G-CSF Administration Research Articles

Autophagy-dependent TIGIT+ Treg are critical for the maintenance of tolerance

Abstract Regulatory T cells (Treg) play a crucial role in the maintenance of peripheral tolerance. Quantitative and/or qualitative defects in Treg result in diseases such as autoimmunity and graft-versus-host disease (GVHD), a serious complication of bone marrow transplantation (BMT). We recently reported that G-CSF mobilisation of stem cells in donor mice increases autophagy related genes in Treg and it is associated with an improved GVHD outcome after BMT. Autophagy is a self-degradative process for cytosolic components which promotes cell homeostasis and survival. Here, we demonstrate that the lineage-specific disruption of autophagy within FoxP3+ Treg (Atg7fl/fl.FoxP3cre+ mice) resulted in a profound loss of Treg within the bone marrow (BM). This resulted in dysregulated effector T cell activation and expansion, and in aged mice, the development of enterocolitis and severe scleroderma. We show that the BM compartment is highly enriched in TIGIT+ Treg and that this subset is differentially lost in the absence of autophagy. Moreover, G-CSF administration to naive mice results in the mobilization of BM TIGIT+ Treg explaining the autophagy signature of peripheral Treg in G-CSF treated mice. Importantly, lethally irradiated B6D2F1 recipients of splenic grafts from G-CSF mobilized B6.Atg7fl/fl.FoxP3cre+ donors failed to reconstitute the Treg compartment within the BM. This resulted in GVHD exacerbation and reduced survival compared to recipients of B6.WT.FoxP3cre+ grafts. Collectively, these data indicate that autophagy-dependent TIGIT+ Treg in the BM are critical for the maintenance of tolerance and to control GVHD; thus the induction of autophagy may represent an attractive immune-restorative therapeutic strategy.

A schematic case episode format to aid in “Choosing Wisely."

181 Background: The recommendations of the Choosing Wisely campaign are evidence –based strategies to reduce cost without sacrificing outcomes. Yet tying the recommendations to indicators of use at the physician and case level has been challenging. As practices become responsible for total cost of care, an easy to use analytic method to determine appropriate use will be critical. We here describe a tool for rapid assessment of individual cases to achieve that objective. Methods: The population was a payer defined cohort of lung cancer patients treated at Texas Oncology (TxO). TxO maintains a payer patient list, updated daily with demographics and diagnosis details for all enrolled patients. The list of lung cancer patients was cross-referenced against billing data from TxO's financial data warehouse (FDW). The FDW data is generated monthly, based on billing details from TxO's practice management system, and includes procedure codes and dates of service. Radiation therapy, chemotherapy, and GCSF administrations for each enrolled lung cancer patient were identified in the FDW data based on CPT codes. SAS software (version 9.4 for Windows) was used to generate a time series plot for each patient, based on date of service for each procedure. The time series plots were inserted into an Excel report template, along with general patient information, using a Visual Basic script, for review by TxO's medical director and quality committees. Results: Each patient-specific time series schematic displays elapsed weeks on the x-axis, beginning with week 1 to end of treatment. Three variables are displayed on the y-axis, using distinct colors and symbols: dates of radiation therapy (orange #), dates of GCSF administration (red x), and dates of chemotherapy administration (green ^). Each of the three y-axis variables is assigned a constant value that is plotted along a straight line. A graphic representation for a patient with stage III lung cancer could look as shown in the Table. Conclusions: Treatment episodes can be distilled into a meaningful format that allows rapid case review and the opportunity for continuous learning. Additional diseases and graphics will be available for presentation. [Table: see text]

Administration of G-CSF from day +6 post-allogeneic hematopoietic stem cell transplantation in children and adolescents accelerates neutrophil engraftment but does not appear to have an impact on cost savings.

G-CSF post-allogeneic HSCT accelerates neutrophil engraftment, but evidence that it impacts on cost-related outcomes is lacking. We performed a retrospective child and adolescent single-center cohort study examining G-CSF administration from Day +6 of allogeneic HSCT vs. ad hoc G-CSF use where clinically indicated. Forty consecutive children and adolescents undergoing allogeneic HSCT were included. End-points were as follows: time to engraftment; incidence of acute and chronic GvHD; number of patients alive at Day +100; 180-day TRM; post-transplant days in hospital; and cost of antimicrobials, TPN, and G-CSF usage. Neutrophil engraftment occurred earlier in the group that received G-CSF from Day +6. There was no difference between groups in any of the other end-points with the following exception: the cost of GCSF was significantly higher in the D + 6 G-CSF group. However, median G-CSF cost in this group amounted to only €280. There was a trend towards reduced cost of antimicrobials in the D + 6 G-CSF group, although this did not reach significance (p = 0.13). The median cost per patient of antimicrobial agents between groups differed by €1116. This study demonstrated the administration of G-CSF on Day +6 in pediatric HSCT to be safe. A further study using a larger cohort of patients is warranted to ascertain its true clinico-economic value.

G-CSF Administration after the Intraosseous Infusion of Hypertonic Hydroxyethyl Starches Accelerating Wound Healing Combined with Hemorrhagic Shock.

Objective. To evaluate the therapeutic effects of G-CSF administration after intraosseous (IO) resuscitation in hemorrhagic shock (HS) combined with cutaneous injury rats. Methods. The rats were randomly divided into four groups: (1) HS with resuscitation (blank), (2) HS with resuscitation + G-CSF (G-CSF, 200 μg/kg body weight, subcutaneous injection), (3) HS with resuscitation + normal saline solution injection (normal saline), and (4) HS + G-CSF injection without resuscitation (Unres/G-CSF). To estimate the treatment effects, the vital signs of alteration were first evaluated, and then wound closure rates and homing of MSCs and EPCs to the wound skins and vasculogenesis were measured. Besides, inflammation and vasculogenesis related mRNA expressions were also examined. Results. IO infusion hypertonic hydroxyethyl starch (HHES) exhibited beneficial volume expansion roles and G-CSF administration accelerated wound healing 3 days ahead of other groups under hemorrhagic shock. Circulating and the homing of MSCs and EPCs at wound skins were significantly elevated at 6 h after G-CSF treatment. Inflammation was declined since 3 d while angiogenesis was more obvious in G-CSF treated group on day 9. Conclusions. These results suggested that the synergistical application of HHES and G-CSF has life-saving effects and is beneficial for improving wound healing in HS combined with cutaneous injury rats.

