Large Volume Leukapheresis Research Articles

Collection efficiency and safety of large-volume leukapheresis for the manufacturing of tisagenlecleucel.

In patients with relapsed or refractory B cell acute lymphoblastic leukemia or B cell non-Hodgkin lymphoma (r/r B-ALL/B-NHL) with low CD3+ cells in the peripheral blood (PB), sufficient CD3+ cell yield in a single day may not be obtained with normal-volume leukapheresis (NVL). Large-volume leukapheresis (LVL) refers to the processing of more than three times the total blood volume (TBV) in a single session for PB apheresis; however, the efficiency and safety of LVL for manufacturing of tisagenlecleucel (tisa-cel) remain unclear. This study aimed to investigate the tolerability of LVL. We retrospectively collected data on LVL (≥3-fold TBV) and NVL (<3-fold TBV) performed for patients with r/r B-ALL/B-NHL in our institution during November 2019 and September 2023. All procedures were performed using a continuous mononuclear cell collection (cMNC) protocol with the Spectra Optia. Although pre-apheresis CD3+ cells in the PB were significantly lower in LVL procedures (900 vs. 348/μL, p < .01), all patients could obtain sufficient CD3+ cell yield in a single day with a comparably successful rate of final products (including out-of-specification) between the two groups (97.2% vs. 100.0%, p = 1.00). The incidence and severity of citrate toxicity (no patients with grade ≥ 3) during procedures was not significantly different between the two groups (22.2% vs. 26.1%, p = .43) and no patient discontinued leukapheresis due to any complications. LVL procedures using Spectra Optia cMNC protocol was well tolerated and did not affect the manufacturing of tisa-cel.

Vascular access for autologous peripheral blood stem cells collection by large volume leukapheresis: Single center experience.

Peripheral blood stem cell (PBSC) harvesting requires reliable and safe vascular access. In our institution, a change of practice was implemented and the central venous catheter (CVC) placement for all autologous PBSC collections was abandoned in favor of a careful evaluation of peripheral venous access (PVA) for each individual patient. The aim of this prospective study was to evaluate the rate of patients with adequate peripheral veins for autologous PBSC collection and compare patient characteristics, collection efficacy, and complication rate between patients with PVA and CVC. Peripheral veins were assessed by the apheresis nurse team in all patients referred between January 2020 and July 2021 to autologous PBSC collection. Only in case of difficult venous access, CVC was inserted. Large volume leukapheresis (LVL) procedures, which processed ≥3 total blood volumes, were performed. In 65 (57%) patients PVA was used, while 49 (43%) patients required placement of short-term CVC. Peripheral venous access was successfully used significantly more often in males (69.8%) (P = 0.010), and patients with multiple myeloma (71.0%) than in patients with non-Hodgkin's lymphoma (35.9%) and Hodgkin's lymphoma patients (33.3%) (P < 0.001). There was a significant difference in the type of prior administered chemotherapy; in the patients who received cytostatics free chemotherapy, PVA was used more often (75.0%) (P = 0.007). In terms of the efficacy and safety of LVLs, there were no differences between procedures performed using PVA and CVCs. Peripheral venous access is feasible for autologous PBSC collection in more than a half of patients, in particular in those with multiple myeloma. Changes in the treatment of multiple myeloma, using new proteasome inhibitors-based and immunomodulatory agents that do not adversely affect peripheral veins, have enabled the use of PVA even at the high blood flow rates required by LVL. Peripheral venous access is not associated with safety issues or with a lesser collection efficiency, and it is cost-effective as well. Each patient referred to autologous PBSC collection needs to be evaluated individually by the experienced apheresis team for the most appropriate venous access.

Evaluating the efficiency and safety of large-volume leukapheresis using the Spectra Optia continuous mononuclear cell collection protocol for peripheral blood stem cell collection from healthy donors: A retrospective study.

Large-volume leukapheresis (LVL) refers to processing of more than three volumes of blood in a single session for peripheral blood stem cell collection. Recently, continuous mononuclear cell collection (cMNC) protocol has been developed using the Spectra Optia system, which is a widely used apheresis device. LVL using the novel protocol has been investigated in patients. However, the efficiency and safety of LVL in healthy donors using this protocol has not been characterized. Therefore, this study aimed to evaluate the efficiency and tolerability of CD34+ collection of LVL with the cMNC protocol in healthy donors. We retrospectively collected data on LVL (>3 total blood volume) and normal-volume leukapheresis (NVL) performed in healthy donors between October 2019 and December 2021. All procedures were performed using the cMNC protocol. Although pre-apheresis CD34+ cell count was lesser in LVL (23.5 vs. 58.0/μL, p < .001), CD34+ collection efficiency was comparable between LVL and NVL (61.2% vs. 61.4%, p = .966). Platelet loss was significantly higher in LVL compared to NVL (38.0% vs. 29.4%, p < .001), with no correlation between attrition of platelet and processing blood volume. Moreover, the incidence of citrate toxicity during procedures was comparable between the two groups (31.6% vs. 21.4%, p = .322). All LVL procedures could be completed without any adverse events. Allogeneic LVL procedure using Spectra Optia cMNC protocol was well tolerated by the donors and resulted in efficient collection of CD34+ cells, which was comparable to that of NVL.

