Autologous Peripheral Blood Progenitor Cell Research Articles

Long Term Remission of Capillary Leak Syndrome Associated with Monoclonal Gammopathy with Progression to Multiple Myeloma After Autologous Stem Cell Transplantation: a Case Report and Review of the Literature.

Clarkson's disease is a very rare entity characterised by acute episodes of systemic oedema and severe hypotension associated with paraproteinaemia. Its classical treatment relies on methylxanthine combined with terbutaline. Although this prophylactic therapy reduces the mortality rate, relapses are frequent. Eighty percent of patients with Clarkson's disease present with monoclonal gammopathy of unknown significance (MGUS). The risk of progression to multiple myeloma is 1% per year. Here, we present a 49-year-old woman who suffered multiple such episodes requiring treatment in the intensive care unit. Treatment with terbutaline and theophylline was ineffective. She was diagnosed with multiple myeloma (MM) 8 years after the first of these acute episodes. Antimyeloma treatment with bortezomib and dexamethasone was started, followed by autologous haemopoietic transplantation, with no further acute episodes since then. Our case is, to our knowledge, unique because eradication of MM was followed by complete disappearance of acute episodes of capillary leakage. Our case report is also the first to support the use of bortezomib and dexamethasone in this setting. Furthermore, autologous peripheral blood progenitor cell transplantation consolidated the MM stringent complete remission achieving a very long progression-free survival (> 11 years) of both MM and Clarkson's disease.

Multicenter Phase III Trial to Evaluate CD34+ Selected Versus Unselected Autologous Peripheral Blood Progenitor Cell Transplantation in Multiple Myeloma

High-dose chemotherapy followed by autologous transplantation has been shown to improve response rates and survival in multiple myeloma and other malignancies. However, autografts frequently contain detectable tumor cells. Enrichment for stem cells using anti-CD34 antibodies has been shown to reduce autograft tumor contamination in phase I/II studies. To more definitively assess the safety and efficacy of CD34 selection, a phase III study was completed in 131 multiple myeloma patients randomized to receive an autologous transplant with either CD34-selected or unselected peripheral blood progenitor cells after myeloablative therapy. Tumor contamination in the autografts was assessed by a quantitative polymerase chain reaction detection assay using patient-specific, complementarity-determining region (CDR) Ig gene primers before and after CD34 selection. A median 3.1 log reduction in contaminating tumor cells was achieved in the CD34 selected product using the CEPRATE SC System (CellPro, Inc, Bothell, WA). Successful neutrophil engraftment was achieved in all patients by day 15 and no significant between-arm difference for time to platelet engraftment occurred in patients who received an infused dose of at least 2.0 x 10(6) CD34(+) cells/kg. In conclusion, this phase III trial demonstrates that CD34-selection of peripheral blood progenitor cells significantly reduces tumor cell contamination yet provides safe and rapid hematologic recovery for patients receiving myeloablative therapy.

Low baseline platelet count predicts poor response to plerixafor in patients with multiple myeloma undergoing autologous stem cell mobilization

Background aimsBaseline platelet count has been shown to be a sensitive predictor of autologous peripheral blood progenitor cell collection yield in patients with multiple myeloma mobilized with granulocyte colony-stimulating factor (G-CSF). Patients who mobilize poorly with G-CSF are often treated with plerixafor to enhance mobilization. There are no surrogate markers available to predict response to plerixafor. MethodsWe retrospectively analyzed data from 73 patients with multiple myeloma who did not have adequate mobilization with G-CSF alone and were treated with plerixafor as a rescue agent. ResultsWe found that baseline platelet count directly correlated with peripheral blood CD34+ (PB-CD34+) count after plerixafor treatment (r = 0.36, P < 0.0001) and the number of PB-CD34+ cells collected on the first day of apheresis and inversely correlated with the number of apheresis sessions needed to collect the target number of PB-CD34+ cells (P = 0.0015). Baseline platelet count of 153 000/µL or less was associated with 90% specificity of predicting poor response to plerixafor with a sensitivity of 33%. ConclusionsBaseline platelet count is a good predictor of mobilization response to plerixafor in patients with multiple myeloma.

Evaluation of the Role of Plerixafor in Peripheral Blood Progenitor Cell Mobilization and Collection in Autologous Donors with Multiple Myeloma and Non-Hodgkin Lymphoma