Mobilization of CD8+ Central Memory T-Cells with Enhanced Reconstitution Potential in Mice By a Combination of G-CSF and GMI-1271-Mediated E-Selectin Blockade

T-cells are critical mediators of immune defense against pathogens and cancer. Adoptive T cell immunotherapy and T-cell engineering have promising clinical applications but T cell survival and exhaustion are current limitations. Central memory cells (TCM CD62L+ CCR7+) and their precursors, stem central memory T-cells (TSCM) possess the stem-like properties needed to reconstitute and prolong an effective immune response long-term. These cells have been shown to significantly improve therapeutic efficacy of adoptive T-cell therapy. The challenge remains to harvest good quality TCM-cells for these immunotherapy approaches.The bone marrow (BM) is the major reservoir of CD8+ TCM and their precursors. We have previously shown that E-selectin is expressed in the BM vasculature and drives activation and differentiation of hematopoietic stem cells during G-CSF induced mobilization to the blood. We find therapeutic blockade of E-selectin promotes HSC self-renewal and reconstitution in vivo. We now examine the impact of E-selectin blockade on CD8+ T cell mobilization from the bone marrow to the blood and hypothesize that E-selectin blockade may also dampen the activation/differentiation of this subset.First we administered a standard G-CSF regime (filgastim 250ug/kg/day for 3 days) to mice and then dosed some cohorts with GMI-1271 (40mg/kg BID) from 12 to 72 hours within this 3 day period. Administration of G-CSF alone results in a near complete disappearance of bone marrow resident CD8+ TCM cells, and their apparent migration (increase in numbers) to the blood, while CD8+ subsets in the lymph nodes and spleen were barely affected by G-CSF. Furthermore among T-cell subsets, CD8+ but not CD4+ TCM were specifically mobilized into the blood when GMI-1271 was co-administered for the last 12 to 24 hours of G-CSF. These findings are consistent with reports demonstrating the bone marrow to be a major reservoir for CD8+ but not CD4+ central memory T-cells.Administration of GMI-1271 caused a marked enhancement in mobilization into the blood of CD8+ TCM/SCM (CD62Lhi, CCR7+) cells over treatment with G-CSF alone (p<0.05). To determine the functional consequences of this skewed mobilization following GMI-1271 co-administration, 25 uL of mobilized blood was transplanted into irradiated congenic B6.SJL recipients together with 2x105 congenic BM cells to analyze long-term donor T-cell engraftment in the recipient mice. We found G-CSF mobilized donor blood did not contribute CD8+ TCM cells that can persist post-transplant (<0.5% at 20 weeks post-transplant). In contrast when donor mice were mobilized with G-CSF together with E-selectin blockade (GMI-1271), we found elevated levels of donor blood derived CD8+ T-cells demonstrating robust long-term CD8+ T-cell persistence / regeneration (5.3 ±3.2% of total recipient T-cells, p=0.04). This dramatic boost in donor CD8+ T-cell reconstitution in mobilized blood following GMI-1271 co-administration is likely to be due to the long-term persistence and in vivo amplification of CD8+ TCM cells from donor mobilized blood. Similar in vivo enhancing effects of GMI-1271 were also observed with other mobilizing agents such as combined CXCR4 and VLA-4 blockade and GM-CSF resulting in a significant 4.9-fold boost in donor CD8+ reconstitution with GMI-1271. Importantly, only 12 hours of E-selectin blockade was sufficient to achieve this boost in CD8+ TCM numbers in the blood following G-CSF.In a previous report we have shown that therapeutic blockade of E-selectin promotes HSC self-renewal in vivo. Thus, it is possible that E-selectin blockade boosts mobilization of CD8+ TCM/SCM with stem-like properties into the blood by loosening factors retaining CD8+ TCM/SCM in the bone marrow and/or blocking the E-selectin-mediated activation and differentiation of this T-cell subset.In summary, our studies identify E-selectin blockade as a novel target to improve harvesting of CD8+ TCM/SCM cells with stem-like properties. Blockade of this target with GMI-1271 significantly improves the in vivo reconstitution potential and regenerative properties of CD8+ T-cells from donor blood allowing a valuable source of desired T-cells for use in adoptive immunotherapy and T-cell engineering. DisclosuresWinkler:GlycoMimetics Inc: Research Funding. Barbier:GlycoMimetics Inc: Research Funding. Davies:GlycoMimetics Inc: Research Funding. Smith:GlycoMimetics, Inc.: Employment. Fogler:GlycoMimetics, Inc.: Employment. Magnani:GlycoMimetics Inc: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees.

A Novel and Pivotal Role of the Mannose-Binding Lectin (MBL) Pathway of Complement Cascade (ComC) Activation in Triggering Mobilization of Hematopoietic Stem/Progenitor Cells (HSPCs)

