Efficacy Of Stem Cell Mobilization Research Articles

High Efficacy of Stem Cell Mobilization With Etoposide+Cytarabine Plus G-CSF in Patients With Multiple Myeloma.

BackgroundEfficient mobilization of CD34+ hematopoietic stem cells plays a vital role in successful autologous stem cell transplantation (ASCT) in patients with multiple myeloma (MM), especially in cases with high-risk cytogenetic recommended for tandem ASCT. However, the optimal mobilization strategy remains a matter of debate in the era of lenalidomide. The combination of etoposide with Cytarabine plus G-CSF as a novel mobilization regimen in MM has not been reported previously.MethodsThis research retrospectively studied mobilization efficacy and safety using etoposide combined with Cytarabine (etoposide 50–100 mg/m2, qd d1–3; AraC 0.5 g/m2, q12h d1~3) plus G-CSF (5 µg/kg/day, from d5 until the day of apheresis) in 128 patients with MM. 70(54.7%) patients received lenalidomide-based induction regimens treatmentResultsA median of 27.75×106 CD34+ cells/kg was collected in the first apheresis, and 28.23×106 CD34+ cells/kg were collected overall. Of the 128 patients, all achieved adequate collection (≥2×106 CD34+ cells/kg), 121(94.5%) achieved optimal collection for single ASCT (≥5×106 CD34+ cells/kg), and 114(89.1%) harvested optimal collection for tandem ASCT (≥10×106 CD34+ cells/kg). In particular, the target yield of optimal collection for tandem ASCT was reached in 82.8% (106/128) by a single apheresis procedure. 14 patients obtained deeper response post mobilization. In multivariate analysis, cycles of prior chemotherapy independently affected the optimal achievement of CD34+ cells (p=0.004, OR 0.695, 95% CI 0.544~0.888). Previous lenalidomide exposure did not significantly impair CD34+ cells collection. Although 68% episodes of antibiotic usage were observed, no severe infection or treatment-related mortality occurred.ConclusionStem cell mobilization with Etoposide + Cytarabine plus G-CSF was highly efficient and safe in patients with MM, which could be considered in high-risk MM patients who were referred for tandem ASCT.

Iron overload is correlated with impaired autologous stem cell mobilization and survival in acute myeloid leukemia.

Patients with acute myeloid leukemia (AML) undergoing consolidation with autologous stem cell transplantation (ASCT) depend on the successful mobilization of peripheral blood stem cells. However, the factors affecting the mobilization potential in AML patients and, in particular, the effect of transfusion-related iron overload on peripheral blood stem cell mobilization are largely unknown. We investigated the association of varying levels of iron overload and stem cell mobilization efficacy in consecutive AML patients after two induction cycles. A total of 113 AML patients in early first complete remission underwent the mobilization procedure. While 84 (74.3%) patients had serum ferritin levels exceeding 1000 μg/L, 26 (23.0%) patients had levels even higher than 2000 μg/L. Iron overload correlated with the number of preceding red blood cell transfusions and inversely correlated with circulating CD34+ cell levels (p = 0.04) at apheresis. Finally, the median progression-free and overall survival rates of patients with ferritin levels of higher than 2000 μg/L were shorter with 332 days versus 2156 days (p = 0.04) and 852 days versus 2235 days (p = 0.04), respectively. Our data suggest that transfusion-related iron overload is suppressing the mobilization potential and is associated with inferior outcome in AML.

