- NK cells than were infused, suggesting these specific cells were generated de novo and were not representative of cells infused with the graft (data not shown).Correlations between day 7 or day 14 cell subset numbers and time to engraftment were analyzed. The median absolute number of monocytes at day 7 was 0.075/µl (95% CI: 0.03-0.153). Patients with day 7 monocyte counts above the median demonstrated earlier engraftment than patients below the median (17.5 vs 26.5 days; p= 0.011). Using linear regression with engraftment as the variable of interest and the absolute number of day 7 monocytes as the predictor, the coefficient was 0.6 (95% CI: 0.075- 0.78, P=0.025) (Fig 1B). As expected, higher day 14 monocytes also correlated with earlier engraftment. With respect to the NK cell subset, the absolute number of NK cells at day 7 was not significantly correlated with engraftment time, but day 14 NK cell numbers were predictive of engraftment kinetics. The median number of NK cells at day 14 was 2.78/µl (95% CI: 1.87-5.79). Patients with day 14 NK counts above the median had earlier engraftment than those below the median (16 vs 26.5 days; (P=0.0034). The regression coefficient was 0.5 (95% CI, 0.077- 0.77; P=0.024). Finally, although total T cell numbers at day 7 and day 14 had no correlation with engraftment time, the two evaluable PGF patients had a greater inversion of CD4:CD8 ratio (0.038 & 0.058) than others (median: 0.71, 95% CI: 0.22-1.42) at day 14.Conclusions: This study provides the first clear evidence that RTIP of PBMC at days 7 and 14 can detect the kinetics of circulating de novo generated monocytes and NK cells and also reflects the in vivo immune environment in patients following dCBT. Importantly, detecting and measuring specific cell subsets using RTIP permits earlier identification of patients at high risk of graft failure versus patients with delayed engraftment and warrants further development as a clinically feasible diagnostic method to guide clinical intervention. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Relationships Among Commonly Used Measures of Cord Blood Potency, ALDHbr Cell Content, and Colony Forming Cell Content in Cord Blood Units Prior to Cryopreservation: Towards An Improved Metric for Potency of Banked Cord Blood,

Abstract 4054 Background:Despite adequate total nucleated cell (TNC) dosing, graft failure and engraftment delays remain a significant issue for patients undergoing cord blood transplantation (CBT). Previously, we have shown that recovery of colony forming units (CFU) after cord blood unit (CBU) thaw is superior to TNC and CD34+ content in predicting engraftment. However, the CFU assay requires weeks and is not standardized. Aldehyde dehydrogenase (ALDH), an enzyme expressed in stem and progenitor cells (ALDHbr cells), is detected by a functional, flow-based intracellular assay and can be completed within a few hours. In this study, we determined the statistical relationships among TNC, CD34+, ALDHbr cells and CFU in a large cohort of fresh CBUs prior to cryopreservation with the ultimate goal of defining the best parameter(s) to predict potency defined by successful engraftment. Methods: As part of routine banking procedures, post-processing TNC, mononuclear cell count (MNC), CD34+cells and CFU content were enumerated on fresh CBUs donated to the Carolina Cord Blood Bank (n=5268) or identified for directed donation (n=14) between 9/07–7/09. ALDHbr cell content was measured using Aldecount (Aldagen, Inc.). Correlations among these various CBU characteristics were determined. Results: Of the 5282 fresh CBUs analyzed, the median values for TNC, CD34+, MNC, ALDHbr cells and CFU content were: 11.8×108 (range, 2.9–55.5×108), 3.36 x106 (range, 0.17–98.2×106), 5.5×108(range, 1.7–20.0×108), 4.3x 106 (0–36.6×106) and 34.0×105 (range, 0.6–193.3×105), respectively. There were significant correlations (p<0.0001) among all of the cord blood (CB) parameters measured as presented in Table 1. In standardized, univariate modeling which allowed for ranked comparisons of the CB parameters using AIC values, CFU was best predicted by TNC (AIC=11,501) followed closely by ALDHbr (AIC=12,012) and, to a lesser degree, MNC (AIC=12,412) and CD34+ (AIC=12,612). In multivariate modeling of CB parameters (Table 2), results were similar to the standardized univariate analysis.Table 1:Correlation coefficients for parameters measured on fresh CB (all p<0.0001)TNCCFUMNCALDHbrCD34+TNC1.000.700.820.580.50CFU0.701.000.610.660.60MNC0.820.611.000.470.41ALDHbr0.580.660.471.000.70CD34+0.500.600.410.701.00Table 2:Standardized multivariate models of CBU parametersModels - Slope Estimate (p value)PredictorTNCCFUALDHbrCD34+TNC–0.35*0.23*NSCFU0.22*–0.26*0.23*MNC0.56*0.12*−0.05 (p=0.002)NSALDHbr0.15*0.26*–0.54*CD34+NS0.19*0.46*–Volume0.09*NS−0.03 (p=0.01)0.03 (p=0.01)*p<0.0001; NS - not significant Conclusions:In this large dataset of fresh CBUs, the parameters measured (TNC, MNC, CFU, CD34+ and ALDHbr) are well correlated and predictive of each other. Furthermore, total ALDHbr measured on fresh CB was well correlated with other CB parameters, including CFUs that also assesses viable cells and is the unit characteristic that best predicts engraftment. Given the technical difficulties associated with measuring CFUs, ALDHbr content has promise as a potential CFU surrogate as a potency assay for CBUs. ALDHbr measurements were modestly, but significantly more reliable in predicting CFU content than CD34+ or MNC content in this cohort of fresh CBU. We and others have previously shown that different types of cells survive freezing and thawing of the CBU with different efficiencies. Consequently, we are studying which CBU parameter best correlates with engraftment after thawing of CBUs or from segments attached to CBUs in order to improve assessment of potency during the process of CBU selection for CBT. Disclosures:Balber:Aldagen, Inc: Consultancy, Equity Ownership.

Comparative analyses of megakaryocytes derived from cord blood and bone marrow

Thrombocytopenia is typically observed in patients undergoing cord blood transplantation. We hypothesized that delayed recovery of the platelet count might be caused by defects in the megakaryocytic differentiation pathway of cord blood progenitors. To test this hypothesis, we compared the features of in vitro megakaryocytopoiesis between cord blood progenitors and those in bone marrow cells after isolation of CD34+ cells as progenitors. The proliferative responses of the progenitors in cord blood are higher than those in bone marrow cells in the presence of interleukin (IL)-3, stem cell factor (SCF) and thrombopoietin (TPO). However, the ability to generate mature megakaryocytes was higher in bone marrow progenitors than in cord blood in the same in vitro culture system, when examined by the expression of CD41, polyploidy and proplatelet formation. Furthermore, an earlier induction of c-mpl protein, a receptor for TPO, was observed in the progenitors from bone marrow than in those from cord blood in the presence of SCF and IL-3. Therefore, the ability to generate mature megakaryocytes in bone marrow progenitors is superior to that in cord blood, and the delayed engraftment of platelets after cord blood transplantation might be attributed to the features of cord blood megakaryocyte progenitors.