Purpose Autologous hematopoietic stem cell transplantation is commonly used as treatment for patients with Multiple Myeloma (MM) or Non-Hodgkin Lymphoma (NHL). The mobilization and collection of peripheral blood progenitor cells (PBPC) to reach the required cell dose for transplantation is problematic for some patients when standard mobilization agents such as G-CSF, alone or combined with chemotherapy, are used. These patients are considered poor mobilizers with &lt; 20 CD34+ cells /μl on the planned day of collection and the optimal strategy to collect a sufficient number of PBPC in poor mobilizers is still not fully known. Plerixafor, a CXCR4 inhibitor, when added to the standard mobilization regimen, has been shown to improve mobilization and increase the probability of collecting a PBPC graft &gt; 2 X106CD34 /kg in standard mobilizers. In our center, Plerixafor became available in October 2012. However, how best to use plerixafor to improve care for poor mobilizers while ensuring the most cost-effective use of health care resources for this patient population remains an important question. To begin answering this question, we evaluated the impact of plerixafor usage in our population of poor mobilizers in our institution using a historical cohort for comparison. Material and method The population of poor mobilizers in the setting of MM or NHL (defined as &lt; 20 CD34 / μl on the planned day of collection) between January 2009 and December 2017 at Hôpital de l'Enfant Jesus (HEJ) du CHU de Québec was retrospectively studied and analyzed. To ensure that all poor mobilizers were included, we screened all flow cytometry CD34+ cell count results &lt; 50 /μl on the planned day of collection and reviewed all the charts of patients with &lt; 20 CD34 / μl regardless of whether they were collected or not. This chart review allowed a comprehensive description of the characteristics of our poor mobilizers, a comparison of the capacity to collect an adequate PBPC with or without plerixafor and to evaluate its impact on different variable with regard to PBPC collection and subsequent engraftment. The data were compiled in an ACCESS database using a retrospective review of digital patient charts. The data was analyzed by the Centre de Recherche du CHU de Quebec (CRCHUQ). Data were stratified by disease type and CD34 level on the planned day of PBPC collection. Results 207 patient charts were reviewed; 70 poor mobilizers with 95 mobilizations attempts, 30 of them using plerixafor, were included. The baseline characteristics of both groups were similar as well as the length of hospitalization for transplant and relapse free survival at one year. The addition of Plerixafor to our management of poor mobilizers did not significantly increase the proportion of patients proceeding to autologous PBPC transplant (80,4 vs 75% p=0,7599). During the mobilization attempts, the maximal CD34 /μl reached was similar in both groups (18,86 vs 15,4 p=0,1541). Plerixafor was associated with a superior collection efficiency (% of CD34 collected) (40,85 vs 60,91% p=0,007), but this can be explained by a greater use of central venous catheters for collection in this group (70 vs 95,1% p = 0,0275). Data stratified by initial diagnosis and CD34 level on the planned day of PBPC collection (&lt;10 c/μl versus [10c/μl - 20c /μl [) showed the same tendency. The collection duration was not significantly reduced in the plerixafor group, but there was a trend for a shorter mean duration of 22 minutes also explained by a greater use of central venous catheters in this group. The preplanned subgroups analyses have shown the same trend when stratified by diagnosis, with a reduction of 53 minutes in the multiple myeloma subgroup with a greater total CFU-GEM / kg per collect (46,35 vs 70,55 p=0,0163). Conclusion The use of plerixafor at HEJ did not improve significantly the proportion of poor mobilizers reaching an adequate PBPC graft and did not improve transplant outcomes compared to a historical control. Better strategies are needed to improve care for true poor mobilizers. The optimal use of plerixafor needs to be further studied. Table. Disclosures Blais-Normandin: Sanofi: Other: unrestricted research grant. Laroche:Sanofi: Other: unrestricted research grant.

G-CSF administration favours SDF-1 release and activation of neutrophils and monocytes in recipients of autologous peripheral blood progenitor cells

G-CSF is a growth factor widely used to mobilise CD34+ progenitor cells for clinical applications. The present study aimed to assess expression of G-CSF receptor (CSF3R) on neutrophils and monocytes, as well as SDF-1 and G-CSF serum levels in relation to efficacy of G-CSF-induced mobilisation for autologous transplantation. For this purpose, 105 patients with haematological disorders and 46 healthy controls were investigated.Before mobilisation patients were characterised with significantly higher percentage of CSF3R expressing neutrophils (p < 0.001) and monocytes (p = 0.002), than controls. G-CSF administration resulted in a decrease of CSF3R+ neutrophils (p < 0.001) and monocytes (p < 0.001), while presence of G-CSF receptor on neutrophils tended to negatively affect mobilisation yield (p = 0.075).G-CSF concentration increased during mobilisation (p < 0.001). On the 5th day of mobilisation a positive correlation was observed between G-CSF and SDF-1 serum levels (p < 0.001) and the number of CD34+ cells released from bone marrow seemed to be related to both G-CSF (p = 0.036) and SDF-1 levels (p = 0.084).As compared to Hodgkin’s lymphoma patients, those with multiple myeloma had lower basal percentage of CSF3R+ neutrophils (p = 0.014) while Non-Hodgkin’s lymphoma cases exhibited higher G-CSF (p = 0.026) and SDF-1 (p = 0.006) concentration on mobilisation day 5. Hodgkin’s lymphoma patients were also characterised with worse mobilisation efficacy than multiple myeloma (p = 0.022) and Non-Hodgkin’s lymphoma (p = 0.013) patients.These results suggest that both SDF-1 and G-CSF play a role in HSC release into peripheral blood and show that G-CSF administration affects expression of CSF3R on monocytes and neutrophils, implying potential role of these cell subpopulations in mobilisation process.