Stem Cell Mobilization after Various Induction Regimens in Patients with Multiple Myeloma

Stem cell mobilization after various induction regimens in patients with multiple myelomaIntroduction: Rapid development of novel therapies for multiple myeloma has led to a significant improvement in response to the treatment. Stem cell mobilization before autologous stem cell transplantation is a source of considerable costs of transplant procedure. Whether modern induction regimens affect outcome of stem cell mobilization has not been extensively studied.Aim: The goal of this study was to compare efficacy of stem cell mobilization after different induction regimens in patients with multiple myeloma. The primary goal was to compare CTD (cyclophosphamide, thalidomide, dexamethasone), CVD (cyclophosphamide, bortezomib, dexamethasone) and VTD (bortezomib, thalidomide, dexamethasone) and regimens in terms of succesful stem cell collection.Methods: All patients with multiple myeloma who have been planned for stem cell collection and were treated with one of the above mentioned regimens were included in this retrospective analysis. The demographic data, amount of stem cells collected, number of days needed to reach the target collection were recorded. All patients received high dose cyclophosphamide 2,5 g/m2 prior to stem cell collection and were primed with G-CSF twice daily from day 5. The collection was started at day 10. Collection goal was 8x106/kg CD34+ cells.Results: 15 patients received CTD, 25 patients CVD and 16 patients VTD regimen before stem cell collection. Groups were comparable according to age, gender and myeloma stages. Mean collected cells at the end of collection were 9.2 (SD 2.8) for CTD, 12.3 (SD 5.6) for CVD and 10.1 (SD 2.1) for VTD (p=0.066). Mean daily harvest was 3.4, 8.0 and 7.6 x106/kg respectively (p=0.01). Mean days needed to reach desired harvest were 3, 2.25 and 1.6 days (p=0.001). No collection failure was observed.Conclusion: The best collection results were seen in patients after induction with CVD or VTD regimen. VTD regimen also required the least days for collection and seems to be most beneficial for cost of collection. CTD regimen shows the least efficacy in stem cell collection before autologous transplantation. All patients managed to harvest for at least one stem cell transplant. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Efficacy of stem cell mobilization in patients with newly diagnosed multiple myeloma after a CTD (cyclophosphamide, thalidomide, and dexamethasone) regimen

The CTD (cyclophosphamide, thalidomide, and dexamethasone) regimen is known to be an effective primary therapy in patients with newly diagnosed multiple myeloma (MM). However, stem cell yields after CTD remain inconsistent. The aim of the present study is to identify the influence of the CTD regimen on the outcome of peripheral blood stem cell (PBSC) collection. Fifty-four patients received four cycles of CTD, and PBSCs were mobilized with cyclophosphamide and G-CSF or with G-CSF alone. Each patient from whom ≤4.0×10(6) CD34(+) cells/kg were collected received a second mobilization course. The median duration from the start of a CTD regimen to the first collection was 4.3months. Forty-eight patients were mobilized with cyclophosphamide followed by G-CSF, and six patients were mobilized with G-CSF alone. The median day of apheresis was day 3 (range day 2-day 5). The overall response rate at mobilization was 96.3%, including 11.1% complete response, 22.2% very good partial response, and 63.0% partial response. The median number of harvested CD34(+) cells was 12.8×10(6) cells/kg. At the second mobilization, 88.9% of patients reached the minimal stem cell collection target of ≥2.0×10(6) cells/kg, and 75.9% of patients achieved the collection target of ≥4.0×10(6) cells/kg. CTD within four cycles is an effective primary therapy in patients with newly diagnosed MM and only minimally affects subsequent PBSC collection.

Prospective Randomized Trial of Len/Dex Induction Followed by Tandem MEL140 with Autologous Blood Stem Cell Transplantation and Len Maintenance Versus Continued Therapy with Len/Dex in Myeloma Patients Age 60–75 Years: Protocol-Defined Safety Analysis After 100 Patients

