Sufficient Numbers Of Stem Cells Research Articles

Sysmex XN-HPC: study of reference intervals and clinical decision limits in healthy allogeneic donors mobilised with G-CSF

The Sysmex XN series haematopoietic progenitor cell (XN-HPC) is a novel tool for assessing stem cell yield before allogeneic haematopoietic stem cell transplantation. This study aimed to establish a reference interval (RI) for XN-HPC in peripheral blood allogeneic transplant donors following granulocyte colony-stimulating factor (G-CSF) stimulation and determine its clinical significance. All specimens were analysed using Sysmex XN-20. Samples were collected and analysed using non-parametric percentile methods to define the RIs. Quantile regression was used to explore the dependency of the RIs on sex and age. Samples were included in clinical decision limits for apheresis based on receiver operating characteristic curve analysis. The non-parametrically estimated RI for XN-HPC was 623.50 (90% confidence interval [CI90%] 510.00–657.00) to 4,144.28 (CI90% 3,761.00–4,547.00). The RIs for the XN-HPC were not age-dependent but were sex-dependent. The RI for males was 648.40 (CI90% 582.00–709.00)–4,502.60 (CI90% 4,046.00–5,219.00) and for females was 490.90 (CI90% 311.00–652.00)–3,096.90 (CI90% 2,749.00–3,782.00). Comparisons based on XN-HPC values between the poor and less-than-optimal groups, good and less-than-optimal groups, and good and non-good groups had areas under the curve of 0.794 (P < 0.001), 0.768 (P < 0.001), and 0.806 (P < 0.001), respectively, indicating a good predictive value for mobilisation effectiveness. XN-HPC data exceeding 3974 × 106/L suggested that a sufficient number of stem cells could be collected clinically. Values > 5318 < 106/L indicated 100% mobilisation effectiveness. We established an RI for XN-HPC in peripheral blood allogeneic transplant donors following G-CSF stimulation and determined clinical decision thresholds for mobilisation efficiency.

Cytarabine Plus Etoposideis+Peg-Rhg-CSF Is More Effective Than Cyclophosphamide + G-CSF As a Stem Cell Mobilization Regimen in Poorly Mobilized MM and Lymphoma

There are several methods for hematopoietic stem cell (HSC) mobilization in lymphoma and multiple myeloma (MM)，of which Cyclophosphamide(Cy) plus granulocyte-colony stimulating factor(G-CSF)is a commonly used HSC mobilization regimen,but this regimen often fails to collect a sufficient number of stem cells, especially for poorly mobilized patients.However,previous studies have shown that the mobilization effect of cytarabine plus etoposide and G-CSF seems to have a higher mobilization efficacy. The purpose of this study is to retrospectively compare the mobilization effects of using cytarabine ，etoposide plus polyethylene glycol recombinant human granulocyte stimulating factor(PEG-rhG-CSF)(EAP) and Cy+G-CSF in proven poor mobilizer or predicted poor mobilizer of MM or lymphoma. A total of 43 patients were mobilized by EAP regimen and the 44 patients by Cy+G-CSF, including 56 patients with MM and 31 patients with lymphoma.Both groups were comparable. The target yield of 2× 10 6 CD34 + cells/kg was achieved in 90.7% (EAP) and 70.7% (Cy) of patients(P=0.003) by 1.3 and 1.6 apheresis (means).By single apheresis in 76.7% (EAP) and 29.5% (Cy) of patients achieve the ideal yield of 5× 10 6 CD34 + cells/kg .EAP mobilization resulted in higher peak concentration of CD34+ cells in blood in the first apheresis (median 59.4 vs 9.5/µL, p＜0.001)and second apheresis(median 23.2 vs 8.5/µL, p=0.001)compared to Cy mobilization. The median time of antibiotic administration in EAP mobilization was shorter than that in Cy mobilization (median 1.2 vs 3.1/d, p=0.041). The toxicity is different,the median time of agranulocytosis (median 1.2 vs 3.1/d, p=0 .003) after EAP mobilization were shorter than those after Cy，however, the median time for thrombocytopenia gr.4 (median 0.5 vs 0.2/d, p=0.001) was longer in the EAP group. In view of these results, we conclude that mobilization with EAP is excellent mobilization regimen with acceptable toxicity and could be considered in MM and lymphoma patients with factors of poor mobilization.