Multigene Measurable Residual Disease Assessment Improves Acute Myeloid Leukemia Relapse Risk Stratification in Autologous Hematopoietic Cell Transplantation

We report here the largest study to date of adult patients with acute myeloid leukemia (AML) tested for measurable residual disease (MRD) at the time of autologous hematopoietic cell transplantation (auto-HCT). Seventy-two adult patients who underwent transplantation between 2004 and 2013 at a single academic medical center (University of California San Francisco) were eligible for this retrospective study based on availability of cryopreserved granulocyte colony–stimulating factor (GCSF)–mobilized autologous peripheral blood progenitor cell (PBPC) leukapheresis specimens (“autografts”). Autograft MRD was assessed by molecular methods (real-time quantitative PCR [RQ-PCR] for Wilms tumor 1 (WT1) alone or a multigene panel) and by multiparameter flow cytometry (MPFC). WT1 RQ-PCR testing of the autograft had low sensitivity for relapse prediction (14%) and a negative predictive value of 51%. MPFC failed to identify MRD in any of 34 autografts tested. Combinations of molecular MRD assays, however, improved prediction of post–auto-HCT relapse. In multivariate analysis of clinical variables, including age, gender, race, cytogenetic risk category, and CD34+ cell dose, only autograft multigene MRD as assessed by RQ-PCR was statistically significantly associated with relapse. One year after transplantation, only 28% patients with detectable autograft MRD were relapse free, compared with 67% in the MRD-negative cohort. Multigene MRD, while an improvement on other methods tested, was however suboptimal for relapse prediction in unselected patients, with specificity of 83% and sensitivity of 46%. In patients with known chromosomal abnormalities or mutations, however, better predictive value was observed with no relapses observed in MRD-negative patients in the first year after auto-HCT compared with 83% incidence of relapse in the MRD-positive patients (hazard ratio, 12.45; P = .0016). In summary, increased personalization of MRD monitoring by use of a multigene panel improved the ability to risk stratify patients for post–auto-HCT relapse. WT1 RQ-PCR and flow cytometric assessment for AML MRD in autograft samples had limited value for predicting relapse after auto-HCT. We demonstrate that cryopreserved autograft material presents unique challenges for AML MRD testing because of the masking effects of previous GCSF exposure on gene expression and flow cytometry signatures. In the absence of information regarding diagnostic characteristics, sources other than GCSF-stimulated PBSC leukapheresis specimens should be considered as alternatives for MRD testing in AML patients undergoing auto-HCT.

Infusion of Ex Vivo Expanded Allogeneic Cord Blood-Derived Natural Killer Cells in Combination with Autologous Stem Cell Transplantation for Multiple Myeloma: Results of a Phase I Study

Background: Multiple myeloma (MM) is an incurable disease thought to be characterized by immune dysregulation and exhaustion, whereby proliferation of malignant plasma cells is not checked by the native immune system. Long term remissions in some patients after allogeneic stem cell transplant (SCT) suggest a graft versus myeloma effect; however the treatment-related toxicity limits the widespread use of this modality. Allogeneic natural killer (NK) cells are active in various hematologic malignancies and may have a role against MM, without concomitant graft versus host disease (GVHD). Umbilical cord blood is a potential source for allogeneic NK cells and ex vivo expanded umbilical cord blood-derived NK (CB-NK) cells demonstrate activity comparable to that of peripheral blood-derived NK cells. We describe here the results of a phase I, first-in-human study of ex vivo expanded allogeneic CB-NK cells in conjunction with high dose chemotherapy and autologous SCT.Methods: Patients with symptomatic MM who were appropriate candidates for high dose chemotherapy and autologous SCT were eligible. CB units with at least 4/6 match at HLA-A, B and DR were chosen for each patient. When possible, CB units with potential NK alloreactivity were prioritized. On day (-19) CB units were thawed and mononuclear cells (MNCs) were isolated by ficoll density gradient centrifugation. MNCs were cultured in a gas permeable bioreactor with irradiated (100 Gy) K562-based aAPCs expressing membrane bound IL-21 "Clone 9.mbIL21" (2:1 feeder cell:MNC ratio) and IL-2 (100 IU/mL). On day 7, cells were CD3-depleted via immunomagnetic depletion and remaining cells were re-stimulated with aAPC feeder cells and cultured for an additional 7 days. NK cell purity was determined after 14 days of culture. Due to pre-clinical data demonstrating synergy between lenalidomide and NK cells, patients received lenalidomide (10 mg orally daily) from days (-8) to day (-2). Melphalan 200 mg/m2 was given intravenously on day (-7). Freshly expanded CB-NK cells were infused on day (-5). Autologous peripheral blood progenitor cells (PBPC) were infused on day (0).Results: 12 patients have been enrolled thus far with 3 patients each on the following CB-NK cell dose levels: 5 e6 NK cells/kg, 1 e7 NK cells/kg, 5 e7 NK cells/kg and 1 e8 NK cells/kg. 11/12 patients had at least 1 high-risk feature of progressed/relapsed disease (n=7), high-risk cytogenetics (n=2) or International Staging System III disease (n=3). Successful NK expansion to target dose was achieved in all patients with median NK purity (CD56+/CD16+/CD3-) of 98.9% (96.8-99.7). Expanded cells demonstrated cytotoxicity against classic K562 and MM cell line targets. There were no infusional toxicities and no occurrence of GVHD. One patient (1 e8 NK cells/kg) failed to engraft due to a poor PBPC graft quality; this patient was rescued with a back-up autologous PBPC graft. There have been no other significant adverse events and no second primary malignancies. 11/12 patients are evaluable beyond day 100. Best response has been 8/12 nCR or better, 2/12 VGPR and 1/12 PR. 4/12 patients have progressed at a median of 330 days. By DNA microsatellite chimerism analysis, donor CB-NK cells were detected in 2 patients at the 1 e7 NK cells/kg dose and all 3 patients in the 1 e8 NK cells/kg dose, for at least 5 days after infusion. By a more sensitive flow cytometric chimerism assay using HLA class I-specific antibodies for donor or recipient, donor CB-NK cells were detected in 3 evaluable patients at doses of 1 e7 NK cells/kg, 5 e7 NK cells/kg and 1 e8 NK cells/kg for at least 12 days after infusion. Further analysis of these cells indicated persistence of an activated phenotype (NKG2D+/NKp30+) in vivo.Conclusion: CB-NK cells can be activated and expanded to clinical scale. This is the first clinical study of CB-NK cells for MM. When infused in the setting of myeloablative chemotherapy, up to 1 e8 allogeneic CB-NK cells/kg are well tolerated with no infusional toxicities or GVHD. These cells can persist for at least 12 days in vivo and demonstrate an active phenotype. Though clinical data are early, responses are encouraging in this high-risk patient population. Further updated data will be presented at the annual meeting. DisclosuresOff Label Use: Lenalidomide with high dose chemotherapy and autologous stem cell transplantation. Kaur:UT MD Anderson Cancer Center: Employment. Orlowski:Millennium Pharmaceuticals: Consultancy, Research Funding; BioTheryX, Inc.: Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Research Funding; Genentech: Consultancy; Onyx Pharmaceuticals: Consultancy, Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Spectrum Pharmaceuticals: Research Funding; Acetylon: Membership on an entity's Board of Directors or advisory committees; Janssen Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Array BioPharma: Consultancy, Research Funding; Forma Therapeutics: Consultancy. Cooper:Intrexon: Equity Ownership, Patents & Royalties, Research Funding; ZIOPHARM Oncology: Employment, Equity Ownership, Patents & Royalties, Research Funding. Lee:Cyto-Sen: Equity Ownership; Ziopharm: Equity Ownership; Intrexon: Equity Ownership.