AbstractAbstract 2012In this trial (DSMM XIII) patients of age 60–75 years with newly diagnosed multiple myeloma having symptomatic and measurable disease were randomly assigned to either (A1) 3 cycles of Lenalidomide (25 mg po d1-21/28d) with low-dose Dexamethason (40 mg po d1, d8, d15, d22/28d), followed by stem cell mobilization and then further Len/Dex cycles until progression or (A2) 3 cycles of Len/Dex, followed by stem cell mobilization, tandem high-dose melphalan 140 mg/m2 with autologous blood stem cell transplantation and Len maintenance with 10 mg daily until progression. The maximum time of treatment was defined as 5 years. Antithrombotic prophylaxis consists of LMW heparin for 3 months in both arms. With continued Len/Dex (A1) antithrombotic prophylaxis is also continued after stem cell mobilization with either LMW herparin or aspirin, depending on risk factors. Here we report the severe adverse events (SAEs) after 100 patients who have completed at least 3 cycles induction therapy and the efficacy of stem cell mobilization.149 patients with a median age of 68 years and a proportion of patients with age ≥70 years of 40%, have been randomized since March 2010. With the first 100 patients having completed 3 cycles of induction Len/Dex, 105 SAEs have been reported in 67 patients. SAEs consisted of infections (36), thromboembolic events (7), fever (6), syncope (5), renal failure (4), fractures (4), dyspnoe (4), heart failure (3), pain (3), hyperglycemia (3), rash (1), plasma cell leukemia (1), second malignancy (1) and others. The second malignancy was a melanoma in situ which was completely surgically removed and reported as a SUSAR. Nine patients have died, 2 during study treatment (progression 1, sepsis 1) and 7 off study (progression 4, heart failure 1, sepsis 1, unknown 1).For stem cell mobilization, two alternative protocols were allowed. The decision was left open to the investigator. Either G-CSF only (lenograstim 10 μg/kg for 4 days or end of apheresis) or CE chemotherapy with G-CSF (cyclophosphamide 2500 mg/m2, etoposide 300 mg/m2 plus lenograstim 263 μg from day 5 until end of apheresis) were applied. Mobilization data from 75 patients are available. G-CSF only was used in 10 patients (13%) and resulted in a median of 5.3 × 106 CD34 positive cells/kg (range 1.5 – 8.9) collected with a median of 2 leukaphereses (range 1 – 4). CE plus G-CSF was used in 64 patients (85%) and resulted in a median of 7.5 × 106 CD34 positive cells/kg (range 2.0–38.0) collected with a median of 1 leukapheresis (range 1 – 4). In 2 patients (3%) stem cell mobilization failed after CE plus G-CSF.Len/Dex induction therapy was associated with an acceptable tolerability and feasibility in elderly myeloma patients. Stem cell mobilization was successful in 97% of patients. Disclosures:Straka:Celgene: Consultancy, Honoraria, Research Funding. Off Label Use: Lenalidomide used for initial therapy in multiple myeloma.

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.

Hematopoietic Cell Mobilization for Gene Therapy of Adult Patients with Severe Beta-Thalassemia: The Challenge of Splenectomy and the Role of Plerixafor