Easy 250 EasySep&amp;[trade] Magnet: A Novel Magnetic Platform For Large-Volume CD34+ Hematopoietic Stem and Progenitor Cell Isolation from Leukapheresis Products

Abstract We have developed a novel magnetic platform for large-volume CD34+ hematopoietic stem and progenitor cell (HPSC) isolation from both mobilized and non-mobilized leukapheresis products. As the frequency of CD34+ cells is low, and cell quality and viability can be variable between samples, it takes great effort to isolate these cells with consistently high purities and yields. Working with large volume leukapheresis products can be challenging as the sample often needs to be split and processed in parallel, which can be both time-consuming and labor-intensive. The Easy 250 EasySep™ Magnet simplifies cell isolation procedures when processing leukapheresis samples. It is based on column-free immunomagnetic EasySep™ cell isolation technology and can process sample volumes up to 225 mL. We have developed a negative selection protocol for the enrichment of CD34+ HPSCs (mean purity ± SD of 74.6 ± 10.5%; n = 8) from non-mobilized leukapheresis products. For mobilized leukapheresis products, an EasySep™ positive selection protocol has been optimized for the isolation of highly purified CD34+ HPSCs (mean purity ± SD of 92.0 ± 1.7%; n = 7). Notably, these protocols can be completed in as little as 25 minutes, making the Easy 250 EasySep™ Magnet the fastest and simplest method for enriching highly purified HPSCs from large-volume leukapheresis products—critical for addressing the growing clinical demands for CD34+ cell-based therapies.

The effects of large-volume leukapheresis on hemoglobin, platelet and coagulation in patients with hyperleukocytic leukemia.

This study retrospectively analyzed a total of 86 leukapheresis treatments in 55 patients with hyperleukocytic leukemia (HLL). In the leukapheresis treatments, the monitoring collection (MNC) program in COBE spectra continuous flow centrifuge (CFC) for blood component separator was used. In this study, the white blood cell (WBC) suspension volume collected in leukapheresis treatment were 870.72±208.40 mL, and significantly larger than that reported in previous study. Compared with before leukapheresis, there were no difference in patient with HLL on the peripheral blood platelet (PLT) count and hemoglobin (HGB) levels. The index plasma fibrinogen (FIB) concentration was slightly reduced by leukapheresis, however, it did not affect a lot the maintenance of normal hemostatic function in patients with HLL. Our data provided evidences that large-volume leukapheresis had no obvious effects on HGB level and coagulation functions in patient with HLL. So large-volume leukapheresis by CFC could be widely used in clinic.

Impact of mobilization regimen on multiday collection of peripheral blood CD34+ cells by large volume leukapheresis.

The collection yield of hematopoietic progenitors cell (HPC) by leukapheresis is critical for a successful transplantation, which often requires multiday collections to achieve the collection goal. Collection procedures of 181 patients who underwent leukapheresis for more than 1 day were reviewed. Patients were separated into six groups based on the mobilization regimen: G-CSF on day 1 (D1) and day 2 (D2) (G-G); G-CSF on D1 and G-CSF and plerixafor on D2 (G-GP); G-CSF and plerixafor on day D1 and D2 (GP-GP); G-CSF and plerixafor on D1 and G-CSF on D2 (GP-G); chemotherapy and G-CSF on D1 and D2 (GC-GC); and chemotherapy, G-CSF, and plerixafor on D1 and D2 (GCP-GCP). Patient's pre-collection CD34 count (pre-CD34) on D1 and D2 were compared in the same individual and among groups. We found D2 pre-CD34 were significantly decreased in G-G, GP-G, and GP-GP groups and significantly increased in G-GP group (p < .001) using a repeated measures ANOVA analysis. D2 pre-CD34 remained at similar levels as D1 in GC-GC and GCP-GCP groups. A multiple regression analysis showed that the mobilization regimen was the only factor that significantly affected pre-CD34 D2/D1 ratio (p < .001). There was a significant difference in the pre-CD34 D2/D1 ratio (p < .001) among these six groups with the lowest in GP-G group (0.40 ± 0.45), and the highest in G-GP group (2.35 ± 0.36). Mobilization regimen has significant impact on pre-collection CD34 count. Apheresis facilities may change mobilizing drugs accordingly to achieve a specific HPC goal.

Mobilized and apheresis-collected endothelial progenitor cells with plerixafor.