Multigene MRD Assessment Improves AML Relapse Risk Stratification in Autologous Hematopoietic Cell Transplantation

▪While commonly used in other hematological malignancies, high dose chemotherapy followed by autologous hematopoietic cell transplantation (auto-HCT) has not been widely adopted in acute myeloid leukemia (AML) due to concerns regarding high post-transplant relapse rates. These relapses may be due, in part, to autograft contamination with AML. High sensitivity methods to detect residual AML have demonstrated the ability to correctly identify patients in morphological complete remission (CR) at risk of relapse. We sought to determine if testing of the graft prior to transplantation could predict post auto-HCT relapse. Prior studies investigating the utility of measurable residual disease (MRD) in auto-HCT have been limited by small patient cohorts and/or by the use of a single biomarker to detect AML, a heterogeneous cancer. We report here the largest retrospective study to date of adult auto-HCT AML patients tested for measurable residual disease (MRD) by both molecular methods (RQ-PCR) and multi-parameter flow cytometry (MPFC).Seventy-two patients transplanted between 2004-2013 at a single academic medical center (UCSF) were eligible for this study based on availability of cryopreserved GCSF mobilized autologous peripheral blood progenitor cell (PBPCs) specimens. All samples were collected on IRB approved research protocols.The median age at time of transplant was 48 (24-69), 54% were female, cytogenetics were known in 94% (of which 24% were favorable, 69% intermediate, 7% poor), 99% were in CR at the time of HCT (64/72 in CR1, 6 in CR2, 1 in CR3). Following auto-HCT 1 year RFS was 50% (1 year relapse rate 43%) and 2 year RFS was 40% (2 year relapse rate 47%).Wilms tumor 1 (WT1) is expressed in up to 90% of AML, but sufficiently over-expressed in peripheral blood to have utility as a sensitive marker of MRD in less than 50% of cases. RQ-PCR detectable WT1 above the previously established European LeukemiaNet threshold was found in 9 patients, of whom 6 relapsed in the first year after auto-HCT (19% sensitivity, 93% specificity, PPV: 67%, NPV: 60%).We have previously reported that multi-gene testing can augment WT1 based MRD detection in AML. We were unable however to use one component of our previously reported AML MRD panel, PR3, due to increased baseline expression in these GCSF stimulated PBPC products. Increased PR3 expression following GCSF administration, correlating with neutrophilia, was also observed in an independent cohort of healthy donors.Testing for PRAME, MSLN, CCNA1, t(8,21), Inv16, t(15:17) and NPM1 mutations A, B and D, as a supplement for WT1, in pre-HCT PBPCs resulted in substantially improved ability to predict post auto-HCT relapse (52% sensitivity, 80% specificity, PPV: 67%, NPV: 69%). Addition of these extra genes allowed for correct identification of 10 additional MRD+ patients who relapsed within 1yr after transplant, and added 5 false positive patients (1 of whom suffered early non-relapse mortality, and another who did relapse but more than 1 year after HCT).Finally, in expert hands, flow cytometry can identify residual AML with high sensitivity. Forty PBPC samples from the above cohort were also assessed for MRD using MPFC. CD34 positive cells comprised 0.05-12.5% of autograft specimen mononuclear cells. Due to immunophenotypic changes likely attributable to GCSF mobilization, and without leukemia associated immunophenotypes from diagnosis available, MPFC was unable to identify MRD in any of 40 patients tested.In summary, no single MRD test on autografts could completely predict post-HCT AML relapse. Auto-HCT presents unique challenges for AML MRD testing due to masking effects of GCSF on MPFC and RQ-PCR gene expression signatures. Additionally, detection of any AML autograft contamination must be extremely sensitive to be useful for predicting relapse given the absence of any potentially protective graft versus leukemia effect following an auto-HCT. Here we show combinations of molecular MRD assays can overcome some, but not all, of these limitations (Figure). [Display omitted] DisclosuresRadich:Incyte: Consultancy; Ariad: Consultancy; Novartis: Consultancy; Novartis: Other: Lab Contract. Andreadis:Cellerant: Consultancy; Novartis: Consultancy; McGraw Hill: Other: Publishers; Pharmacyclics: Honoraria. Damon:Sunesis: Research Funding; McGraw Hill: Other: Chapter Royalties; Atara: Consultancy; Sigms Tau: Research Funding. Logan:Jazz Pharmaceuticals: Consultancy; Amgen: Consultancy; Pharmacyclics: Consultancy. Martin:Sanofi: Consultancy.