Abstract 166▪FN2▪This icon denotes a clinically relevant abstractIn preparation for a trial for gene therapy of thalassemia, we investigated the safety and efficacy of stem cell mobilization in adult patients with severe thalassemia using either GCSF or Plerixafor. We first assessed mobilization with G-CSF or G-CSF following pretreatment with Hydroxyurea (HU). HU was used in non splenectomized patients in order to reduce extramedullary hemopoiesis in spleen and the splenic size and in splenectomized patients in order to decrease the high number of platelets and mainly WBCs before G-CSF, so as to reduce the chance of complicated leukocytosis during mobilization. We present data on 23 patients from the G-CSF study (EudraCT number 2005-000315-10, NCT00336362) and on 14 patients from the Plerixafor study (EudraCT number 2009–014136-37, NCT01206075). The bulk of the CD34+cell enriched leukaphereses product in either study was cryopreserved for the future gene therapy trial. Mobilization with G-CSF was safe and effective in non-splenectomized patients (CD34+cellsX106/kg/2aphereses: 6.67±2.87, n=6). HU+G-CSF-treated subjects (n=4), mobilized successfully only when an optimal wash out period of approximately two weeks was maintained before G-CSF administration (CD34+cellsX106/kg/2aphereses: 7.34±1.19 vs 1.86±0.76, respectively, p=0.03). HU reduced the spleen size before G-CSF (376±96cm3 vs 556±218cm3) and resulted in less splenic enlargement during mobilization as compared to non-HU pretreated subjects (26.8% vs 60%, p=0.1), although this difference did not reach statistical significance. Splenectomized patients responded excessively to G-CSF by developing early hyperleukocytosis (day 3 mean WBCs: 80.0±7.5×103/μl) without a corresponding rise in blood CD34+cells. This necessitated a significant G-CSF dose reduction or hold resulting in poor yields in the majority of cases (CD34+cells/kg/2aphereses: 2.0±1.7, n=4). One-month HU-pretreatment (n=9), prevented the G-CSF-associated hyperleukocytosis during mobilization (mean WBCs: 60.4±20.8×103/μl) allowing for safe and successful CD34+cell collections (CD34+cells/kg/2aphereses: 6.6±2.3, p=0.02 vs G-CSF-alone), but again, only when an optimal 2-week wash out period was maintained. Despite the safe and effective mobilization with the optimal HU+G-CSF combination of splenectomized patients, HU pretreatment significantly prolonged the mobilization procedure. Plerixafor induced a rapid and effective mobilization in both the splenectomized and non-splenectomized patients (CD34+celsX106/kg/1 or 2 aphereses: 7.29±1.82, n=8 and 5.38±1.99, n=6, respectively) and it was very well tolerated. In SPL subjects, the mean yield per apheresis with plerixafor was higher, with a trend to significance, over its G-CSF counterpart and similar to the mean yield obtained by the optimally HU+G-CSF-treated patients in the G-CSF study (CD34+cells/kgX106/apheresis: 4.19±3.13 vs 1.01±0.85 vs 3.32±1.16, p=0.06 and p=0.6 respectively). One patient who was remobilized with the combination of G-CSF+plerixafor due to previous failure to collect 32×106/kg/2aphereses with G-CSF-alone, yielded 6.5χ10^6 CD34+cells/kg by one apheresis. Importantly also for the non-SPL patients treated with plerixafor, a mean increase of splenic volume of 10.9±14.3% was encountered which was significantly lower than the 60% mean spleen volume increase during G-CSF mobilization (p=0.03). There was no significant difference in the clonogenic capacity (CFU-GM, BFU-E) of CD34+cells mobilized by G-CSF, HU+G-CSF or Plerixafor. However, there was a trend for Plerixafor to mobilize more primitive HSC subpopulations (CD34+/CD38−, CD34+/CD38−/HLADR−) as compared to HU+G-CSF- or G-CSF-alone-mobilized cells.These results suggest that either G-CSF or Plerixafor could be used for mobilizing non-splenectomized patients. Plerixafor seems to represent the agent of choice for mobilization of splenectomized patients with thalassemia. Disclosures:Off Label Use: Plerixafor is used off-label for mobilization of thalassemic subjects.

CD34+ Subclasses of Blood Stem Cell Grafts Collected After Plerixafor Injection in Non-Hodgkin Lymphoma (NHL) Patients Mobilizing Poorly with Chemotherapy Plus G-CSF