Endothelial progenitor cells (EPCs) are immature cells able to proliferate and contribute to endothelial repair, vascular homeostasis, neovascularization, and angiogenesis. It therefore seems likely that circulating EPCs have therapeutic potential in ischemic and vascular diseases. In this study we evaluated the efficiency of EPC mobilization and collection by large volume leukapheresis in subjects with hematological diseases, treated with plerixafor in association with G-CSF. Twenty-two patients with lymphoid malignancies underwent rHuG-CSF and plerixafor treatment followed by leukapheresis. Blood samples before and after treatment and apheresis liquid sample were taken and analyzed by flow cytometry in order to quantified EPC. The percentage of CD34+ cells and EPCs among circulating total nuclear cells (TNCs) increased significantly by approximately 2-fold and 3-fold, respectively, after plerixafor treatment. Consequently, the absolute number of CD34+ cells and EPCs were increased 4-fold after plerixafor treatment. The median PB concentration of EPCs before and after treatment were 0.77/μL (0.31-2.15) and 3.41/μL (1.78-4.54), respectively, P< .0001. The total EPCs collected per patient were 3.3×107 (0.8×107 -6.8×107 ). We have shown that plerixafor in combination with G-CSF allows the mobilization and collection of large amounts of EPCs along with CD34+ cells in lymphoid neoplasm patients. The possibility to collect and to store these cells could represent a promising therapeutic tool for the treatment of ischemic complications without the need of in vitro expansion.

A Randomized Study Comparing the Effects of G-CSF and G-CSF/GM-CSF for the Mobilization of Peripheral Blood Stem Cells by Mitoxantrone and High-Dose Cytarabine Chemotherapy

We investigated the efficiency of mitoxantrone (MIT) and high-dose cytarabine (Ara-C) chemotherapy followed by G-CSF and G-CSF/GM-CSF treatments for the mobilization of peripheral blood stem cells (PBSCs) in patients with leukemia and lymphoma. MIT was intravenously injected at 10 mg/(m2·d) for 2 to 3 days, followed by Ara-C injected intravenously at 2 g/m2 every 12 hours for 1 to 2 days. When white blood cell count recovered from the lowest value, 5 to 7.5 μg/ (kg·d) G-CSF was administered in 23 patients for 5 to 7 successive days. Another 27 patients received 3-5 μg/ (kg·d) G-CSF and 3-5μg/ (kg·d) GM-CSF. Autologous peripheral blood mononuclear cells were collected. Levels of CFU-GM and CD34+ cells were determined after unfreezing. The CD34+ cells and CFU-GM yields of 27 patients in G-CSF plus GM-CSF combination group [(8.79±3.11)×106/kg, (3.52±1.34)×105/kg, respectively] were significantly higher than those of patients receiving G-CSF alone (n=23) [(6.14±2.06)×106/kg, (2.03±1.06)×105/kg, respectively (P &lt; 0.05)]. No obvious changes of T lymphocyte subsets in patients were observed when using G-CSF/GM-CSF, but levels of CD34+ cells increased gradually (P&gt;0.05). The end-point separation blood volume was all above trebling TBV. No severe complications were observed during the mobilization and collection. Autologous PBSCT obtained quick hematopoietic reconstitution. In conclusion, MA chemotherapy combined with G-CSF alone and G-CSF/GM-CSF can safely and effectively mobilize autologous PBSCs, while G-CSF plus GM-CSF is superior to G-CSF alone. Large volume leukapheresis is an important method to enhance the production rate of stem cells and decrease harvesting time.

Impact of Daratumumab-Containing Induction on Stem Cell Mobilization and Collection, Engraftment and Hospitalization Parameters Among Multiple Myeloma Patients Undergoing Autologous Stem Cell Transplantation