Both autologous bone marrow mononuclear cell and peripheral blood progenitor cell therapies similarly improve ischaemia in patients with diabetic foot in comparison with control treatment

The aim of our study was to compare the effect of bone marrow mononuclear cell and peripheral blood progenitor cell therapies in patients with diabetic foot disease and critical limb ischaemia unresponsive to revascularization with conservative therapy. Twenty-eight patients with diabetic foot disease (17 treated by bone marrow cells and 11 by peripheral blood cell) were included into an active group and 22 patients into a control group without cell treatment. Transcutaneous oxygen pressure and rate of major amputation, as the main outcome measures, were compared between bone marrow cells, peripheral blood cell and control groups over 6 months; both cell therapy methods were also compared by the characteristics of cell suspensions. Possible adverse events were evaluated by changes of serum levels of angiogenic cytokines and retinal fundoscopic examination. The transcutaneous oxygen pressure increased significantly (p < 0.05) compared with baseline in both active groups after 6 months, with no significant differences between bone marrow cells and peripheral blood cell groups; however, no change of transcutaneous oxygen pressure in the control group was observed. The rate of major amputation by 6 months was significantly lower in the active cell therapy group compared with that in the control group (11.1% vs. 50%, p = 0.0032), with no difference between bone marrow cells and peripheral blood cell. A number of injected CD34+ cells and serum levels of angiogenic cytokines after treatment did not significantly differ between bone marrow cells and peripheral blood cell. Our study showed a superior benefit of bone marrow cells and peripheral blood cell treatments of critical limb ischaemia in patients with diabetic foot disease when compared with conservative therapy. There was no difference between both cell therapy groups, and no patient demonstrated signs of systemic vasculogenesis.

Defining “poor mobilizer” in pediatric patients who need an autologous peripheral blood progenitor cell transplantation

The definition of poor mobilizers is not clear in pediatric patients undergoing autologous peripheral blood hematopoietic progenitor cell (HPC) mobilization. Most studies conducted in children define those variables related to the collection of HPC after leukapheresis, but the information regarding exclusively the mobilization process is scarce. In our experience, most children (92.2%) reach the target CD34(+) cell dose for autologous peripheral blood progenitor cell transplantation ifCD34(+) cell count was higher than 10/μL. No differences were observed between those with >20 CD34(+) cells/μL and 11-20 CD34(+) cells/μL. In this study, we analyzed the variables that influence CD34(+) cell count; we found that prior use of radiotherapy was the main variable related to poor mobilization. Patients diagnosed with Ewing sarcoma, treated with radiotherapy and mobilized with standard doses of granulocyte colony-stimulating factor (G-CSF) were also at a high risk of mobilization failure. In these patients, we should consider mobilization with high dose G-CSF and be prepared with new mobilization agents to avoid delay on their course of chemotherapy.

Autologous Peripheral Blood Progenitor Cells Cryopreserved in 7.5% DMSO Give Faster Hematopoietic Reconstitution Than Cells Cryopreserved in 10% DMSO.