Background. The complement cascade (ComC) is part of the innate immunity system, which is not adaptable and does not change over the course of an individual's lifetime; however, it can be recruited and brought into action by the adaptive immune system. The ComC has several pleiotropic effects, and, as we have previously demonstrated, it is required for mobilization of HSPCs during infection or tissue/organ injuries and responding to pharmacological mobilizing agents (Blood 2004, 103, 2071-2078). The ComC is activated by three pathways: the classical, alternative, and mannose-binding lectin (MBL) pathways. While a requirement for ComC activation and, in particular, the pivotal role of the distal part of complement activation and generation of C5 cleavage fragments was previously demonstrated by us (Leukemia 2009, 23, 2052-2062), mice with mutations to components of the classical and alternative pathways in which the distal pathway of C5 activation remained intact did not show impairment of HSPC mobilization (Leukemia 2010, 24, 1667-1675). However, no studies so far have been performed to address the role of the MBL pathway of ComC activation in triggering the mobilization of HSPCs. The MBL pathway is homologous to the classical pathway, but contains opsonin, MBL, and ficolins instead of C1q. MBL functions by pattern recognition, as opsonin binds to mannose residues on the surface of pathogens and certain cells, and activates the MBL-associated serine proteases, MASP-1, and MASP-2, which can then split C4 (into C4a and C4b) and C2 (into C2a and C2b) to form the classical C3-convertase, as in the classical pathway. Interestingly, it is known that ~10% of the population has defective activation of the MBL pathway.Hypothesis. We hypothesized for first time that the MBL ComC-activation pathway is involved in triggering ComC-mediated mobilization of HSPCs and that MBL deficiency results in poor mobilization.Materials and Methods. In our experiments, 2-month-old, MBL-deficient mice (MBL-/-) and normal wild type (WT) littermates were mobilized for 6 days with G-CSF or AMD3100. Following mobilization, we measured in peripheral blood (PB) i) the total number of white blood cells (WBC), ii) the number of circulating clonogenic colony-forming unit granulocyte/macrophage (CFU-GM) progenitors, and iii) the number of Sca-1+ c-kit+ lineage- (SKL) cells. In parallel, we evaluated activation of the MBL pathway in WT animals after administration of G-CSF and AMD3100.Results. We found that pattern recognition by the MBL ComC activation pathway is involved in pharmacological G-CSF- and AMD3100-induced mobilization of HSPCs, and activation of the MBL pathway was confirmed by ELISA in WT animals. As predicted, MBL KO mice were found to be poor mobilizers.Conclusions. We identified a previously unrecognized role of the MBL pathway in triggering ComC activation in the process of HSPC mobilization. This finding explains the pivotal role of the MBL pathway in triggering activation of the proximal part of the ComC and explains why, even with a deficiency in activation of classical and alternative pathway components, mobilization of HSPCs proceeds normally as long as the MBL pathway is intact. On the other hand, if the MBL pathway of the ComC is defective, neither classical nor alternative pathways can trigger optimal mobilization of HSPCs. Taking into consideration that ~10% of normal people are poor activators of the MBL pathway, we are currently investigating whether MBL deficiency correlates with poor mobilization in these patients. DisclosuresNo relevant conflicts of interest to declare.

Hypogammaglobulinaemia and Infectious Toxicity Related with the Use of Maintenance Strategies with Rituximab in B Lymphomas

IntroductionThe use of maintenance treatment with rituximab in B lymphomas has improved the results in terms of progression free survival in follicular lymphoma (PRIMA trial). This improvement has been seen in other types of indolent lymphomas including chronic lymphatic leukemia or specific cases of aggressive lymphomas with or without low grade associated component. However, the rituximab maintenance, even though is a well-tolerated strategy, is not free of toxicity. Our objective is reporting the rituximab toxicity associated to the maintenance with rituximab in a series of B-cell lymphomas treated with immunochemotherapy in first line in Hospital Son Espases (HUSE) and Hospital del Mar (HM).Methods and materialsTo avoid selection bias, we retrospectively identified from the Pharmacy registries of HUSE and HM those patients who had been treated with rituximab maintenance between 2002 and 2014 in first line of induction treatment with conventional immunotherapy or immunochemotherapy. Second lines or more intensive treatments were excluded. We obtained information about diagnosis, prognostic and outcome in all the patients, including levels of immunoglobulins as well as the seriousness of infections or other related side effects. Polymorphisms FcgRIIIa were analyzed using RT-PCR and sequencing.ResultsWith the previous criteria 82 patients were included. Table 1 shows the diagnostic characteristics and the treatment of the patients. The median of age was 65 years; most cases were follicular lymphoma treated with R-CHOP or R-B and receiving maintenance with rituximab every 2 months (66%). Table 2 describes toxicity related to rituximab maintenance: hypogammaglobulinaemia IgG, IgA or both in the 27%, 21% and 11% of the cases respectively. When we analyzed the role of polymorphisms of FcgRIIIa we found that the patients with homozigous polymorphism VV were exempt of hypogammaglobulinaemia IgG or IgA. Moreover the polymorphism FF was associated meaningfully with a greater incidence of hypogammaglobulinaemia IgA (p=0.022). Only the 8% of the cases of hypogammaglobulinaemia needed treatment with intravenous immunoglobulins and only 6% had grade 3-4 infections. 21% of the patients treated with rituximab maintenance developed grade 3-4 neutropenia, generally reversible with the administration of G-CSF and without serious infections. Increase of mortality was not observed related with the presence of hypogamaglobulinaemia IgG (p=0.34), IgA (p=0.59) or both (p=0.59) or grade 3-4 neutropenia secondary to rituximab (p=0.38).ConclusionsBetween one-quarter or one-fifth of the patients who received maintenance therapy with rituximab during 2 years associate hypogammaglobulinaemia IgG, IgA and/or neutropenia as toxicity. Nevertheless these complications are generally mild, without related severe infections or increase of mortality.Table 1Clinical characteristics of the seriesMedian of age65 (27-85)Sex41 (50%) / 41 (50%)ECOG PS >17 (9%)Type of lymphoma:- Folicular lymphoma- DLBCL- CLL/SLL- Marginal lymphoma- Mantel cell lymphoma- Other46 (56%)13 (16%)8 (10%)6 (7%)5 (6%)4 (5%)Stage III-IV71 (87%)B symptoms29 (35%)Induction treatment:- R-CHOP/R-CVP- R-B- Rituximab alone- Schema with Fludarabine-R- Intensive schemas- Others50 (61%)13 (16%)9 (11%)4 (5%)3 (4%)3 (4%)Type of maintenance (2 years):- Every 2 months- Every 3 months- Every 6 months (4 weeks)54 (66%)15 (18%)1 3 (16%)Table 2Rituximab maintenance attributable toxicityPolimorphism FcgRIIIAType of toxicityTotalFFFVVVpHypogammaglobulinaemia- IgG- IgA- Double IgG + IgA22 (27%)17 (21%)9 (11%)7 (23%)11 (37%)3 (10%)15 (31%)6 (12%)6 (12%)0 (0%)0 (0%)0 (0%)0.340.0220.74Treatment with immunoglobulins iv7 (8%)3 (10%)4 (8%)0 (0%)0.79Grade 3-4 infections:5 (6%)2 (7%)3 (6%)0 (0%)0.87Grade 3-4 neutropenia:17 (21%)8 (27%)8 (17%)1 (25%)0.59 DisclosuresNo relevant conflicts of interest to declare.