Abstract 2990 Background:Mobilization of blood stem cells is difficult in a subgroup of patients with standard methods. Plerixafor, a CXCR4 antagonist, has been used for stem cell mobilization in combination with G-CSF for some years. Mobilization method used may affect not only efficacy of stem cell mobilization and collection but also graft content which on the other hand may have effect on post-transplant outcomes. No data is available on CD34+ subclasses in grafts collected after plerixafor administration in patients who mobilize poorly with chemotherapy plus G-CSF.Patients and Methods: Altogether blood stem cell grafts collected from 26 NHL patients were studies. Thirteen patients (8 M, 5 F, median age 51 yrs) were mobilized with a combination of chemotherapy and G-CSF ad received plerixafor due to poor mobilization followed by stem cell apheressis. Thirteen patients (10 M, 3 F, median age 56 yrs) were mobilized with chemotherapy plus G-CSF without plerixafor and served as controls. Samples from the first collection after plerixafor and from the first apheresis of control patients were studied by flow cytometry using the following antibodies: CD34, CD38, CD 117, CD133, CD19 and CD45. Viability of CD34+ cells after freezing was assessed with 7-aminoactinomycin D staining. Also in vitro growth of granulocyte/macrophage progenitors (GM-CFU) were assessed from all grafts. Patients were followed after high-dose chemotherapy in regard to hematopoietic reconstitution. Results:The number of viable cells in the grafts was comparable between the plerixafor and the control groups (Table 1). The number of the most primitive stem cells (CD34+CD38−CD133+) was higher in plerixafor mobilized grafts (Table 1). Most of the CD34+ cells were myeloid progenitors, as defined by their CD117 antigen co-expression. No differences in GM-CFU were observed between the groups. All except one patient had received high-dose therapy. The median number of CD34+ cells collected from the patients was comparable (3.1 vs. 3.3 × 106/kg). The median time to reach neutrophils > 10 × 109/L was 10 days from the stem cell infusion in both groups and time to unsupported platelets was also comparable (16 d vs. 13 d). Platelet counts at 1 month, 3 months and 6 months were comparable between the groups. Absolute lymphocyte counts were higher in plerixafor group but the differences were not statistically significant. One early toxic death occurred in the plerixafor mobilized group and one death due to disease recurrence in both groups with a median follow-up of 301 and 348 days from stem cell infusion in prelixafor and control groups, respectively. Conclusions:Plerixafor added to chemomobilization in NHL patients resulted in higher number of the most primitive CD34+ cells in the graft with comparable in vitro growth and engraftment potential after BEAM chemotherapy when compared to patients mobilized without plerixafor. Longer follow-up of higher patient numbers are needed to evaluate whether differences in graft content have an effect on patient outcomes.TableCD34+ subclasses and in vitro growth in graft collected after plerixafor injection in NHL patients mobilizing poorly with chemotherapy plus G-CSF and in patients mobilized without plerixafor.Mobilization without plerixafor, median (range), n=13Mobilization with plerixafor, median (range), n=13pCD34+ cell content of the graft (x 106/kg)1.80 (0.31–4.74)1.45 (0.4–4.40)0.858Proportion of CD34+, CD133-, CD38+ cells from all CD34+ cells (%)26.3 (13.0–44.5)25.9 (7.9–44.9)0.663Proportion of CD34+, CD133+, CD38+ cells from all CD34+ cells (%)70.7 (54.1–86.6)68.4 (51.9–90.0)0.473Proportion of CD34+, CD133-, CD38- cells from all CD34+ cells (%)0.4 (0.1–0.7)0.4 (0.1–2.1)0.407Proportion of CD34+, CD133+, CD38- cells from all CD34+ cells (%)1.6 (0.0–5.9)3.5 (1.0–11.6)0.010The most primitive stem cell (CD34+, CD133+, CD38-) content of the graft (x 106/kg)0.02 (0.00–0.12)0.07 (0.01–0.15)0.005GM-CFU (x 104/kg)16.2 (3.0–63.3)23.1 (6.0–43.0)0.817GM-CFU, granulocyte macrophage colony forming unit Disclosures:Jantunen:Genzyme: Honoraria, Membership on an entity's Board of Directors or advisory committees.