AbstractAbstract 3018 Background:Cryopreservation of autologous peripheral blood progenitor cells (PBPCs) in 10% DMSO is a routine in most transplantation centers. During ASH 2011 Meeting, we presented the results of in vitro research, concerning the optimization of DMSO concentrations for recovery and clonogeneic potential of PBPCs after thawing. We concluded, that reduction of DMSO concentration from 10% to 7.5% may have favorable impact for cell clonogeneicity. Accordingly, we implemented new cryoprotective mixture (7.5% instead of 10% DMSO) into clinical practice. The purpose of this study was to clinically evaluate the changed protocol. Patients and Methods:In our department, between Jan 2012-Aug 2012, 56 patients were transplanted with autologous PBPCs cryopreserved in 7.5% DMSO solution (median of age: 57 years, range: 21–66). We compared the data concerning hematopoietic engraftment and the frequency of side effects with historical control – 52 subsequent patients treated with transplantation of PBPSc cells cryopreserved in 10% DMSO (median of age: 57 years, range: 21–66) in a preceding period. Both study groups did not differ significantly with regard to the diagnosis (mostly lymphoproliferative disorders) or disease status at transplantation. As well, the number of transplanted CD34+ cells was comparable: median 6.5′106/kg (range 1.5–24.7) for 7.5% DMSO and 7.5′106/kg (2.1–24.6) for 10% DMSO group, p=0.68. All received G-CSF (filgrastim) starting on day +7 after transplantation. Results:The volume of infused DMSO was significantly lower in patients who obtained PBPBc cryopreserved in 7.5% DMSO (median 22.5 ml, range 7.5–45) than 10% DMSO (median: 30 ml, range, 10–160); p=0.02. The time to leukocyte recovery >1′109/L was faster for 7.5% DMSO (median: 11 days, range: 9–12) than in 10% DMSO (median 11 days, range: 10–13), p=0.03. Similarly, reconstitution of neutrophils >0.5′109/L was faster for 7.5% DMSO group: median 11 days (range 9–13 days) vs. 11 days (range 10–13 days), respectively; p = 0.04. We didn’t observe significant difference with regard to platelet recovery >50′109/L (median 12 days, range: 0–21 days for 7.5% DMSO vs. median 12.5, range: 0–19 days for 10% DMSO). Hospital stay since HSCT was shorter in case of 7.5% group (median: 14 days, range: 11–21) than 10% group (median: 15 days, range: 13–25); p=0.04. Number of RBC and platelets transfusions as well as transfusion-related complications did not differ between the groups. Adverse events after transplantation were mild and transient, usually grade 1 nausea, and occurred in 20 (38%) patients in 10% DMSO group compared to 24 (43%) in 7.5% DMSO group (p=0.7). Conclusions:The analysis of newly implemented cryopreservation protocol suggest that reduction of the DMSO concentration from 10% to 7.5% is associated with faster leukocyte and neutrophil recovery as well as shorter hospital stay. These findings require verification in a prospective, randomized trial. [Display omitted] Disclosures:No relevant conflicts of interest to declare.

Genetic variant of the G-CSF receptor gene is associated with lower mobilization potential and slower recovery of granulocytes after transplantation of autologous peripheral blood progenitor cells

Peripheral blood mobilized by cytokines (i.e. granulocyte colony stimulating factor, G-CSF) and chemotherapy has become a major source of hematopoietic stem and progenitor cells for transplantation (PBPCT). In this study the effect of the G-CSF receptor (CSF3R) gene polymorphism was investigated. The presence of the CSF3R variant (T allele, rs3917924) was related to CD34+ mobilization yield and the pace of granulocyte recovery after autologous PBPCT. The mobilization yield was higher in patients lacking the CSF3R variant (OR=4.756, p=0.046) and those with multiple myeloma (OR=10.534, p=0.019). The pace of granulocyte recovery was found to be associated with the CSF3R polymorphism and was significantly slower in patients carrying the CSF3R-T variant than in CC homozygotes (median of 17 vs. 13days, p<0.001). This association was confirmed (OR=4.445, p=0.014) by multiple regression analysis considering patient age and sex, the number of transplanted CD34+ cells, diagnosis and CSF3R polymorphism. These results imply that CSF3R gene polymorphism plays a significant role in PBPCT.

Stem Cells in Arthroscopy

Stem Cells in Arthroscopy

Successful collection and engraftment of autologous peripheral blood progenitor cells in poorly mobilized patients receiving high‐dose granulocyte colony‐stimulating factor

Granulocyte Colony-Stimulating Factor (G-CSF) alone or in conjunction with chemotherapy is commonly used to mobilize hematopoietic progenitor cells (HPC) into peripheral blood for progenitor cell harvest for autologous HPC transplantation. However, in up to 30% of patients, HPC are not effectively mobilized. In this study, we report the efficacy and safety profiles of a mobilization strategy using high-dose (up to 36 μg/kg) G-CSF in poorly mobilized patients. Retrospective medical record reviews were performed for 392 patients who underwent autologous peripheral blood progenitor cell collection. A total of 56 patients were given high-dose G-CSF due to very ineffective mobilization and 35 of these patients underwent autologous HPC transplantation. The efficacy of mobilization, apheresis collection, and infusion were reviewed and analyzed. More than 2.5 × 10(6) CD34/Kg were collected in 88% of patients (49 of 56) who were placed on high-dose G-CSF due to very ineffective mobilization. Of the 35 patients who underwent HPC transplantation using the progenitor cells that were mobilized with high-dose G-CSF due to very ineffective mobilization, all had rapid and complete neutrophil and platelet engraftment comparable with good mobilizers. We conclude that collection of HPC using hyperstimulation with G-CSF is an effective alternative approach for HPC harvest for poorly mobilized patients.