Introduction: Advances in induction regimens have significantly improved depth of response and duration of remission in multiple myeloma (MM) patients who are eligible for high-dose therapy and autologous stem cell transplantation (ASCT). Proteasome inhibitor-based induction regimens are standard as part of induction and it has been shown not to have any detrimental effect on stem cell (SC) collection and engraftment. Daratumumab (DARA) is an IgG1k monoclonal antibody directed against CD38 with potent antimyeloma activity. Based on the results of prospective studies DARA is now approved as part of induction therapy. Available data indicate a potential impact of DARA on SC collection but there is limited data on engraftment, duration of hospitalization and infection risk. In this retrospective analysis we evaluated the effect of DARA-based induction on ASCT parameters.Methods: The analysis included consecutive newly diagnosed MM patients that received ASCT between 2016 and 2020, as part of their upfront treatment regimen in our institution (Department of Clinical Therapeutics, Athens, Greece). Per institutional protocol, after 4-6 cycles of induction, pts received low dose cyclophosphamide (2.5 g/m2) followed by G-CSF (10 mcg/Kg/day) to mobilize and collect SCs. Plerixafor was administered on-demand in case of poor mobilization and insufficient first day collection. Large volume leukapheresis was performed in pts with low CD34+ counts in order to increase CD34+ yield. Pts received G-CSF 480 μcg once daily from day +4 after SC reinfusion to ANC >1500/mm3. All pts received antiviral and antifungal but no anti-bacterial prophylaxis.Results: 200 eligible pts were included in the analysis; 40 (20%) pts received DARA as part of PI-based upfront treatment and 160 (80%) pts received PI-based upfront treatment without DARA. Baseline demographics (age, gender, performance status) and disease characteristics (ISS and R-ISS stage, cytopenias, eGFR, lytic bone disease etc) were not different between the two groups. Response after induction was also similar (CR+VGPR rate was 93% vs 95% for non-DARA and DARA-containing regimens respectively).Use of DARA at induction was associated with lower total mean number of collected CD 34+ SCs (10.48 x 10^6/kg vs 16.58 x 10^6/kg, p<0.0001), or SC collection on day 1 (7.99 x 10^6/kg vs 16.27 x 10^6/kg, p<0.0001). Fewer pts in the DARA-treated group achieved the planned yield of at least 5 X 10^6 CD34+/kg, compared to DARA-untreated group (87.5 % vs 96.2%, p=0.047). DARA-treated pts required more often additional SC mobilization with on demand administration of plerixafor (42.5% vs 7.6%, p<0.0001). In order to compensate for a poorer mobilization and lower quality graft (CD34% 0.66% vs 1.26% in apheresis product, p<0.0001) DARA-treated group underwent more often >1 day of SC collection (37.5% vs 6.3%, P <0.0001), resulting in longer duration of collections (689 vs 452 min, p<0.0001) and larger total apheresis volumes (723 vs 557 ml, p<0.0001). However, 97% and 98% of pts in the two groups respectively were able to move to at least a single ASCT. Following ASCT, DARA-treated pts had a slightly delayed hematopoietic recovery (11 vs 10 days to PMN>500/mm3, p<0.001 and 12 vs 11 days for PLT counts> 25x10^9/mm3, p<0.001) and required more transfusions (2 vs 1 for RBCs, p=0.031 and 4 vs 2 for platelets, p<0.001). Rates of neutropenic fever were higher (80% vs 67%, p=0.182), required antibiotics for longer duration (10 vs 8 days, p=0.042) and more often 2 or more lines of antibiotic therapy (53% vs 39%, p=0.003), experienced more often septic shock (12.5% vs 1.3%, p=0.003) and as a results DARA-treated pts had a slightly prolonged hospitalization (21 vs 19 days, p=0.02). However, D100 mortality was not statistically different (<2% in both groups).Conclusion: DARA-containing induction before ASCT is associated with poorer mobilization and frequent need for use of plerixafor. However, similar percentage of patients can move to at least a single ASCT. The use of DARA-containing induction was also associated with slightly increased risk of infectious complications, antibiotics use and blood product transfusions but no increase in the risk of D100 mortality. These data point to the need for certain modifications to ASCT protocol for patients treated with DARA-containing regimens at induction, such as preemptive use of plerixafor, and perhaps prophylactic antibiotics. DisclosuresTerpos: Amgen: Consultancy, Honoraria, Research Funding; BMS: Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Genesis: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Novartis: Honoraria; Takeda: Consultancy, Honoraria, Research Funding; Sanofi: Consultancy, Honoraria, Research Funding; GSK: Honoraria, Research Funding. Gavriatopoulou: Takeda: Honoraria; Karyopharm: Honoraria; Sanofi: Honoraria; GSK: Honoraria; Genesis: Honoraria; Janssen: Honoraria; Amgen: Honoraria. Dimopoulos: Amgen: Honoraria; BMS: Honoraria; Janssen: Honoraria; Takeda: Honoraria; BeiGene: Honoraria. Kastritis: Takeda: Honoraria; Pfizer: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Genesis Pharma: Honoraria; Janssen: Consultancy, Honoraria, Research Funding.

Successful autologous peripheral blood stem cell collection using large volume leukapheresis in patients with very low or undetectable peripheral blood CD34+ progenitor cells

Autologous stem cell transplantation provides some patients with hematolymphoid and solid organ malignancies an opportunity for cure. Management of peripheral hematopoietic stem cell (HSC) collections differs among institutions, especially if a very low pre-procedure peripheral blood CD34+ cell count (PBCD34) is demonstrated. This study retrospectively analyzed results of large-volume peripheral HSC collections in 91 patients over approximately two years. Fifteen patients with PBCD34 < 10 × 10e6/l (eleven with undetectable PBCD34) were compared to 76 patients with higher counts on the first collection day (adequate mobilizers). The poor mobilizer group had significantly lower pre-collection WBC and platelet counts as well as collection yields. However, most patients with PBCD34 < 10 × 10e6/l (80 %) collected the minimum target for HSC transplant (2.0 × 10e6 CD34+ cells/kg) in <5 consecutive days of collection, and those who did collect the minimum successfully underwent autologous transplantation, with hematopoietic engraftment and long-term survival comparable to the adequate mobilizers. Successful HSC collection may often be achieved regardless of d 1 PBCD34 counts.