Evaluation of CD34+ Cell Count at Different Time Points Following Plerixafor Administration in Autologous Hematopoietic Stem Cell Transplantation

Background: In apheresis, collecting an adequate number of CD34+ cells is required for successful autologous hematopoietic stem cell transplantation (auto-HSCT) procedure. It is difficult to harvest a sufficient number of stem cells in certain patients due to their old age and history of intensive chemotherapy. Plerixafor could mobilize stem cells and facilitate peripheral blood hematopoietic stem cell collection. However, not enough information is available on the appropriate time intervals from plerixafor administration to apheresis. Objectives: In this study, we aimed at evaluating the level of peripheral blood CD34+ cells at plerixafor administration time and every three hours to identify the peak time of circulating CD34+ cells. Methods: Circulating CD34+ cells were enumerated by flow cytometry on day 4 post mobilization. Plerixafor was administered to patients with poor mobilization based on the count of peripheral blood hematopoietic stem cells. The number of circulating CD34+ cells was evaluated before and 3, 6, 9, and 12 hours after plerixafor administration to assess the time it takes for stem cells to reach their peak level. Results: The highest level of stem cell concentration was found in 9 hours after plerixafor administration with an increasing trend. A statistically significant relationship was also observed between factors including platelet count on the first day of GCSF injection and the day of stem cell infusion, leukocyte count on admission, and basal levels of CD34+ cells in peripheral blood and the amount of harvested stem cells. Conclusions: We demonstrated that plerixafor causes an incremental trend in CD34+ cells mobilization, reaching its peak after 9 hours. Further research should be performed to provide insights into graft cells’ population and hematologic and immunological recovery.

Calcified nodule formation by dental pulp cells derived from rats after subcutaneous injection of an immunosuppressant

Introduction: The purpose of this study was to assess the significant proliferation of dental pulp-derived stem cells in vitro from rats with the systemic administration of immunosuppressant in subcutis. There must be a sufficient number of stem cells for tooth regeneration. However, number of mesenchymal stem cells in the dental pulp tissue is a small. Then, the proliferation of stem cells must be accelerated for hard tissue formation. The subcutaneous injection of the immunosuppressant would enhance the hard tissue forming ability of dental pulp cells of rat. It was hypothesized in this study that differentiation of stem cells into blasts would be effectively promoted by suppression of the systemic immune response. Materials and methods: The dental pulp cells of rats with immunosuppressant injection subcutaneously were cultured with or without addition of the immunosuppressant in the medium containing dexamethasone for calcified nodule formation. Ca2+ by decalcification of calcified nodules were quantitatively analysed. Statistical comparisons between the quantities of Ca2+ were performed using two-way unrepeated ANOVA followed by post hoc analysis with Tukey-Kramer’s test. Differences of p &lt; 0.01 were considered significant. Results: The proliferation and differentiation of stem cells among dental pulp cells was inhibited by the presence of immunosuppressive agents in the culture medium. However, stem cells obtained from rats after systemic administration of an immunosuppressive agent exhibited a high ability to form calcified nodules. Conclusions: To promote proliferation and differentiation of stem cells, systemic administration of an immunosuppressant to individuals prior to harvesting stem cells would be recommended.

Dystrophin Dp71ab is monoclonally expressed in human satellite cells and enhances proliferation of myoblast cells

Dystrophin Dp71 is the smallest isoform of the DMD gene, mutations in which cause Duchenne muscular dystrophy (DMD). Dp71 has also been shown to have roles in various cellular processes. Stem cell-based therapy may be effective in treating DMD, but the inability to generate a sufficient number of stem cells remains a significant obstacle. Although Dp71 is comprised of many variants, Dp71 in satellite cells has not yet been studied. Here, the full-length Dp71 consisting of 18 exons from exons G1 to 79 was amplified by reverse transcription-PCR from total RNA of human satellite cells. The amplified product showed deletion of both exons 71 and 78 in all sequenced clones, indicating monoclonal expression of Dp71ab. Western blotting of the satellite cell lysate showed a band corresponding to over-expressed Dp71ab. Transfection of a plasmid expressing Dp71ab into human myoblasts significantly enhanced cell proliferation when compared to the cells transfected with the mock plasmid. However, transfection of the Dp71 expression plasmid encoding all 18 exons did not enhance myoblast proliferation. These findings indicated that Dp71ab, but not Dp71, is a molecular enhancer of myoblast proliferation and that transfection with Dp71ab may generate a high yield of stem cells for DMD treatment.

Prevention of irradiation-induced damage to salivary glands by local delivery of adipose-derived stem cells via hyaluronic acid-based hydrogels

Most patients with head and neck cancer experience salivary gland dysfunction after chemo- and radiotherapies. There is currently no available treatment for this condition. Intervention with stem cell therapy has emerged as a promising approach. Although some progress has been made with systemic delivery of stem cells, it remains challenging to deliver a sufficient number of stem cells to the damaged tissue. Furthermore, local delivery of stem cells involves frequent cell loss. Herein, we evaluated the ability of adipose-derived stem cells (ASCs) with or without hydrogel to prevent salivary gland damage. Salivary gland cells were isolated from irradiated mouse submandibular glands. These cells exhibited higher expression levels of amylase, mucin, and aquaporin-5 when co-cultured with ASCs. Local delivery of ASCs into the salivary gland, using in situ-forming hyaluronic acid-based hydrogel (HA gel) as a carrier, revealed that ASCs remained at the injection site for a longer duration, compared with ASCs that were injected without HA gel. The salivary gland exhibited better function and morphology when ASCs were injected using an HA gel. In conclusion, retention of locally delivered ASCs by HA gels can enhance the paracrine effect of ASCs, thereby preventing irradiation-induced damage to the salivary gland and subsequent dysfunction.