Romiplostim After Autologous Peripheral Blood Progenitor Cell Transplantation: Results of a Pilot Study

Romiplostim After Autologous Peripheral Blood Progenitor Cell Transplantation: Results of a Pilot Study

Impact of Constitutional Polymorphisms in VCAM1, CD44, CXCL12, CXCR4 and CSF3R in Progenitor Cell Mobilization in Patients with Hematological Malignancies

Abstract 2997Introduction. The identification of genetic variants predictive of response to G-CSF mobilization might be useful in deciding the best strategy to obtain haematopoietic progenitors (HP) in patients scheduled for autologous peripheral blood progenitor cell transplant. Recently, one study has demonstrated the relationship among polymorphisms in genes implicated in trafficking and homing of CD34+ cells and the degree of mobilization after G-CSF therapy among healthy donors (Haematologica 2011; 96: 102–109). Aims. To evaluate if polymorphisms in five genes (CD44 rs13347 C>T, CSF3R rs3917924 A>G, CXCR4 rs2680880 A>T, CXCL12 rs1801157 G>A, and VCAM1 rs1041163 T>C) previously associated with the number of G-CSF mobilized CD34+ cells in healthy donors, can also predict the mobilization efficacy in a group of patients with hematological malignancies. Patients and Methods. We retrospectively evaluated 183 patients who were treated with s.c. G-CSF at 10 mcg/kg during 4 days. HP collection was initiated or not at day 5 according to the CD34+ number in peripheral blood (PB). Patients were selected among two groups: (1) poor mobilizers (n=109), who failed a mobilization attempt, presenting with <10 CD34+cells/mcl of PB, and (2) good mobilizers (n=74), those achieving >2 ×106CD34+ cells/kg in a first and only apheresis. The genetic variants were genotyped by allelic discrimination polymerase chain reaction (PCR) assays using TaqMan®Genotyping Assays (Applied Biosystems). Results. Patients diagnosed with lymphoma, myeloma and acute leukaemia were 40%, 38% and 21% of poor mobilizers, and 38%, 46% and 16% of good mobilizers, respectively. On the overall group, the genetic variant TT rs1801157 in CXCL12 was significantly associated with poor mobilization (p=0.040). Among lymphoma patients, the presence of the C allele in VCAM1 was significantly associated with mobilization rate (49% vs 19% among poor and good mobilizers respectively, p=0.011). In this lymphoma group, a trend towards poor mobilization was also observed in relation with homocygosis for the T allele of CXCL12 (12% vs 0% in poor and good mobilizers, respectively, p=0.066). The analyzed variables had no impact on the mobilization capacity in patients with myeloma or acute leukaemia. Discussion. Genetic variants in VCAM and CXCL12 seem to be related with the mobilization yield after G-CSF, particularly in lymphoma patients. Other polymorphisms in adhesion molecules related to the degree of CD34+ cell mobilization in healthy donors have not shown a relevant role in patients with hematological malignancies, probably reflecting the predominant effect of disease biology and/or of previous treatments.Funding. This study was supported in part by a research grants 04515/GERM '2f 06; RECAVA RD06/0014/0039, and FIS 10/02594 Disclosures:No relevant conflicts of interest to declare.

Treatment of Myelofibrosis Using I.V. Treosulfan and Autologous Peripheral Blood Progenitor Cell Transplantation Is Feasible and Results in Hematologic Responses – Long-Term Follow-up and Final Analysis of a One-Armed Clinical Study and of Individual Cases

Abstract 4514 Introduction:Myelofibrosis with myeloid metaplasia is a chronic myeloproliferative disorder. The only curative therapy to date is myeloablation followed by allogeneic transplantation. Autologous transplantation after myeloablation with high-dose oral busulfan provides a palliative approach which might lead to a long-term relief of symptoms of the disease and is associated with acceptable morbidity and mortality (Anderson et al., Blood, 2001; 98:586–593). However, busulfan pharmacokinetics after oral administration can vary among patients. On the other hand treosulfan, a bi-functional alkylating drug, can be administered safely i.v. with reliable pharmacokinetics and might be suitable for conditioning of patients with myelofibrosis. Methods and Results:In a pilot phase, we treated three patients on an individual basis with an autologous transplantation after myeloablation with treosulfan. Following the encouraging results we initiated a clinical trial. Included were patients with myelofibrosis and a high risk disease according to the 1997 criteria of Cervantes. The study is registered at www.kompetenznetz-leukaemie.de.As of today we have transplanted 11 patients according to the protocol, 9 in Heidelberg, 1 at the Diakoniekrankenhaus Rotenburg and 1 at the University Hospital Rostock. Age at transplantation ranged from 50 to 69 years. For 10 of 11 patients we have been successful in collecting more than 5×106 CD34+ cells/kg body weight (one patient with 4,9 ×106) after mobilization with G-CSF at 16 μg/kg BW s.c. daily for 4 days. The conditioning regimen was treosulfan for three days (total dose 42 g/m2). Hematologic reconstitution was considerably protracted with a median of 26 days (leucocyte count > 1/nl) compared to autologous transplantation for other indications. One patient died on day 34 post transplantation after cerebral hemorrhage. He suffered from severe transfusion refractory thrombocytopenia already before the transplantation.The twelve-month follow-up showed symptomatic relief, mostly as freedom from transfusions in 7 of 11 patients. The two survivors are alive after 9 years, one with allogeneic transplantation 2 years and the other with allogeneic transplantation 8 years after autologous transplantation. The median survival of the other 9 patients was 3.5 years after autologous transplantation (0.1 – 7.5 years). Conclusions:We conclude that myeloablation with treosulfan and autologous PBPCT for patients with myelofibrosis can provide significant palliation. It is associated with an acceptable morbidity and mortality, taking into account the high-risk nature of the disease. The approach is feasible for high-risk patients that cannot or chose not to undergo allogeneic transplantation. Disclosures:Buss:medac GmbH: Research Funding. Off Label Use: Treosulfan, off-label use as an agent for high-dose chemotherapy with subsequent autologous stem cell transplantation. Topaly:medac GmbH: Research Funding. Fruehauf:medac GmbH: Research Funding.