Mobilization of Auto-Stem Cells in the Thalassemia Major and Relationship with Iron Overload

Objectives: The hematopoietic stem cells transplantation (HSCT) is the only curable treatment in current for thalassemia major (TM).The quantity of auto stem cell of TM patients are very important for rescue the patients in case of failure of HSCT, and auto stem cells of TM will be used as target cells for gene therapy (GT) in the near future. Iron overload (IOL) can damage the hematopoiesis of TM. How to collect the auto stem cells in TM patients? How about the affects of iron overload in the mobilization and collection of auto stem cell? The aim of this study was to analyze quantity of auto stem cells in the TM and the affects of iron overload (IOL). Methods: We retrospectively analyzed a total of 134 patients and 25 normal donors undergoing PBSCs collection between January 2012 and December 2019. Patients with serum ferritin levels over 1,000 ng/mL and with a history of red blood cell transfusions prior to stem cell collection were defined as a group having transfusion-associated iron overload (IOL). In total one hundred and thirty-four patients and twenty-five donors were subcutaneous administrated G-CSF (Granulocyte Colony Stimulating Factor) 10 mg/kg/d Injection for 5 days, PBSCs was collected using a large volume leukapheresis (LVL) procedure. The data of WBC, MNC and CD34+ in product were analyzed by SPSS 20 software. Results: In 134 patients, median age is 8.60 range: (1-17),and 25 Normal donors, median age is 7.5 range: (1-18). The results demonstrated that cells either in patients with thalassemia or normal donors were effectively mobilized by G-CSF (10 mg / kg/d),as these children showed a marked increase in white blood cells and MNC cells in peripheral blood, reaching a peak in 4 to 5 days after the injection of G- CSF alone. Comparison of the MNC [(6.7±3.5) ×10⁸/kg vs (7.2±4.77) ×10⁸/kg] and CD34+cells [(10.29±4.5)×10⁶/kg vs (10.9±6.8)×10⁶/kg] in collected product in TM patients and normal donors groups revealed no significant difference. There was no significant difference between different Ferritin level (Mild, Intermediate and Severe) on WBC, MNC, and CD34+ in each group at the time point studied. But there was significant difference on MNC number (Normal liver iron (7.73±2.69)×10⁸/kg, Mild liver iron overload (7.66±4.64)×10⁸/kg, Intermediate (6.18± 2.84) ×10⁸/kg compared to Severe liver iron (4.45±3.34)×10⁸/kg, P=0.039. Significant difference was also can be seen in number of MNC of product in patients with intermediate cardiac iron overload compared to normal cardiac iron overload and mild cardiac iron overload (2.76±0.97) ×10⁶/kg vs(6.86±3.39) ×10⁶/kg and (6.88±3.72)×10⁶/kg, P=0.030. There was a statistically significant difference positive correlation between median age (P=0.011) and MNC/Kg (P=0.030). Conclusion:The quantity of mobilization of PBSC in TM patients had no significant difference compare to normal donor. The MNC in product was statistically decreased in intermediate cardiac iron overload and severe liver iron overload (IOL) subgroup. It indicated that iron overload (IOL) in organs may negatively relate to proliferation and mobilization of stem cells in TM. Further quality study of PBSC such as stem cell culture and ability of proliferation should be more evaluated. The sufficient quantity and high quality auto stem cell from TM patient can be more used in the future. Disclosures No relevant conflicts of interest to declare.

Prospective identification of potential factors influencing stem cell mobilization and the necessity for plerixafor use in newly diagnosed multiple myeloma patients undergoing autologous stem cell transplantation

IntroductionTo study the efficacy and safety of single large volume leukapheresis by using generic G-CSF or G-CSF plus Plerixafor in achieving adequate stem cell yield and various factors influencing thereof in newly diagnosed multiple myeloma patients undergoing autologous stem cell transplant . MethodThis prospective study was undertaken among 55 newly diagnosed multiple myeloma patients undergoing autologous stem cell transplant and aged between 18 and 75 years. Mobilization and harvesting of stem cells were performed by using GCSF or GCSF plus Plerixafor and large volume leukapheresis, respectively. A stem cell yield of ≥2×106kg–1 and the number of apheresis procedures were primary efficacy endpoints, while the ideal stem cells yield >5×106kg–1, the engraftment day and D100 response/graft sustainability were secondary endpoints. ResultThe primary endpoint was achieved in all cases in both the groups by using a single LVL leukapheresis procedure. Fulfillment of all the secondary endpoints was satisfactory and comparable in both the groups. Age, pre-apheresis CD34+ count and number of interruptions during the LVL were significant factors influencing the stem cell yield (p<0.05). Adverse drug reactions during the apheresis and post-ASCT period were manageable. ConclusionThe LVL is safe and cost-effective in attaining a minimum of CD34+ cells in a single procedure with manageable adverse reactions. Judicious intervention during the procedure may be helpful in ensuring the adequate yield.