Analysis of factors associated with successful allogeneic peripheral blood stem cell collection in healthy donors

BackgroundThe collection of a sufficient number of stem cells is important for success of allogeneic hematopoietic stem cell transplantation (HSCT). This study aimed to investigate the factors associated with successful allogeneic peripheral stem cell (PBSC) collection in healthy donors. MethodsWe retrospectively reviewed clinical data of allogeneic PBSC collection in 175 donors from 2007 to 2017 at the National Cancer Center, Korea. This study analyzed factors associated with the CD34+ cell yield such as the characteristics of donors, including age, laboratory results before apheresis, and data of procedures on the first day. The CD34+ cell dose of ≥ 4.0 × 106/kg have recently been the accepted minimum recommended dose in allogeneic HSCT settings, and this was the target dose in our study. ResultsThe factors associated with the CD34+ cell yield were age (p = 0.007), baseline platelet (PLT) (p = 0.014), and pre-collection hematopoietic progenitor cells (HPCs) (p = 0.001) by multivariate analysis. This study represented that age, baseline platelet count, and pre-collection HPC count are important predictive factors as shown in other previous studies. ConclusionOur data suggest that young age, high baseline platelet counts and high HPC counts before collection might be useful for identifying successful mobilizers.

Results of a Prospective, Multicenter, Phase I/II Clinical Study in Pediatric and Adult Patients Using TCR Alpha/Beta and CD19 Depleted Haploidentical Hematopoietic Stem Cell Grafts Following Reduced-Intensity Conditioning

Results of a Prospective, Multicenter, Phase I/II Clinical Study in Pediatric and Adult Patients Using TCR Alpha/Beta and CD19 Depleted Haploidentical Hematopoietic Stem Cell Grafts Following Reduced-Intensity Conditioning

One-Day Low-Dose Etoposide Is an Effective Chemomobilization Regimen with Minimal Toxicity for Autologous Stem Cell Transplantation in Patients with Multiple Myeloma: A Multicenter Study from Tertiary Hospitals in Korea

PurposeAutologous stem cell transplantation (ASCT) is widely used as a part of induction treatment for transplantation-eligible patients with multiple myeloma. For successful ASCT, mobilizing hematopoietic stem cells from bone marrow to peripheral blood is essential because collecting a sufficient number of stem cells using apheresis is mandatory. As a method for mobilization, chemomobilization consisting of high-dose chemotherapy plus granulocyte-colony stimulating factor (G-CSF) or G-CSF alone (G-mobilization) has been used. However, the mobilization failure still remains a problematic issue. Given repeated mobilization attempts increase medical costs and the risk of morbidity related with apheresis, the mobilization process should be efficient. Chemomobilization is more effective than G-mobilization, however, chemomobilization has the risk of complication such as febrile neutropenia or chemotherapy-induced second malignancy. Thus, we compared the efficacy and safety of our new chemomobilization regimen, one-day low-dose etoposide with that of two-day low-dose etoposide, one-day high-dose cyclophosphamide, and G-mobilization.MethodsWe retrospectively analyzed 234 patients who underwent ASCT for MM between 2008 and 2018 in four tertiary hospitals in Korea. One-day low-dose etoposide regimen (E1) was the intravenous (IV) administration of etoposide (375 mg/m2) over 4 hours whereas two-day low-dose etoposide (E2) was the same dose of etoposide on 1st and 2nd day in outpatient clinic. G-CSF administration was started around on 10th day until the end of collection. In G-mobilization regimen (G), the injection of G-CSF was started four days (day -4) before initiation of apheresis (day 0), and G-CSF once a day was maintained untill the end of collection. One-day high-dose cyclophosphamide regimen (C) was the IV administration of cyclophosphamide (3.5 g/m2) on 1st day and the daily injection of G-CSF from 2nd day until the end of stem cell collection. Peripheral blood stem cell collection was started when CD34-positive cells or hematopoietic progenitor cells were more than 5000/μL in peripheral blood, or white blood cell count was more than 5000/μL. In this study, we defined the ‘adequate mobilization’ as CD34-positive cells more than 4ⅹ106/kg, and ‘mobilization failure’ as a collected CD34-positive cells less than 2ⅹ106/kg. Neutrophil and platelet engraftment was defined as more than 500/μL and 20,000/μL on consecutive two days, respectively.Results31 patients received single dose etoposide (E1) between 2016 and 2018 whereas 28 patients received double dose etoposide (E2) between 2011 and 2018. The other two regimens were used in 105 (C, 2008-2015) and 70 patients (G, 2008-2017) according to physicians' decision. The comparison of four regimens showed the median CD34-positive cells of E1 regimen was 5.57ⅹ106/kg that was comparable to that of E2 and C (Table 1). The number of CD34-positive cells on 1st day of apheresis was 4.03ⅹ106/kg in E1 regimen, and it was higher than that of C and G (3.32 and 1.79ⅹ106/kg, respectively). As a result, E1 regimen achieved ‘adequate mobilization’ in 100% of patients (n=30) like E2 regimen (n=28, 100%). Mobilization failure did not occur in E1 and E2 regimens whereas 4-10% of patients experienced mobilization failure in C and G regimens (Table 1). All patients receiving E1 and E2 regimen but one patient in E1 could collect more than 4ⅹ106/kg of CD34-positive cells within three cycles of apheresis. The occurrence of febrile neutropenia was extremely lower in E1 regimen (7%) than E2 and C regimens (43% and 34%, respectively, p<0.001). Both neutrophil and platelet engraftment were the fastest in E1 (the median 9 days after ASCT, p<0.001).ConclusionsOne-day low-dose etoposide administration could be effective for chemomobilization in myeloma patients with reduced risk of complication compared to two-day low-dose etoposide, high-dose cyclophosphamide and G-mobilization. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Contributions of Bioactive Molecules in Stem Cell-Based Periodontal Regeneration.