Peripheral Blood Stem Cell Mobilization In Hiv Positive Patients with Lymphoma Candidates to Autologous Transplantation: Predictive Factors Analysis for Failure or Suboptimal Stem Cell Collection

AbstractAbstract 2250 Background:Autologous stem cell (SC) transplantation (ASCT) is a potentially curative treatment for several hematologic malignancies and has been demostrated feasible and effective in HIV-related lymphoma (ARL). Peripheral blood SC collection could represent a major issue in the use of ASCT in HIV infected patients (pts). Aim:To evaluate the feasibility and efficacy of SC mobilization in HIV positive (pos) pts with lymphoma and identify factors influencing harvest results. Potential “ongoing” predictors of collection were also assessed. Patients and Methods:We retrospectively analysed 98 consecutive pts with ARL, candidates to ASCT, who underwent SC mobilization at 3 Italian and 2 Spanish centers from 2000 to 2010. A collection less than 2×106 CD34+ cells/kg was defined as “mobilization failure”, between 2–5 as “suboptimal collection” and more than 5 as “good collection”. Several parameters were evaluated for correlation with outcome: age, sex, lymphoma histopathology, disease status, WBC and Plt count at start of mobilization, type of mobilizing therapy, marrow disease, previous mobilization failure, n° of previous chemotherapy (CT) lines, months from first detection of HIV positivity, CD4 count and HIV-viremia. Moreover, circulating CD34+ and WBC count on the first day of CD34+ monitoring and their ratio (SC ratio = CD34/WBC) were assessed as “ongoing” outcome predictors. Results:A total of 127 attempts of SC harvest in 98 pts were analysed. Median age was 41.5 ys (28-65). Lymphoma diagnosis was DLBCL in 42% of cases, Burkitt 10%, plasmablastic 10%, HL 31%, anaplastic 5%, follicular lymphoma 1% and PEL 1%. Disease status was complete remission in 36%, chemosensitive disease in 53% and refractory disease in 10% of cases. In 3 cases bone marrow was involved and mobilizations failed. In 18% of cases pts received mobilizing therapy after 1 previous CT line, in 67% after 2 and in 16% after 3 or more. All pts but 2 were on antiretroviral therapy. Median CD4 count was 231/mcl (50-1146) and HIV-viremia was detectable in 22%. Median time from first HIV detection was 79.5 ms (3-295). In 24% of cases G-CSF alone (10-20 mcg/Kg) was used as mobilizing treatment, while CT + G-CSF (5-10 mcg/Kg) in 76%, including single-agent Cyclophosphamide (CTX) 1.5 gr/ms (13%), CTX >3 gr/ms (27%), platinum containing regimens (20%), ifosfamide containing regimens (11%) and others (5%). Mobilization failure occurred in 40% of procedures, a collection between 2–5 × 10^6 CD34/Kg in 24% and > 5 in 35%. Finally, of 98 pts who underwent SC mobilization, 22% failed to collect enough cell to perform ASCT, 12 pts even after repeated attempts, 33% had a suboptimal and 45% a good collection (4 and 5 pts respectively after repeated mobilizations). At univariate analysis failure was significantly associated with refractory disease, Plt < 150.000/cmm, CTX 1.5 gr/ms as mobilizing treatment, previous mobilization failure and circulating CD34+ cell < 7.4/mcl on the first day of monitoring; whereas CTX > 3 gr/ms, CD4 count and SC ratio > 0.002 were associated with a reduced risk of failure. In multivariate analysis refractory disease (p<0.0001) and CTX 1.5 gr/ms (p=0.003) were indipendent predictors for failure and SC ratio > 0.002 (p<0.0001) a protective factor. A good collection was predicted at univariate analysis by Plt and CD4 count, age, months from first HIV detection, CT + G-CSF as mobilizing therapy, CTX > 3 gr/ms, WBC count and circulating CD34+ cells >29,7/mcl at the first day of monitoring and SC ratio > 0,002, whereas G-CSF alone and previous mobilization failure were negative predictive factors. Multivariate analysis confirmed CTX > 3 (p<0.0001), CD34+ cells > 29,7 (p=0.0003) and SC ratio > 0,002 (0.0036) as indipendent factors for good collection. Conclusions:In this series of 98 ARL and 127 SC mobilization attempts, a substantial number of pts failed SC harvest (22%) whereas 33% had a suboptimal and 45% a good collection. Lymphoma status and mobilizing treatment seems the strongest predictors for outcome, with refractory disease and low CTX dose (1.5 gr/ms) significantly associated with failure and CTX > 3 gr/ms predictor for good collection. A high ratio between circulating CD34+ cells and WBC on the planned day of first apheresis might represent a useful “ongoing” parameter to predict the outcome. These data might help to decide the mobilizing strategy in ARL and could provide the framework to rationally explore the use of new mobilizing agents Disclosures:No relevant conflicts of interest to declare.