Mobilization kinetics of CD34+ hematopoietic stem cells stimulated by G-CSF and cyclophosphamide in patients with multiple sclerosis who receive an autotransplant

Background aimsAutologous hematopoietic stem cell transplantation (AHSCT) is an alternative for multiple sclerosis (MS) patients who do not respond to conventional treatment. Mobilization kinetics of CD34+ cells in MS patients has not been studied. MethodsPatients with MS mobilized with granulocyte colony-stimulating factor (G-CSF) and cyclophosphamide (Cy) were prospectively studied. Three counts of CD34+ cells were done in peripheral blood: at baseline before mobilization, at the start, and immediately at the end of apheresis. Complete blood counts were performed at the times of CD34+ cell counting. Standard statistical descriptive analysis of MS patients’ salient features was performed, and after log 10 transformation of the data, Pearson test was performed to assess correlation between variables and CD34+ cell count. In addition, multiple linear regression of relevant data was carried out for multivariate analysis. ResultsData of 51 consecutive MS patients with median age of 48 (31–64) years were analyzed. The CD34+ cell count increased 26-fold after mobilization. During large volume leukapheresis (LVL), the number of CD34+ cells in peripheral blood increased from 51.29 CD34+/μL at the start to 62.3 CD34+/μL at the end. A negative correlation between CD34+ cell count after leukapheresis and age (r = –0.32, P = 0.02) was observed. Neither the CD34+ baseline count nor sex correlated with the CD34+ count in peripheral blood immediately at the end of apheresis. ConclusionsMobilization with G-CSF and Cy in MS patients resulted in effective CD34+ hematoprogenitors release from the bone marrow and in intra-apheresis recruitment.

Coagulopathy, Hypoxemia, and Mortality Outcomes in Newly Diagnosed Acute Myeloid Leukemia with Hyperleukocytosis Treated with Large Volume Leukapheresis

Background: Patients with acute myeloid leukemia (AML) presenting with hyperleukocytosis have frequent complications and early mortality. Pulmonary, central nervous system, and cardiovascular complications are common. Attempts to improve outcomes in the 10% of AML patients arriving with hyperleukocytosis have included leukapheresis. No prospective randomized study has supported the use of leukapheresis, and retrospective reports have revealed persistently poor outcomes as well as emerging concerns of acquired coagulopathy and worsening hypoxemia after leukapheresis. While many centers use a leukapheresis protocol processing two blood volumes, Johns Hopkins protocol routinely processes three-five blood volumes. This study aimed to assess coagulopathy, hypoxemia, and mortality with large volume leukapheresis. Methods: 32 patients with newly diagnosed AML treated with large volume leukapheresis for WBC depletion are included in this report. Demographic, clinical, laboratory, and apheresis-related data were collected. Coagulopathy and hypoxemia-related metrics were evaluated within 6 hours before leukapheresis and within 6 hours of completion of leukapheresis. Descriptive and inferential statistics (chi square and Mann-Whitney U test) were used to compare pre and post-leukapheresis findings and assess clinical outcomes. Results: Twenty-nine of 32 (93.8%) patients presented with symptomatic leukostasis (with pulmonary and/or CNS symptoms in 26/29). Median blood volume processed was 14.8 liters (range 4-23.4L). Mean platelet count decreased from 60x109/L to 37x109/L after leukapheresis (p&lt;0.001). Comparing supplemental oxygen requirements before and after leukapheresis, 1 patient had decreased requirements, 6 increased, and 25 were unchanged. Median change in prothrombin time (PT) was an increase of 1 second (range: -2.4 to +7.7 seconds; p=0.13). Twenty-six of 26 evaluable patients had a decline in fibrinogen post-pheresis, with median reduction of 84 mg/dL (range: -12 to -483 mg/dL; p=0.04). Twelve of 18 (66.7%) patients 65 years and older died by day 30, compared with 3/14 &lt;65 years-old (p=0.01). Fifteen of 32 (46.9%) arrived with acute renal failure on admission, 10 of whom died by day 30. All 8 patients requiring hemodialysis during their initial admission died prior to day 30. 12/32 (37.5%) required mechanical ventilation, 10 of whom died by day 30. Overall, 10/32 (31.3%) died by day 7, and 15/32 (46.9%) died by day 30. The most common primary cause of death was multiorgan failure including both renal failure and hypoxemia in 8 patients. Two patients died of confirmed intracranial hemorrhage, and 2 died with clinical suspicion for intracranial hemorrhage but were too unstable for imaging prior to death. Two deaths were attributed to ischemic stroke, and 1 patient died with isolated refractory hypoxemia. Conclusions: Patients with AML presenting with hyperleukocytosis have a very high mortality, particularly when complicated by symptomatic leukostasis. Similar to Van de Louw's report, we observed worsening coagulopathy and a subgroup with increased oxygen requirements after leukapheresis. While our sample size is too small to draw broad conclusions, we were not able to identify a group clearly benefiting from leukapheresis. We did not find evidence that larger volume leukapheresis decreased complications or mortality. These results should lead to caution when considering leukapheresis for patients with newly diagnosed AML, particularly in those presenting with a severe coagulopathy. Table Disclosures Webster: Pfizer: Consultancy; Amgen: Consultancy; Genentech: Research Funding. Gojo:Amphivena: Research Funding; Amgen Inc: Consultancy, Honoraria, Research Funding; Juno: Research Funding; Merck: Research Funding; Jazz: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria. Pratz:Boston Biomedical: Consultancy; Millenium/Takeda: Research Funding; Agios: Consultancy, Research Funding; Astellas Pharma: Consultancy, Research Funding; AbbVie: Consultancy, Research Funding. Smith:Celgene: Consultancy; Jazz: Consultancy; Pfizer: Consultancy; Agios: Consultancy; Novartis: Consultancy. DeZern:Astex Pharmaceuticals, Inc.: Consultancy; Celgene: Consultancy. Levis:Amgen: Consultancy, Honoraria; Agios: Consultancy, Honoraria; Astellas: Consultancy, Research Funding; FUJIFILM: Consultancy, Research Funding; Menarini: Consultancy, Honoraria; Novartis: Consultancy, Research Funding; Daiichi Sankyo Inc: Consultancy, Honoraria.