Azacitidine with Oral Cytarabine and Etoposide Induces Immediate Hematological and Cytogenetic Response in Higher Risk Myelodysplastic Syndromes Even in Single Azacitidine Resistance

Background: Although azacitidine (AZA) induces high hematological response and delayed progression to acute myeloid leukemia (AML) in patients with myelodysplastic syndromes (MDSs), its cytotoxic and cytogenetic effects are relatively poor. Moreover, the prognosis of patients at very high risk in the revised international prognostic scoring system (IPSS-R), with poor cytogenetics including complex abnormalities and monosomy 7, and with AZA resistance is extremely poor. Therefore, AZA-based combination therapy with low toxicity is warranted. In the past, effective low-dose etoposide (VP16) and cytarabine ocfosphate (SPAC) combination therapy for refractory MDS and AML was reported. Here, we report 10 higher-risk MDS and AML patients treated with AZA in combination with oral VP16 and SPAC.Patients and methods: AZA (75 mg/m2 daily for 7 days) in combination with SPAC (300 mg daily for 14 days) and VP16 (25 mg daily for 14 days) was administered to 10 patients (median age, 80 years; range, 48 to 90) with newly diagnosed (n=2) and previously AZA-treated (n=8) MDS and AML between December 2013 and April 2015. The diagnoses were RCMD (n=2), RAEB1 (n=2), RAEB2 (n=1), CMML (n=1), and AML-MRC (n=4). IPSS-R categories were low (n=1), intermediate (n=3), high (n=2), and very high (n=4). In 8 AZA-treated patients, its median cycle before this study was 11 (1-30). Results: In the pretreatment period, neutrophil count, platelet count, hemoglobin level, bone marrow blast rate, and WT1 were 805/µl (170-2700), 53.0 (25-222) × 109/l, 8.4 g/dl (7.2-13.5), 6.5% (1.4-30.0), and 1200 (200-31,000) copies/µg RNA, respectively. Five out of 10 had complex karyotype abnormalities. After 1 course of treatment, the overall response rate was 100%. The rate of any hematological improvement (HI) was 90% (9 out of 10) including HI-neutrophil (HI-N) for 67%, HI-platelet (HI-P) for 75%, and HI-erythroid (HI-E) for 0%. Marrow CR (mCR) and cytogenetic response rates were 67% (6 out of 9) and 83% (5 out of 6), respectively. Notably, 3 out of 4 complex abnormalities were also improved. Moreover, WT1 was decreased from 1200 to 60 copies/µg RNA. Interestingly, all (4 out of 4) of the patients with erythroblast hyperplasia or strong erythroid dysplasia exhibited HI-P, HI-N, blast decrease, and at least one log decrease of WT1 copy number. Owing to high marrow suppression including 100% of grade 3 to 4 neutropenia and all patients but 1 with grade 3 to 4 thrombocytopenia, the time to the second course was longer (43 days) than for single AZA. The times to elevation of neutrophils (>500/µl) and platelets (>50×109/L) were 35 and 37.5 days, respectively. Although 70% (7 out of 10) of patients suffered from infection including febrile neutropenia, no other grade 3 to 4 non-hematological adverse events (AEs) resulting in therapy-related death occurred. At present, four of 10 are continuing AZA without disease progression (PD), 2 patients discontinued it due to impaired quality of life (QOL) and the remaining 4 patients had PD (median 6 months). With a median follow-up time of 24 months from starting AZA alone or in combination, median OS was not reached.Conclusion: Our AZA-based combination therapy induced immediate cytotoxicity of malignant MDS clone with poor cytogenetics and single AZA resistance. SPAC, VP16 and AZA might promote effect of each other. However, prolonged marrow suppression resulted in severe infection, impaired QOL and a delayed treatment schedule. Therefore, response duration appeared to be relatively short, resulting in shorter survival. Appropriate dose setting and treatment duration, effective infection control methods such as G-CSF administration, and prophylactic antibiotics were warranted. However, this method did not require an intravenous drip and had good cost performance. Finally, our cohort is very small, so further experience is needed. DisclosuresNo relevant conflicts of interest to declare.

Safe and Effective Treatment of Patients with Peripheral T-Cell Lymphoma (PTCL) with the Novel HDAC Inhibitor, Belinostat, in Combination with CHOP: Results of the Bel-CHOP Phase 1 Trial