Correlation of Cytokines Levels and Adhesion Molecules Expression with Stem Cell Mobilization Efficacy in Healthy Donors

Peripheral blood stem cells (PBSC) are standard source of hematopoietic stem cells for allogeneic transplantations. Mobilization of PBSC in healthy donors is induced by a short term administration of G-CSF. The biological basis of the mobilization procedure in not completely discovered. Several factors were identified influencing the mobilization efficacy however their predictive potential for detection of poor mobilizers is limited. We performed a prospective study to evaluate differences in cytokines levels and adhesion molecules expression between good and poor mobilizers. The aim was to find out whether some of these factors can predict mobilization efficacy. Sixty healthy donors (25 related, 35 unrelated) were included in the study. The median age was 39 years (26–67). All donors were treated with G-CSF 10 μg/kg/day (filgrastim, Neupogen) for 5 days. Aphereses were started on day+5. Blood levels of certain cytokines (SDF-1, ICAM, VCAM, MMP-9, IL-6, IL-8, fractalkine, TNFα, VEGF, E-selectin) were tested before G-CSF application (day+0) and at first apheresis (day+5). Adhesion molecules expression (CD11a, CXCR4, CD44, CD117, CD26, CD49d) on CD34+ cells was measured at day +5. Cytokines were assayed by multiplex xMAP or ELISA technology. CD34 positive cells and adhesion molecules were evaluated with the flow cytometry using standard protocols. In response to the G-CSF stimulation the following cytokines significantly increased: ICAM (p<0.0001), VCAM (p<0.0001), MMP-9 (p=0.0039) IL-6 (p=0.0133), TNFα (p<0.0001) and E-selectine (p<0.0001). SDF-1 (p=0.0001), IL-8 (p=0.0013) decreased and fractalkine and VEGF remained unchanged. There was a positive correlation of day+5 SDF-1 (p=0.0011) and VCAM (p<0.0001) levels with CD34+ count at day+5. As for IL-6 borderline negative correlation (p=0.0861) between day+0 cytokine level and day+5 CD34+ count was found. Afterwards the donors were divided into two groups according to the CD34+ count at day +5. The cut-off of 40.0 CD34+ cells/μl was used for distinguishing of poor mobilizers. Twenty two percent (13 donors) mobilized below and 78 % (47 donors) above the cut-off. Between good and poor mobilizers there were significant differences of ICAM levels at day+0 (p=0.0369) and day+5 (p=0.0023), VCAM at day+5 (p=0.117) and IL-8 at day+5 (p=0.0473). Using logistic regression ICAM and IL-6 measured at day+0 (before stimulation) were tested as predictors of mobilization efficacy. The ICAM level below cut-off of 100 ng/mL implies approx. 5× higher risk of poor mobilization (odds ratio 4.8, p=0.0206). Conversely the IL-6 level above cut-off of 32 pg/mL means approx. 16× higher risk of poor mobilization (odds ratio 15.6, p=0.0112). Immunophenotyping of CD34+ cells suggested an inverse relationship of CD34+ counts with two adhesion molecules expression: CD11a (p=0.0002), CXCR4 (p=0.0075). However the expression of all tested antigens was similar in both donors groups. G-CSF stimulated PBSC mobilization results in increased plasma levels of some cytokines mostly evident for ICAM, VCAM, TNFα and decreased levels of SDF-1 and IL-8. In cases of ICAM and IL-6 the kinetics of these changes correlates with the quality of PBSC mobilization in peripheral blood. Their levels measured before G-CSF mobilization might serve as predictive factor for mobilization efficacy and graft quality. Contribution of adhesion molecules to stem cell mobilization is less clear and their practical utilization for mobilization course management is low.