Clinical Application of Apheresis in Very Small Dogs Weighing &lt;8 kg to Pediatric Patients

Apheresis in low body weight children and adolescents is challenging due to a variety of technical and clinical issues including vascular access, low total blood volume, and hypotension. Although dogs have been a valuable preclinical model for apheresis, the procedure can be challenging since many pure-bred dogs are extremely small. Therefore, apheresis in these very small breeds presents very similar challenges as seen when performing the procedure in very low body weight people. We describe apheresis of four very small dogs, weighing from 4.6 to 7.6 kg, using either a COBESpectra and Spectra Optia apheresis system (Terumo BCT, Lakewood, CO, USA). Two dogs underwent large volume leukapheresis to collect mononuclear cells in preparation for hematopoietic stem cell transplantation and two dogs underwent therapeutic plasma exchange to treat an immune-mediated disease. In all cases, a dual-lumen hemodialysis catheter placed in the jugular vein provided adequate machine inlet and return flow rates. Machine priming was necessary to maintain hemodynamic stability during the beginning of the procedure, and rinseback was avoided for the same reason. Anticoagulant citrate dextrose solution, solution A was used for the large volume leukapheresis procedures and a combination of anticoagulant citrate dextrose solution, solution A and heparin was used for the therapeutic plasma exchange procedures. As such, serum iCa levels were regularly monitored and 10% calcium gluconate constant rate infusions were used to prevent citrate toxicity. All dogs completed the aphereses with no life-threatening adverse events. We conclude that aphereses in very small dogs is feasible if close attention is paid to hemodynamic stability and citrate toxicity.

Performance prediction algorithm for autologous PBSC collection in adults and pediatric patients using large volume leukapheresis.

The number of CD34+ cells collected in apheresis procedures depends mainly on the collection efficiency of the device and the blood volume processed. Large volume leukapheresis (LVL) can improve CD34+ cell yield and has previously been investigated using the COBE Spectra device (Terumo BCT, USA). This was a retrospective analysis of LVL performance in patients undergoing continuous mononuclear cell collection (CMNC) using the new Spectra Optia apheresis system (Terumo BCT, USA) at the University Hospital Center, Zagreb, from March 2016 to September 2016. CD34+ cell yield predictability, determined using a customized algorithm, was also assessed. In total, 67 procedures performed in 46 adults and 14 performed in 11 children were included in the analysis. In adults, 30 (65.2%) patients successfully reached their target preapheresis CD34+ cell count on day 1, with a median (interquartile range [IQR]) CD34+ collected cell dose of 4.8 × 106 /kg (2.3-10.6 × 106 /kg). In the pediatric group, 81.8% successfully collected the target CD34+ cell dose on the first day, with a median (IQR) CD34+ collected cell dose of 11.1 × 106 /kg (3.2-16.3 × 106 /kg). The customized algorithm showed a strong and significant linear correlation with actual CD34+ cell dose (P < 0.0001). The results of this study support the use of LVL and the customized prediction algorithm in apheresis procedures. The ability to tailor the procedure to meet the needs of the individual patient may help to minimize the blood volume processed, shorten the duration, reduce the volume of infused anticoagulants, and improve patient comfort.

Comparison and cost analysis of three protocols for mobilization and apheresis of haematopoietic progenitor cells.

Autologous bone marrow transplantation is a component of the malignant hemopathy therapy. The preferred mobilization and collection method is apheresis. The aim of this study is to compare three protocols analyzing the effect of plerixafor, higher dose of G-CSF and large volume leukapheresis (LVL). A retrospective cohort study including 119 patients referred for mobilization. Three protocols were compared: (a) G-CSF 10 μg/kg/day subcutaneous (sc) × 4 days mobilizing 1 to 1.5 blood volumes. (b) G-CSF 10 μg/kg/day sc × 4 days + plerixafor 0.24 mg/kg/day sc preventively or as a rescue agent mobilizing 1 to 1.5 blood volumes. (c) G-CSF 20 μg/kg/day sc × 4 days ± plerixafor 0.24 mg/kg/day sc preventively or as a rescue agent mobilizing 3 to 4 blood volumes. The average number of days of apheresis was reduced to 1.37 with protocol 3. The average cost per patient was reduced by 67% compared with protocol 2 and increased by only 5% compared with protocol 1, reducing the failure rate to 0%. Adding preemptive or rescue plerixafor (protocol 2) to G-CSF 10 μg/kg/day alone (protocol 1) did not improve the days of apheresis nor the number of CD34+ cells collected but had higher cost and failure rate. Using LVL, plerixafor and G-CSF 20 μg/kg/day (protocol 3) decreased the number of sessions to 1.37, reduced the failure rate to 0% and led to a significant increase in the number of CD34+ cells collected without toxicity and with a similar cost to protocol 1.