Periodontal disease is a widespread disease, which without proper treatment, may lead to tooth loss in adults. Because stem cells from the inflammatory microenvironment created by periodontal disease exhibit impaired regeneration potential even under favorable conditions, it is difficult to obtain satisfactory therapeutic outcomes using traditional treatments, which only focus on the control of inflammation. Therefore, a new stem cell-based therapy known as cell aggregates/cell sheets technology has emerged. This approach provides sufficient numbers of stem cells with high viability for treating the defective site and offers new hope in the field of periodontal regeneration. However, it is not sufficient for regenerating periodontal tissues by delivering cell aggregates/cell sheets to the impaired microenvironment in order to suppress the function of resident cells. In the present review, we summarize some promising bioactive molecules that act as cellular signals, which recreate a favorable microenvironment for tissue regeneration, recruit endogenous cells into the defective site and enhance the viability of exogenous cells.

Stem cell mobilization in patients with dialysis-dependent multiple myeloma: Report of the Polish Myeloma Study Group.

High-dose chemotherapy with autologous hematopoietic stem cell transplantation (auto-HSCT) improves the outcome of patients with multiple myeloma (MM). It seems that auto-HSCT is also a feasible therapeutic option in MM dialysis-dependent (MMDD) patients. However, to perform transplantation, a sufficient number of stem cells must be collected. Given that data on mobilization of auto-HSC efficacy and safety in dialysis-dependent patients are limited, we report data from all Polish Centers belonging to the Polish Myeloma Study Group. Twenty-eight dialysis-dependent MM-patients were enrolled into this retrospective analysis. The study population comprised patients diagnosed between 2004 and 2015 in whom an attempt to collect auto-HSC was made (68%: women, median age: 56). Patients received granulocyte-colony stimulating factor (G-CSF) alone or in combination with chemotherapy and autologous peripheral blood stem cells (auto-PBSCs) were collected by leukapheresis. The success rate in terms of obtaining sufficient number of CD34(+) cells/kg for an auto-HSCT (≥2 × 106 cells/kg body weight) during the first mobilization attempt was 92% (26/28 patients), and for 2 auto-HSCTs (≥4 × 106 cells/kg) - was 75% (21/28 patients). After the second mobilization attempt (undertaken in 8 patients), a sufficient number of CD34(+)/kg cells for an auto-HSCT was obtained for all patients and the number of CD34(+)/kg collected cells was sufficient for 2 auto-HSCT in 6 additional patients. Hematologic toxicity and infections were the most frequent complications. Higher doses of cytarabine (>1.6 g/m2 ) and cyclophosphamide (> 2 g/m2 ) should be avoided in MMDD patients due to toxicity. Further studies are needed to establish mobilization regimens, confirm their safety, and dosing in MMDD patients.

Difficulties of Mobilization and Harvesting of Hematopoietic Stem Cells in Heavily Pre-Treated Patients

AbstractThis study presents a new method of combined growth factor mobilization with increased dose (15 μg/kg) of filgrastim and standard mono dose of plerixafor in subjects with multiple myeloma, Hodgkin or non-Hodgkin lymphoma. Efficient mobilization is a key factor in the treatment of this group of patients, because autologous transplantation can be performed in cases in which we succeed to obtain a minimally sufficient number of stem cells. Autologous stem cell transplantation is included in the current standard treatment of multiple myeloma and in the relapsed cases of lymphomas.