Background: Peripheral T-cell lymphoma (PTCL) is a heterogeneous group of non-Hodgkin lymphomas associated with poor prognosis and repeated recurrence for most subtypes. Currently, anthracycline-based therapies such as cyclophosphamide, doxorubicin, vincristine and prednisone (CHOP) or CHOP-like therapies are recommended as the first-line treatment for PTCL, but the prognosis remains poor with most patients relapsing within 5 years. Thus, improved treatment strategies are still needed.Belinostat is a potent, pan-histone deacetylase inhibitor that was recently approved in the United States for the treatment of patients with relapsed or refractory PTCL (R/R PTCL). Approval was based on results from the pivotal Phase 2 BELIEF study (O'Connor et al, JCO, 2015) of belinostat in R/R PTCL, which demonstrated durable clinical benefit (objective response rate [ORR] 25.8%) and tolerability. Since belinostat (Bel) and each of the components of the CHOP regimen target different aspects of the cell cycle with different mechanisms of action, there is potential for a synergistic effect of a Bel-CHOP combination treatment regimen for patients with PTCL.Methods: Patients with PTCL received CHOP in association with 1000 mg/m2 of belinostat on various schedules, repeated every 21-days for up to 6 cycles. The cohort schema followed a traditional "3+3" dose escalation design. The objective of Part A of the study was to determine the Maximum Tolerated Dose (MTD) of the Bel-CHOP combination. Once the MTD was determined, at least 10 more patients were to be treated in the Expansion Phase (Part B). Belinostat was to be administered as a 1000 mg/m2 IV infusion once daily for up to 5 days, depending on the assigned cohort (Fig 1). The starting cohort was Cohort 3 (CHOP + 1000 mg/m2 of daily belinostat on Days 1-3).Patients received primary prophylaxis with growth factor (G-CSF) support. Dose-limiting toxicities (DLT) were considered during the 1st cycle and included: non-hematological toxicity Grades 3-4, platelet count < 25 X 109/L at any time or ANC < 0.5 X 109/L lasting more than 7 days despite G-CSF administration. The primary endpoint of the study was the determination of the MTD of the Bel-CHOP combination. Secondary endpoints included safety, tolerability and ORR (complete response [CR] + partial response [PR]) and pharmacokinetics.Results: A total of 23 patients were enrolled in the study, 11 of which were treated in Part A. One patient in Part A was deemed inevaluable because the patient died due to disease progression before completing Cycle 1. The MTD was determined to be 1000 mg/m2 on Days 1-5 (Cohort 5); 12 more patients were then treated at this dose level (Part B). The only DLT experienced in the study was in Cohort 3 (Grade 3 Nausea and Vomiting). At the time of this abstract, 18/23 patients (78%) have completed all 6 cycles of Bel-CHOP, with 87% completing at least 4 cycles. Ten patients (43%) had at least one serious adverse event (SAE) and 18 (78%) had at least one Grade 3 or 4 adverse event (AE). The most frequent Grade 3/4 AEs were hematological in nature: neutrophil count decreased (26%), anemia (22%), neutropenia (17%) and white blood cell count decreased (17%). The ORR for the18 patients that have completed an End of Study Visit is 89% (16/18), with the vast majority achieving a CR [72% (n=13)], and 17% (n=3) a PR. Progressive disease was reported in 2 patients.Conclusions: These results demonstrate that the combination of belinostat with CHOP (Bel-CHOP) is well tolerated, with all components of CHOP and belinostat being given at their standard therapeutic doses. The rates of AEs were consistent with those typically reported with CHOP alone, and clinical activity was demonstrated with a response rate of 89% based on 18 evaluable patients. Thus, Bel-CHOP is a promising new regimen in PTCL that will be further tested in a Phase 3 randomized trial. [Display omitted] [Display omitted] DisclosuresBarta:Seattle Genetics: Research Funding. Bhat:Spectrum Pharmaceuticals, Inc: Employment. Song:Spectrum Pharmaceutical, Inc: Employment. Choi:Apectrum Pharmaceuticals, Inc: Employment. Allen:Spectrum Pharmaceuticals, Inc: Employment. Foss:Spectrum Pharmaceuticals; Celgene: Seattle Genetics: Infinity; Millenium: Consultancy, Honoraria, Research Funding, Speakers Bureau.

FCR for Advanced Chronic Lymphocytic Leukemia: Real Life Results from a Series of 50 Patients Treated at a Single Institution

Introduction: The combination of fludarabine, cyclophoshamide and rituximab (FCR) represents the current standard of treatment of fit patients with chronic lymphocytic leukemia (CLL). However, despite the success of the FCR regimen, a number of questions exist on its optimal administration as well as on its short and long term toxicity. As a consequence, in the daily practice (Knauf et al, Hematological Oncology, 2015) less intensive regimens such as bendamustine plus rituximab (BR) are often adopted also in fit patients. Most data on the FCR regimen derive from clinical trials, while data from real life are poor.Methods: Here we report our experience on a series of 50 previously untreated patients with CLL, managed with FCR at a single institution in Italy. Patients were programmed to receive 6 courses plus 2 additional rituximab infusions. The median age of the whole patient population was 60 years (range 41-75), there were 29 males (58%) and 21 females (42%). 44 patients (88 %) received the classical schedule (Fludarabine 25 mg/sqm and Cyclophosphamide 250 mg/sqm, on days 1-3) , while 6 patients (12%), all aged over 70 years, were given FCR-lite (Fludarabine 15 mg/sqm and Cyclophosphamide 200 mg/sqm). In all patients Rituximab was given at 375 mg/sqm on course 1, and 500 mg/sqm infusions 2 to 6. According to Binet staging system, 20 patients were in stage B and 30 in stage C. Patients were carefully selected according to the absence of hepatic, cardiac, renal and respiratory comorbidity and medical judgment (go patients). 40 patients were evaluable for cytogenetic examination (FISH) and none of them had 17 p abnormality.Results: Overall, 46 patients (92%) received the 6 programmed FCR courses, while in four patients therapy was stopped after 1, 2, 4 and 5 courses due to unexplained cytopenia, while CLL cells were undetectable in the bone marrow. Overall response rate was 90 % (74 % CR, 16 % PR) The median duration of response was 30 months, after a median follow up of 38 months. 8 patients have relapsed from CR achievement and 2 of them achieved sustained response with BR, one is on treatment with ibrutinib, while 5 were refractory to salvage therapy. 5 patients died in the setting of refractory disease, 44 are alive and one has been lost to follow-up. Major toxicities (grade 3-4) included: neutropenia registered in 25 patients (50 %), needing G-CSF administration in all cases and producing delay in the administration in 35 % of patients; anemia requiring blood transfusions was registered in 10% of the cases, while no patient required platelet units. Hepatitis B and Herpes Zoster virus reactivation occurred in 1 (2%) and 2 patients (4%), respectively. Overall, 5 patients were hospitalized, all because of febrile neutropenia successfully managed with antibiotic therapy and G-CSF.Conclusions: We conclude that FCR in the "real life" is effective and well tolerated in patients with advanced CLL, provided that a careful evaluation of is made as to eligibility to therapy, mainly in terms of absence of comorbidities.This work was supported by AIL Napoli, "Sezione Bruno Rotoli". DisclosuresNo relevant conflicts of interest to declare.