The impact of preapheresis white blood cell count on autologous peripheral blood stem cell collection efficiency and HSC infusion side effect rate.

Autologous peripheral blood hematopoietic stem cell (PBSC) collection efficiency (CE) is reportedly affected by the patient's blood properties; however, studies to identify factors correlated with CE have shown inconsistent results. Additionally, variables such as stem cell graft granulocyte content and patient age, sex, and underlying disease, may be associated with hematopietic stem cell (HSC) infusion-related adverse reactions. In this study, we evaluated the correlation of preleukapheresis PB granulocyte count and PBSC harvest variables with CD34+ collection yield and efficiency, and thawed HSC infusion side effect occurrence. We evaluated data from 361 patients who had undergone autologous PBSC transplant. Large volume leukapheresis was the method for PBSC collection. Complete Blood Count and CD34+ cell enumeration were performed in the preapheresis PB and the apheresis product sample. The PBSC grafts were submitted to non-controlled rate freezing after addition of 5% DMSO plus 6% hidroxyethylstarch as a cryoprotectant solution. The cryopreserved graft was thawed in a 37°C water bath and then infused without further manipulation. The CD34+ yield was associated with preapheresis PB CD34+ count and immature granulocyte count. The PBSC CE was negatively correlated with preapheresis white blood cell (WBC), immature granulocyte and granulocyte count. The leukapheresis product total nucleated cell (TNC) and granulocyte content was correlated with the thawed graft infusion side effect occurrence. This study has shown that preapheresis PB WBC and granulocyte counts were associated with leukapheresis CE. Additionally, the leukapheresis product TNC and granulocyte content was correlated with thawed graft infusion side effect occurrence.

Expression of adhesion molecules on CD34+ cells from steady-state bone marrow before and after mobilization and their association with the yield of CD34+ cells.

BackgroundCell adhesion molecules (CAMs) expressed on hematopoietic progenitor cells (HPCs), endothelial cells, and stromal cells play a pivotal role in the mobilization of CD34+ cells. Herein, we conducted a non-randomized peripheral blood stem cell (PBSC) mobilization study aimed to compare the potential differences in the expressions of several CAMs and chemokines on CD34+ cells obtained from bone marrow aspirate before and after HPC mobilization from patients with hematologic malignancies and healthy donors.MethodsThree-color cytofluorometric analysis was used to compare the expressions of CAMs and chemokines in the bone marrow before and after mobilization.ResultsFor all studied groups, CAM expression among those with good and poor yields of CD34+ cells was significantly correlated with VCAM-1 (P=0.007), CD44 (P=0.027), and VLA-4 (P=0.014) expressions. VCAM-1 (P=0.001), FLT-3 (P=0.001), CD44 (P=0.011), VLA-4 (P=0.001), and LFA-1 (P=0.001) expressions were higher before HPC mobilization than after HPC mobilization. By contrast, the expression of CXCR4 significantly varied before and after mobilization only among those with successful PBSC mobilization (P=0.002).ConclusionWe attempted to identify particular aspects of CAMs involved in CD34+ cell mobilization, which is a highly complex mechanism that involves adhesion molecules and matrix metalloproteases. The mechanism by which CD34+ cell mobilization is activated through proteolytic enzymes is not fully understood. We believe that CXCR4, VLA-4, CD44, and VCAM-1 are the most important molecules implicated in HPC mobilization, particularly because they show a correlation with the yield of CD34+ cells collected via large volume leukapheresis.

Factors influencing platelet clumping during peripheral blood hematopoietic stem cell collection.

Platelet clumping is a common occurrence during peripheral blood hematopoietic stem cell (HSC) collection using the Spectra Optia mononuclear cell (MNC) protocol. If clumping persists, it may prevent continuation of the collection and interfere with proper MNC separation. This study is the first to report the incidence of clumping, identify precollection factors associated with platelet clumping, and describe the degree to which platelet clumping interferes with HSC product yield. In total, 258 HSC collections performed on 116 patients using the Optia MNC protocol were reviewed. Collections utilized heparin in anticoagulant citrate dextrose to facilitate large-volume leukapheresis. Linear and logistic regression models were utilized to determine which precollection factors were predictive of platelet clumping and whether clumping was associated with product yield or collection efficiency. Platelet clumping was observed in 63% of collections. Multivariable analysis revealed that a lower white blood cell count was an independent predictor of clumping occurrence. Chemotherapy mobilization and a lower peripheral blood CD34+ cell count were predictors of the degree of clumping. Procedures with clumping had higher collection efficiency but lower blood volume processed on average, resulting in no difference in collection yields. Citrate toxicity did not correlate with clumping. Although platelet clumping is a common technical problem seen during HSC collection, the total CD34+ cell-collection yields were not affected by clumping. WBC count, mobilization approach, and peripheral blood CD34+ cell count can help predict clumping and potentially drive interventions to proactively manage clumping.