Low-Dose Cyclophosphamide Plus Granulocyte Colony-Stimulating Factor As an Efficacious and Safe Peripheral Blood Stem Cell Mobilizing Regimen in Patients with Hematologic Malignancies

▪Introduction:Collection of adequate numbers of Hematopoietic Stem Cells (HSCs) is a prerequisite for proceeding to autologous bone marrow transplant. However, various studies have showed that approximately 5% to 40% of patients do not meet the minimum threshold of 2×106 CD34+ cells/kg that is associated with timely engraftment and better outcomes. There is little consensus over the optimal mobilization regimen for procurement of peripheral blood CD34 + stem cells (PBSCs). Studies comparing mobilization using high-dose cyclophosphamide(HD-CY) {5-7 g/m2} or intermediate dose cyclophosphamide (ID-CY) {3-4g/m2} plus G-CSF with low dose cyclophosphamide (LD-CY) {1-2g/m2) plus G-CSF have reported higher total PBSC yield with the former but at the cost of higher toxicity [Hiwase et al, Cytotherapy 2007]. Hence the significance of our study to evaluate the efficacy and safety of LD-CY plus G-CSF in the era of novel induction therapies, takes precedence in terms of optimal resource utilization.Methods:We retrospectively analyzed mobilization efficacy and need for supportive care in Myeloma and Lymphoma patients that received LD-CY (2g/m2) plus G-CSF (10µg/kg/day) as the preferred mobilization regimen. Patients treated with novel induction regimens only, with or without radiation were included. LD-CY was given on day 1 with MESNA. G-CSF was started Day +5 from LD-CY and continued until the completion of apheresis. We started to measure peripheral CD34+ (pCD34) when patient's white blood cell count recovered to >1000/µL. When the pCD34 count was ≥10/uL, apheresis was started.Results:Out of 21 patients analyzed, 14 (67%) were Myeloma, predominantly IgG subtype (71%) and 7(33%) Lymphoma predominantly Non-Hodgkins type (71%). Our study population comprised of 53% females, 81% Caucasians with median age of 59 years (30-66). 28.5% patients had radiation treatment in the past. Mean/Median lines of chemotherapy previously received were 2/1 (1-5).71% patients received Lenalidomide (57%) / Thalidomide (14%) based regimens for induction therapies. 52% patients had complete response at the time of chemo mobilization.Successful mobilization (defined as total CD34+ cells collected >2x106/kg) was significantly achieved in 95% patients with mean/ median collection of 12.4/11.5 x106 CD34+ cells/kg (20 out of 21 patients, One sample T- test with significance level, alpha =0.025 [one sided ]showed T statistic 5.36 with p-value <0.0001). First apheresis yielded at least 2 x10 6/kg CD34+ cells in 76 % patients with a mean/median of 5.8/3.8 x106/kg CD34+ cells. 66% of patients were found to be good mobilizers, defined as patients collecting ≥5 x106 CD34+ cells/kg in ≤2 days. 81% patients collected ≥5x106 CD34+ cells/kg and 57 % patients collected ≥10 x106 CD34+ cells/kg. Mean/Median Peak pCD34count was 97.7/60 cells/µL(2-488). Median number of aphereses was 2 (mean 2.6) with the mean and median time to apheresis being 11 days (9-15). None of the patients needed inpatient hospitalization or intravenous antibiotics during mobilization or had any complication related to immunosuppression. 14% and 19% of patients needed packed red blood cell and platelet transfusions respectively during the mobilization period. One Myeloma patient (4.8%) that failed mobilization had previously received five lines of chemotherapy including lenalidomide/thalidomide containing regimens and had stable disease at the time of mobilization. Interestingly, 61% of myeloma patients that successfully mobilized had received lenalidomide containing induction regimen (mean/median no. of cycles- 3.9/4[1-7])Conclusion:Our analysis shows that LD-CY plus G-CSF is efficacious to mobilize sufficient number of stem cells (>2x106 CD34+ cells /kg) in Myeloma and Lymphoma patients treated with novel agents, including the challenging sub-group of lenalidomide treated myeloma patients, with no infectious complications or morbidity noted during the entire period of mobilization. Therefore, we believe LD-CY plus G-CSF may overcome the negative effects of prior lenalidomide exposure on PBSC mobilization [Mark et al, Biol Blood Marrow Transplant 2008] and is superior in terms of optimal resource utilization compared to novel plerixafor based regimens [Chaudhary et al, J Clin Apher 2013]. Prospective studies involving CY based regimens will help elucidate the optimal dose of chemo-mobilization. DisclosuresNo relevant conflicts of interest to declare.

A novel approach for the enumeration of peripheral blood stem cells suitable for transplantation.