Hematologic Changes After Total Body Irradiation and Autologous Transplantation of Hematopoietic Peripheral Blood Progenitor Cells in Dogs With Lymphoma

Dogs with and without lymphoma have undergone hematopoietic cell transplantation in a research setting for decades. North Carolina State University is currently treating dogs with B- and T-cell lymphoma in a clinical setting with autologous peripheral blood progenitor cell transplants, using peripheral blood CD34+ progenitor cells harvested using an apheresis machine. Complete blood counts were performed daily for 15 to 19 days posttransplantation to monitor peripheral blood cell nadirs and subsequent CD34+ cell engraftment. This study documents the hematologic toxicities of total body irradiation in 10 dogs and the subsequent recovery of the affected cell lines after peripheral blood progenitor cell transplant, indicating successful CD34+ engraftment. All peripheral blood cell lines, excluding red blood cells, experienced grade 4 toxicities. All dogs had ≥ 500 neutrophils/μl by day 12, while thrombocytopenia persisted for many weeks. All dogs were clinically normal at discharge.

High-dose Busulfan and Cyclophosphamide as a Conditioning Regimen for Autologous Peripheral Blood Stem Cell Transplantation in Childhood Non-Hodgkin Lymphoma Patients

We analyzed the outcome in 22 children with refractory or relapsed non-Hodgkin lymphoma who underwent autologous peripheral blood progenitor cell transplantation between 1994 and 2009. The conditioning regimen used in all patients consisted of busulfan and cyclophosphamide. Median age was 6 years (range 2 to 16 y). The most common histologic subtype was Burkitt lymphoma. Ten patients were in complete remission and 12 in partial remission at the time of transplant. The median dose of CD34+ cells that was infused was 4.6 × 10/kg (range 2.1 to 58.7 × 10/kg). All the patients were engrafted, with a median time for neutrophils and platelets recovery of 11 (range, 8 to 15 d) and 14 (range, 9 to 60 d) days, respectively. Nonhematologic treatment-related toxicity included severe mucositis in 3 patients and hepatic sinusoidal obstruction syndrome in 1 patient. There were no transplant-related mortalities. With a median follow-up of 60 months (range, 4 to 180 d) the disease-free survival was 90 ± 6.5% for the whole group. This retrospective study shows a high long-term survival using busulfan/cyclophosphamide as conditioning regimen in children with refractory or relapsed non-Hodgkin lymphoma.

Articular Cartilage Regeneration With Autologous Peripheral Blood Progenitor Cells and Hyaluronic Acid After Arthroscopic Subchondral Drilling: A Report of 5 Cases With Histology

The purpose of this study was to evaluate the quality of articular cartilage regeneration after arthroscopic subchondral drilling followed by postoperative intraarticular injections of autologous peripheral blood progenitor cells (PBPCs) in combination with hyaluronic acid (HA). Five patients underwent second-look arthroscopy with chondral core biopsy. These 5 patients are part of a larger pilot study in which 180 patients with International Cartilage Repair Society grade III and IV lesions of the knee joint underwent arthroscopic subchondral drilling followed by postoperative intra-articular injections. Continuous passive motion was used on the operated knee 2 hours per day for 4 weeks. Partial weight bearing was observed for the first 6 to 8 weeks. Autologous PBPCs were harvested 1 week after surgery. One week after surgery, 8 mL of the harvested PBPCs in combination with 2 mL of HA was injected intra-articularly into the operated knee. The remaining PBPCs were divided into vials and cryopreserved. A total of 5 weekly intra-articular injections were given. Second-look arthroscopy confirmed articular cartilage regeneration, and histologic sections showed features of hyaline cartilage. Apart from the minimal discomfort of PBPC harvesting and localized pain associated with the intra-articular injections, there were no other notable adverse reactions. Articular hyaline cartilage regeneration is possible with arthroscopic subchondral drilling followed by postoperative intraarticular injections of autologous PBPCs in combination with HA. Level IV, therapeutic case series.