Potent Stem Cell Mobilization with the Novel CXCR4 Antagonist POL6326 - Results of a Phase IIa Dose Escalation Study in Comparison to G-CSF

Background:Stem cell mobilization (SCM) with G-CSF is efficient but - although overall safe - inconvenient because of the five-day injection regime and certain contraindications. Side effects, sometimes severe, are frequent. These disadvantages fuel the quest for alternative mobilizing agents. Mobilization with the CXCR4-inhibitor plerixafor is rapid, albeit insufficiently efficacious on its own. POL6326, a potent 2nd generation macrocycle CXCR4 antagonist, has demonstrated rapid mobilization kinetics and efficacy in mice. We herein report the results of a Phase IIa dose escalation trial where SCM in response to POL6326 was compared with G-CSF in healthy volunteer stem cell donors.Methods:In this Phase IIa open label trial, healthy volunteer stem cell donors with average mobilization (121±7 CD34+ cells/μL, MW±SEM)after a five-day course of G-CSF, and a wash-out period of at least 6 weeks, received POL6326 at 500-2500 µg/kg as a single 2-hour i.v. infusion. Safety, tolerability, pharmacokinetics (PK) and pharmacodynamics (PD) were assessed in 3-10 subjects/dose group. Subgroups received two doses of POL6326, 1000 and 2500 µg/kg or 1500 and 2500 µg/kg, at least 2 weeks apart (paired intra-individual analysis). For PK and PD blood samples were collected before (0) and at 2, 3, 4, 6, 8 and 24 hrs after infusion start. Complete blood count, CD34+, CFU-C count and PK were assessed at all time points. At 0, 4, 8 and 24 hrs extensive phenotyping of mobilized mature and immature leukocyte subsets was performed. Eight to 14 days after treatment volunteers underwent extensive clinical and laboratory follow-up.Results:POL6326 was very well tolerated. Several volunteers experienced a mild urticarial or itchy macular rash which responded well to H1/H2 blockade. Rating of tolerability/adverse events by volunteers (questionnaire) compared favourably with G-CSF administration. Exposure (Cmax, AUC) was dose-linear. At all doses tested POL6326 mobilized CD34+ progenitor cells and colony-forming cells (CFU-C, Figure 1) exceeding reported peak mobilization with plerixafor in donors at all except the lowest dose levels. In this dataset mobilization after doses of 2000 or 2500 µg/kg did not appear meaningfully stronger than after 1500 µg/kg. The SCM response for CD34+ cells to doses ≥1500 µg/kg was 36.9±2.4/µL (mean±SEM), or 1/3 that of G-CSF (y=0.324x). Good SCM with G-CSF was predictive of good SCM with POL6326 (r=0.63). One/5.7 POL6326-mobilized CD34+ cells was clonogenic (G-CSF: 1 CFU-C/3.4 CD34+ cells) possibly indicating a more immature phenotype of CD34+ cells mobilized by POL6326. POL6326 caused mixed leucocytosis with peak values in the mid-20K/µL. B-lymphocytosis was more and neutrophilia and monocytosis were less pronounced after POL6326 than G-CSF. Compared to G-CSF the subset of plasmocytoid dendritic cell progenitors (pDC) was enriched to a distinct population within the CD34+ cells following SCM with POL6326 as previously described for plerixafor. At the 24 h time point, blood values were well on their way towards normal, and at follow-up all laboratory values had normalized.Summary/Conclusions:The novel CXCR4-antagonist POL6326 is safe, well tolerated, and provides efficient mobilization of HSPCs. Based on the number of mobilized CD34+ cells at higher doses in this study, we conclude that a standard dose of 4x10E6 CD34+ cells/kg can be extracted with a single apheresis for most recipients unless their body weight significantly exceeds the donor weight. However, exploration of alternative dosing regimens may provide even higher mobilization responses. POL6326 can be an effective mobilizing agent for allogeneic donors, including subjects with contra-indications to G-CSF. [Display omitted] [Display omitted] DisclosuresEscot:Polyphor Ltd.: Employment. Douglas:Polyphor Ltd.: Employment. Romagnoli:Polyphor Ltd.: Employment. Chevalier:Polyphor Ltd.: Employment. Dembowsky:Polyphor Ltd.: Consultancy. Hooftman:Polyphor Ltd.: Employment. Bonig:Polyphor Ltd.: Research Funding.

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.

Long-Term HSC Donor Follow-up: Malignancy Incidences after Donation Are Identical Compared to a Matched Control Sample of Registered Donors, but Different to Expected General Population Rates