Stem cells have the capability to proliferate and differentiate into various cells of the body. Few stem cell sources have been approved for transplantation, among them are the hematopoietic progenitor cells which are progenitors of the myeloid and erythroid lineage in the hematopoietic system, that continually provides mature blood cells during the lifespan of the individual. These well-characterized stem cells are clinically relevant in the treatment of diseases such as breast cancer, leukemias, and congenital immunodeficiencies. Peripheral blood stem transplantation is a standard procedure after its first successful transplantation more than 35 years ago. The minimum intended dose of stem cells given to the patient is 2.5 × 106 –5 × 106 cells. In this study, we are establishing a correlation between the number of stem cells enumerated and the weight of the patient as a determinant for suitable transplantation. We have established a conversion factor to deliver the required dose of approximately 3 × 106 stem cells/kg body weight. This will ensure a uniform collection strategy that is sufficient for transplantation irrespective of the weight of the patient. This approach, if incorporated, will lead to a significantly lesser rate of bone marrow transplantation failures as sufficient number of stem cells will ensure engraftment of stem cells.

Stem cell mobilization in HIV seropositive patients with lymphoma

High-dose chemotherapy with autologous peripheral blood stem cell rescue has been reported as feasible and effective in HIV-associated lymphoma. Although a sufficient number of stem cells seems achievable in most patients, there are cases of stem cell harvest failure. The aim of this study was to describe the mobilization policies used in HIV-associated lymphoma, evaluate the failure rate and identify factors influencing mobilization results. We analyzed 155 patients who underwent attempted stem cell mobilization at 10 European centers from 2000-2012. One hundred and twenty patients had non-Hodgkin lymphoma and 35 Hodgkin lymphoma; 31% had complete remission, 57% chemosensitive disease, 10% refractory disease, 2% untested relapse. Patients were mobilized with chemotherapy + G-CSF (86%) or G-CSF alone (14%); 73% of patients collected >2 and 48% >5 × 10(6) CD34(+) cells/kg. Low CD4+ count and refractory disease were associated with mobilization failure. Low CD4(+) count, low platelet count and mobilization with G-CSF correlated with lower probability to achieve >5 × 10(6) CD34(+) cells/kg, whereas cyclophosphamide ≥ 3 g/m(2) + G-CSF predicted higher collections. Circulating CD34(+) cells and CD34/WBC ratio were strongly associated with collection result. HIV infection alone should not preclude an attempt to obtain stem cells in candidates for autologous transplant as the results are comparable to the HIV-negative population.

Brisk Elevation of Serum Lactate Dehydrogenase During G-CSF Priming is a Significant Predictor for the Ensuing Successful Autologous Peripheral Blood Stem Cell Collection

Autologous stem cell transplantation plays a critical role in the management of patients with hematologic malignancies such as multiple myeloma and malignant lymphoma. The minimum number of CD34 positive cells required for transplantation is believed to be 2.0×106 cells/kg. Up to 20% of the patients, however, fail to reach the number after a single collection attempt and require multiple aphereses to obtain a sufficient number of stem cells. To predict the outcome of mobilization, several pre-mobilization factors have been investigated, including peripheral blood CD34+ cell count at the time of harvest, but little is as yet known about the significance of serum lactate dehydrogenase (LDH). We, therefore, sought to evaluate the predictive potential of serum LDH for the autologous stem cell yield. Peripheral blood stem cell mobilization attempts were made from April 2002 to March 2012 for 55 consecutive patients of hematologic disorders. Primary diagnosis of our patients and their mobilization chemotherapy regimens are shown in Table 1. All patient received G-CSF (filgrastim 400μg/sq/day or lenograstim 10μg/kg/day subcutaneously). None was mobilized with G-CSF alone. Aphereses were repeated until the total stem cell yield exceeded 2×106cells/kg, for maximum 3 consecutive days. Forty-four patients successfully produced the sufficient number of stem cells, while 11 patients failed (poor mobilizer). Between the two groups, there were no significant differences in pre-mobilization WBC counts and serum LDH levels. However, the ratio of the LDH level immediately before apheresis to the lowest one (“LDH-ratio”) were significantly lower in poor mobilizers than good mobilizers. Our ROC analysis showed that the cut-off value of “LDH-ratio” was 1.75. An elevation of serum LDH levels to reach “LDH-ratio”>1.75 were found to be a useful predictive factor for successful CD34+ stem cell collection. The delay in reaching “LDH-ratio”>1.75 may suggest that we should modify the mobilization regimen by, for instance, adding prelixafor.Table 1Pts’ characteristicsNo. of Patients55Median age (range)56(35-69)Gender, male (%)31(56)Primary Disease MM19 NHL26 HL4 Others6Median WBC counts ×103/μL (range)2.1(1.5-3.2)Median LDH Level IU/L (range)196(145-445)PBSC mobilization regimen ESHAP27 HD-CY22 G-CSF alone0 Others6MM indicates: multiple myeloma, NHL: non-hodgkin lymphoma, HL: hodgkin lymphoma, HD-CY: high dose cyclophosphamide Open table in a new tab MM indicates: multiple myeloma, NHL: non-hodgkin lymphoma, HL: hodgkin lymphoma, HD-CY: high dose cyclophosphamide