Comparison of CXCR-4 and Adhesion Molecule Expression in Healthy Bone Marrow with Expression in Bone Marrow and Peripheral Blood of Patients Receiving G-CSF Plus AMD3100.

It is known that the crosstalk between adhesion molecules, bone marrow microenvironment, and cytokines facilitates the multi step process of stem cell mobilization from bone marrow to peripheral blood. A combination of G-CSF plus AMD3100 - a CXCR-4 antagonist - has been shown to be safe and efficient in stem cell mobilization of healthy donors and cancer patients. Nevertheless, data predicting the efficacy of this approach are still missing. The present study investigated the correlation of the expression of CXCR-4 (CD184) and adhesion molecules with the kinetics and efficacy of stem cell mobilization in nine patients with Multiple Myeloma (MM) or NHL, respectively. Steady-state mobilization was performed using a combination of G-CSF (Filgrastim, 10μg/kg/d, 8 am) for 4 days followed by AMD3100 (240μg/kg) on day 4 at 10pm. Autologous aphereses were started on day 5. Bone marrow and peripheral blood (PB) before AMD3100 application (day 4) and PB on day 5 were investigated with a 4-color flow cytometric procedure. Bone marrow aspirates of healthy donors (n=20) served as control. The qualitative (%) and quantitative (mean fluorescence intensity, [MFI]) antigen expression of CXCR-4 in relation to CD34 was assessed as well as the expression of certain adhesion molecules including LFA-1, PECAM-1, VLA-1, L-selectin and CD44. First, the median percentage of CXCR-4 surface expression in healthy bone marrow was significantly higher (92%; range: 52 – 99%) than in patients bone marrow (70%; 30 – 88%; p=0.002), PB before AMD3100 (87%; 35 – 97%; p=0.050) and on day 5 (17%; 2 – 74%; p<0.001), whereas cytoplasmic expression was comparable (91%; 53 – 95%) in all cell compartments. The median quantitative CXCR-4 surface expression was significantly decreased in PB on day 5 compared to pre AMD3100 (14 vs. 95; p=0.003). Furthermore, the qualitative expression of LFA-1 and the quantitative expression of LFA-1, PECAM-1, VLA-1, and CD44 were also downregulated in response to AMD3100 (p<0.010). Second, a median of 63/μl (range: 15 – 132/μl) CD34+ cells was measured in the PB on day 5. Thus, a high absolute count of CD34+ cells in the PB on day 5 significantly correlated with lower qualitative and quantitative CXCR-4 expression in the same material (r=0.833; p=0.015). Evaluating CXCR-4 expression in bone marrow, PB before AMD3100 and on day 5 no significant correlation to CD34+ counts could be detected. However, there was one very poor mobilizing patient (15/μl CD34+ cells on day 5) in whom the quantitative CXCR-4 expression in the bone marrow was significantly higher than the median of all patients (MFI 95 vs. 26). Furthermore, some of the adhesion molecules (L-selectin, VLA-4, and CD44) showed a rather positive correlation with CD34 count. In summary, these preliminary data suggest that the amount of CD34+ cells in the peripheral blood after G-CSF plus AMD3100 application seems to be negatively correlated with CXCR-4 expression. A higher quantitative CXCR-4 expression in the bone marrow pre AMD3100 might predict a lower mobilization efficacy.