Introduction:HSC donation is an established procedure with few, manageable short term side effects. As donation is, especially for unrelated volunteer donors, an entirely altruistic act with no direct health benefits, careful and systematic long-term follow-up is crucial. However, interpretation of follow-up data requires a suitable control group. Here, we report first results from a matched pair donor follow-up study. Since 2009, DKMS has extended the routine questionnaire-based donor follow-up to a control group of registered potential stem cell donors who had not actually donated at that time. Donors in the control group (CG) were matched for age, sex and, if possible, HLA to stem cell donors. Eligibility for controls was evaluated using the standard health questionnaire from confirmatory typing stage. After study recruitment, controls were sent identical follow-up questionnaires in the same intervals as real donors. As of July 1st, 2015, our data set for analysis extended to 170,484 questionnaires (58,306 from 21,633 unrelated donors (URD), and 112,178 from 65,543 controls (CG)). Of the URD, 17,472 have donated peripheral blood stem cells (PBSC), 3,735 bone marrow and 145 are donors of both, marrow and PBSC. For statistical reasons, 281 donors requested for donation but deferred at medical pre-examination are included in the donor group.Results:A total of 157 malignancies (excluding ICD-10 C44 neoplasms of skin other than malignant melanoma) have been reported (URD 63, CG 94). In multivariate analysis, significantly more malignancies were observed in females (p <0.001; OR 2.07, 95% CI 1.55 - 2.78), and with increasing age. No difference in the overall malignancy incidence was seen between the URD and control group. In both groups, incidences were significantly lower than the expected rates based on the standard incidences in the German population (standard incidence ratio (SIR) 0.64, 95% CI 0.54 - 0.74). SIR analysis for single ICD malignancy groups revealed significant or close to significant scarcity of reported neoplasms of the lung, oral cavity and colon, all known to correlate with certain health-related behavior (smoking, misuse of alcohol, diet).Due to the administration of GCSF to healthy volunteers for PBSC donation, any hematological malignancies in this group are of particular interest. In the URD group, 1 single case of leukemia (AML) and 2 cases of lymphoma (2 Hodgkin's lymphoma, 1 cutaneous T-cell lymphoma) have been reported from PBSC donors. Because eligibility for the control group was assessed by a confirmatory typing (CT) stage health questionnaire without physical examination, we have included potential donors who where originally cleared at CT stage but deferred during work-up process for medical reasons as part of the URD group for enhanced comparability ('intention to donate'). One case of multiple myeloma was seen in this subgroup (reason for deferral: monoclonal gammopathy). No cases were reported from donors who had donated via bone marrow extraction. In the control group, a total of 8 hematological malignancies were reported (2 Hodgkin's lymphoma, 4 NHL, 1 multiple myeloma, 1 Langerhans cell histiocytosis). No increased incidences of hematological malignancies were observed in either the URD or control group.Discussion:For evaluation of long-term HSC donor follow-up, the applicability of general population data for comparison must be discussed critically. Our data suggest that registered donors as a group are distinct from the general population regarding health-related behavior and attitude or disease susceptibility. Malignancy incidences lower than expected from epidemiological data can most likely be explained by selection effects. People willing to donate stem cells likely represent a healthy sample of the general population. While incomplete reporting cannot be ruled out, it should have affected both URD and CG in the same way. Thus, our approach to include registered potential donors as a control sample is a viable and thorough approach to identify (late) health hazards, further increasing safety and confidence for volunteer stem cell donors. Even more, our study will allow to evaluate donor follow-up data where epidemiological data is insufficient or not applicable, e.g. for autoimmune diseases or general well-being. DisclosuresNo relevant conflicts of interest to declare.

Safety and Functional Studies of Ex Vivo Generated Megakaryocytic Progenitors/Platelets in a Myeloablative Non-Human Primate Model

The ex vivo induction of megakaryocytes/platelets of hematopoietic stem cells represent an effective treatment for thrombocytopenia. We have previously reported a clinically feasible "two-stage culture system" for expanding CD34+ cells and producing megakaryocytes/platelets. With this culture system one cord-blood unit (2 × 106 - 5 × 106 CD34+ cells) yields sufficient megakaryocytes/platelets for treating up to 85 patients (with an average weight of 70 kg and infusion cell number 5.45 × 106 cells/kg). The induced human megakaryocytic cells were capable of producing functional platelets in xenotransplantation mouse model.Here, we further tested the safety and feasibility of ex vivo generated megakaryocytic progenitors/platelets in a myeloablative non-human primate model. Mobilized peripheral blood CD34+ cells of cynomolgus monkeys (Macaca fascicularis) wereisolated after daily administration of G-CSF (100 μg/kg/day) and SCF (50 μg/kg/day) for five days. Expansion and subsequent differentiation of CD34+ cells and megakaryocytic progenitor cells were performed in a modified IMDM basal medium supplemented with various cytokine combinations including stem cell factor, Flt-3 ligand, thrombopoietin, interleukin 3, StemRegenin 1, interleukin 6 for 8 days. The expansion and differentiation processes were closely monitored by flow cytometry for expression of cell surface markers. In vitro morphological identification and CFU assays were carried out for monkey megakaryocytic progenitors. Furthermore, safety and efficacy of induced megakaryocytic progenitor cells were evaluated in vivo by using a thrombocytopenia model of monkeys. Briefly, cynomolgus monkeys were administered with carboplatin at a dose of 8 mg/kg/day on days 1, 2 and 3. On day 7 after the first carboplatin injection, the experimental group monkeys were infused with megakaryocytic progenitor cells (4.6±1.2 × 106/kg ) labeled with anti-monkey IgG-microbead-FITC conjugates. Negative control group was treated with normal saline. Peripheral blood and bone marrow from the tested animals were obtained for analyzing cell differentiation at various times. In vivo bleeding time was recorded to assess the matured platelet function, which was the time length of the bleeding to stop by making a cut in the forearm of the monkeys.After culturing for 8 days, total cells were expanded up to 13.65±3.79-fold. Analysis of the percentage of component cells further showed that CD34+, CD41+, and CD34+/CD41+ megakaryocytic progenitor cells were 46.8%±3.2%, 22.9%±4.6%, and 20.3%±2.8%, respectively. Induced megakaryocytic cells were morphologically distinguishable as they were much larger than CD34+ cells with apparent lobular nuclei. CFU analysis revealed that they increased 12±4.3-fold on day 8 as compared with day 1. In the experimental group, platelet count nadir occurred on day 14 or 15 with 40% of normal value and completely recovered to the normal value on day 26. On the other hand, the nadir of the platelet count in the control group occurred on day 19 or 20 with 20% of normal value and recovered to the normal on day 32. During the nadir phase (from days 14 to 20), the in vivo bleeding time of experimental group was considered normal (6-7.5 minutes) relative to untreated normal monkey values (range 5-7 minutes), whereas it was longer (~8.5 minutes) in control group. Likewise, the nadir of white blood cell (WBC) count (with 45% of the normal value) occurred on day 17, which completely recovered on day 22 for the monkeys infused with induced megakaryocytic progenitors/ platelets. On the other hand, the nadir of WBC count of the control group was 35% of normal value on day 19, which recovered to the normal on day 36. Fifteen days after infusion, flow cytometry and fluorescent microscope analyses showed that about 1% of fluorescent cells remained in bone marrow, indicating successful engraftment of CD34+ stem/progenitor cells in the infused cell preparation. The infused monkeys have survived with no apparent abnormalities for more than one year.Combined, our results strongly suggest that functional human megakaryocytes/platelets can be produced in a large-scale from CD34+ cells for potential clinical application. More importantly, induced non-human primate megakaryocytic progenitors/platelets can be safely administrated to myelosuppressive monkeys, contributing to platelet recovery and early engraftment. DisclosuresQin:Biopharmagen. corp: Employment. Ren:Biopharmagen corp: Employment. Jiang:Biopharmagen.corp: Employment.