Very High Efficacy of Stem Cell Mobilization Using Intermadiate-Dose Cytosine Arabinoside + G-CSF in Patients with Lymphoid Malignancies, Including Predicted and Proven Poor Mobilizers

AbstractAbstract 4131 Background:High-dose therapy followed by autologous peripheral blood stem cell transplantation (autoPBSCT) is widely applied in the treatment of lymphoid malignancies. However, a significant proportion of patients fail to mobilize sufficient number of stem cells after commonly used protocols. Patients and Methods:In this study we evaluated efficacy of first-line mobilization based on cytosine arabinoside (AraC) 1600 mg/m2 + G-CSF (filgrastim) 5–10 ug/kg. Results of 70 subsequent patients (diagnosis multiple myeloma, MM-39, Hodgkin’s lymphoma, HD-9, non-Hodgkin lymphoma, NHL-22) were compared with 45 individuals mobilized with cyclophosphamide (CTX) (4 g/m2) + G-CSF in a preceding period (MM-25; HD-4; NHL-16). In respective groups, 39 (56%) and 30 (67%) patients were classified as “predicted” poor mobilizers, defined as the presence of at least one risk factor: age >60 years, thrombocytopenia, preceding treatment with melphalan, fludarabine, preceding radiotherapy on pelvis, >8 courses of chemotherapy for MM, >3 courses containing cisplatin or carboplatin for lymphomas, preceding high-dose treatment with autoHSCT. In addition, we analyzed efficacy of AraC+G-CSF in 14 patients who had failed previous chemo-mobilization, mostly with CTX+G-CSF. Results:All but three patients (96%) treated with AraC reached >15 CD34+ cells/uL in peripheral blood, compared to 67% in the CTX+G-CSF cohort (p<0.0001). Median peak level was 127 (3–780) CD34+ cells/uL vs. 33 (1–240), respectively (p<0.0001). After AraC, 68 (97%) patients collected >=2×10e6 CD34+ cells/kg and 58 (84%) collected >=5×10e6 CD34+ cells/kg with the median 14.9 (2.3–54.6), which was achieved with a single leukapheresis in 92% patients. Using CTX, 67% patients collected sufficient number of stem cells (p<0.0001) and 76% required 2 or more leukaphareses (p<0.0001). AraC+G-CSF was effective in 37 (97%) predicted poor mobilizers compared with 16 (64%) in the CTX+G-CSF group (p=0.0006). The toxicity related to both regimens was comparable. Among patients who proceeded to autoPBSCT the time to neutrophil recovery after transplantation was the same for AraC and CTX cohorts (median 12 days), while platelet engraftment was faster for patients mobilized with AraC (13 days vs. 14 days, p=0.06).All 14 patients who had failed previous mobilization collected >=2×10e6 CD34+ cells/kg after AraC+G-CSF, with the median 9.3 (2.5–25.9). It was achieved with a single apheresis in 10 (71%) cases. Conclusions:Mobilization based on intermediate doses of AraC+G-CSF is highly effective allowing adequate CD34+ harvest in almost all patients with lymphoproliferative diseases, including predicted and proven poor mobilizers. The level of mobilized CD34+ cells is four times higher compared to protocol based on CTX. Disclosures:No relevant conflicts of interest to declare.

Efficacy of a Dose-Intensified CHOP (Double-CHOP) Regimen for Peripheral T-Cell Lymphomas

Abstract 4855 Background:Peripheral T-cell lymphomas (PTCLs) are a rare and heterogeneous group of non-Hodgkin lymphomas, often resulting in poor prognoses. The CHOP chemotherapy regimen, which includes cyclophosphamide, doxorubicin, vincristine, and prednisone, had been used previously to treat other types of lymphomas. Here, we examined the efficacy and safety of a dose-intensified CHOP regimen (Double-CHOP), which was followed by autologous stem-cell transplantation (ASCT) or high-dose methotrexate (HDMTX), in PTCL patients. Patients and Methods:Twenty-eight PTCL patients, who received 3 courses of Double-CHOP at our institution, were retrospectively studied from 1996 to 2012. Patients with anaplastic lymphoma-kinase-positive anaplastic large-cell lymphoma (ALK+–ALCL) were excluded from this study. The Double-CHOP regimen consisted of 3 courses of intravenous (iv) administration of cyclophosphamide (750 mg/m2, days 1–2, over 2 h), doxorubicin (50 mg/m2, days 1–2, over 30 min), vincristine (1.4 mg/m2, day 1, max 2 mg/Kg body weight), and per os (p.o.) prednisone (50 mg/m2, days 1–5). For patients aged >60, cyclophosphamide and doxorubicin doses were modified. The third cycle of Double-CHOP regimen was used for stem cell mobilization. Consolidating high-dose therapy (HDT) regimen consisted of cyclophosphamide (60 mg/kg, day 6 and 7, iv, over 3 h), etoposide (500 mg/m2; day 4, 5, and 6; iv; over 6–8 h), and ranimustine (250 mg/m2, day 2 and 3, iv, over 1 h). ASCT was performed on day 0 and G-CSF administered from day 1 until neutrophil engraftment. As an alternative to HDT/ASCT, HDMTX (8 g/m2, day 1, iv, over 4 h) was indicated for patients who could not yield a sufficient number of stem cells or were ineligible for HDT/ASCT. Results:Patients’ median age was 58 years (range: 17–69). They had low-intermediate (n = 11), high-intermediate (n = 10), or high (n = 7) risk according to the International Prognostic Index (IPI). The overall complete remission (CR) rate following Double-CHOP treatment was 68%. Of all the CR patients, 10 could successfully tolerate a consolidated HDT followed by ASCT, and 7 received HDMTX. Only a single case of treatment-related mortality was recorded during the study. On a median 31-month follow-up, the estimated 3-year overall survival (OS) rate and 3-year relapse-free survival (RFS) rate after CR were 68% and 60%, respectively. Conclusion:Although this study included elderly and excluded low-risk IPI and ALK+–ALCL patients, both RFS and OS results were superiorly favorable, indicating the efficacy of this Double-CHOP regimen and Double-CHOP followed by ASCT/HDMTX consolidations is safe and may achieve prolonged EFS, especially in patients with poor prognostic factors. However, an effective treatment strategy for refractory or relapsing patients needs to be validated and established. Disclosures:No relevant conflicts of interest to declare.

1082P - Very High Efficacy of Cytarabine + G-CSF Compared to Cyclophosphamide + G-CSF as Hematopoietic Stem Cell Mobilization in Patients with Lymphoid Malignancies Referred for Autologous Transplantation

ABSTRACT High-dose therapy followed by autologous peripheral blood stem cell transplantation (autoPBSCT) is widely applied in the treatment of lymphoid malignancies. However, a significant proportion of patients fail to mobilize sufficient number of stem cells. So far, no standards for mobilization have been elaborated with G-CSF alone or G-CSF in combination with cyclophosphamide (CTX) being most commonly used regimens. In this study we evaluated efficacy of mobilization based on cytosine arabinoside (AraC) 1600 mg/m2 + G-CSF (filgrastim) 5-10 ug/kg. Results of 67 subsequent patients were compared with 45 individuals mobilized with cyclophosphamide (CTX) (4 g/m2) + G-CSF, according to our preceding standard protocol. The median age of patients in the AraC group was 57 years; the indications were: multiple myeloma (MM, n = 37), Hodgkin's lymphoma (HL, n = 9) and non-Hodgkin lymphoma (n = 21). Patients had previously been treated with 8 (3-36) cycles of chemotherapy, 2 (1-6) lines of chemotherapy; 33% had received radiotherapy. The characteristics of the CTX group were comparable. All but two patients (97%) treated with AraC reached > 15 CD34 + cells/uL in peripheral blood, which was required to start leukapheresis, compared to 67% in the CTX + G-CSF cohort (p = 2x10e6 CD34 + cells/kg, required for single autoPBSCT, while 54 (81%) collected > = 5x10e6 CD34 + cells/kg, needed for double transplantation. Respective proportions in the CTX cohort were 56%(p We conclude that mobilization based on intermediate doses of AraC + G-CSF is highly effective allowing adequate CD34+ harvest in almost all patients with lymphoid malignancies, including heavily pre-treated and elderly individuals. The level of mobilized CD34+ cells is four times higher compared to commonly used protocol based on CTX. Hence, AraC + G-CSF may be a preferable option for predicted poor mobilizers, especially when high number of CD34+ cells is required for transplantation. Disclosure All authors have declared no conflicts of interest.

Hematopoietic Stem Cells: Transcriptional Regulation, Ex Vivo Expansion and Clinical Application

Maintenance of ex vivo hematopoietic stem cells (HSC) pool and its differentiated progeny is regulated by complex network of transcriptional factors, cell cycle proteins, extracellular matrix, and their microenvironment through an orchestrated fashion. Strides have been made to understand the mechanisms regulating in vivo quiescence and proliferation of HSCs to develop strategies for ex vivo expansion. Ex vivo expansion of HSCs is important to procure sufficient number of stem cells and as easily available source for HSC transplants for patients suffering from hematological disorders and malignancies. Our lab has established a nanofiber-based ex vivo expansion strategy for HSCs, while preserving their stem cell characteristics. Ex vivo expanded cells were also found biologically functional in various disease models. However, the therapeutic potential of expanded stem cells at clinical level still needs to be verified. This review outlines transcriptional factors that regulate development of HSCs and their commitment, genes that regulate cell cycle status, studies that attempt to develop an effective and efficient protocol for ex vivo expansion of HSCs and application of HSC in various non-malignant and malignant disorders. Overall the goal of the current review is to deliver an understanding of factors that are critical in resolving the challenges that limit the expansion of HSCs in vivo and ex vivo.