Identifying Inherited and Acquired Genetic Factors Involved in Poor Stem Cell Mobilization and Donor-Derived Malignancy

BackgroundMobilized peripheral blood (PB) stem cells (PBSC) are the most frequent source of hematopoietic stem cells (HSC) used for allogeneic HSC transplantation (HSCT). However, genetic factors contributing to donors who mobilize PBSC poorly are not well understood. We previously identified an undetected case of a familial myelodysplastic syndrome/acute leukemia (MDS/AL) predisposition syndrome in a donor with mild pre-donation thrombocytopenia who mobilized very poorly. Familial MDS/AL predisposition syndromes were identified in two other donors found to have hypocellular bone marrows after poor PBSC mobilization. Thus, we hypothesized that we could identify germline MDS/AL predisposition among poor mobilizing donors using a genomic screen of key hematopoietic genes.Patients and MethodsAmong 331 HLA-matched related HSC donors who underwent PBSC mobilization at The University of Chicago from 2001 to 2011, we defined those whose PB CD34+ cells/μl on day 5 post G-CSF administration fell within the lowest quartile as poor mobilizers (PM). Genomic DNA was available for 23 consented PM. This study was approved by The University of Chicago Institutional Review Board. Genomic DNAs isolated from donors' mobilized PBSC product were screened utilizing MarrowSeq, an next generation sequencing (NGS) assay targeting 142 genes involved in inherited and acquired bone marrow failure syndromes and hematologic malignancies. Potentially damaging variants were confirmed by Sanger sequencing. Clinical NGS panel testing was used to identify additional acquired mutations in one donor/recipient pair.ResultsAmong the 23 PM sequenced using MarrowSeq, deleterious mutations were identified in 2 (9%). The first, a 63 year old male donor with mild thrombocytopenia (platelet count 136 K/μL) and macrocytosis (MCV 107.8 fL) was previously reported by our group to have a deleterious germline TERT mutation (c.2908A>G; p.M970V; 48% allelic ratio), which was also identified in the recipient's leukemia. This donor required bone marrow harvest after two failed PBSC mobilization attempts and the recipient died 5 months post-HSCT due to complications of poor engraftment. The second was a 67 year old mildly thrombocytopenic male donor (platelet count 139 K/μl) with a deleterious TET2 mutation (c.3765C>G; p.Y1255*; 48% allelic ratio), which was not present in the recipient's skin fibroblasts, most consistent with the presence of clonal hematopoiesis of indeterminate potential (CHIP) in the donor. Follow-up of this donor six years after collection of these TET2-mutated PBSCs demonstrated persistent mild thrombocytopenia (platelet count 121 K/μl), but otherwise normal blood counts. The recipient achieved remission after reinduction chemotherapy for relapsed AML followed by matched-related HSCT. Post-HSCT, the recipient demonstrated persistent mild thrombocytopenia. Six years later, she developed transfusion-dependent anemia and refractory anemia with excess blasts-2. Karyotype and microsatellite marker analysis of the bone marrow were consistent with an MDS of male donor origin. NGS sequencing from the bone marrow demonstrated the same TET2 mutation found in the mobilized PBSC product used in the transplant and an additional acquired RUNX1 mutation (c.585del; p.T196Qfs*15). Additional data from NGS sequencing of donor and recipient samples over time will be presented.ConclusionsWe demonstrate that both inherited and acquired genetic factors in PBSC from apparently healthy donors can contribute to poor mobilization and donor-derived malignancy post-HSCT. To the best of our knowledge, this is the first example of clonal evolution of TET2-mutated CHIP within a donor resulting in a donor-derived leukemia within the HSCT recipient. Replicative stress to reconstitute hematopoiesis in the recipient and/or the recipient's damaged bone marrow microenvironment may be contributing factors to the development of this donor-derived leukemia while the donor remains well. This study suggests that PM with thrombocytopenia may carry inherited or acquired mutations in hematopoietic genes, and alternative donor options should be considered. Our study also raises concerns about whether we need to screen for CHIP in healthy donors with unexplained cytopenias, especially as donors of increasing age are utilized.This research was supported by the American Society of Hematology through the ASH HONORS Award. Disclosuresvan Besien:Miltenyi Biotec: Research Funding. Godley:Onconova: Research Funding.

P3-5-60 - Examination about febrile neutropenia in patient performed FEC with breast cancer

Background: Chemotherapy takes an important role in mammary cancer treatment, and to obtain therapy effect surely, RDI (relative dose intensity) is important.To preserve RDI, management of febrile neutropenia (FN) is demanded in a side effect. In this country, prolonged model G-CSF is approved, and primary prevention administration was enabled, but examination to relate to choice of the administration patient is not enough. We investigated a risk of FN in patient performed FEC and we examined whether exhaustion of the case that should have considered prolonged model G-CSF administration was possible.Methods: We intended for patient with breast cancer of 186 examples who took FEC medical treatment during from January, 2009 to December, 2014 in our hospital.We analyzed symptom example number that caused FN, patient background, and RDI. We evaluated the adverse event using CTCAE v.4.0.Results: In 186 examples, 10 examples (5.3%) had FN. Preoperative adjuvant therapy 2 of 88 examples (2.3%), postoperative adjuvant therapy 5 of 76 examples (6.5%), recurrence treatment 1 of 9 examples (11.1%), stage4 treatment 2 of 13 examples (15.4%) had FN (P = 0.1). The case that inflammation of mouth was present before FN showed 4 examples. One example had the post-operative wound infection. After FN, 2 examples canceled FEC, and one example made RDI decrease by 80%, and 7 examples maintained RDI 100% (one example uses prolonged model G-SCF together).Conclusion: FN was seen a lot in metastasis recurrence example. It was thought that there was need of mouth care before the FN onset, but we might prevent the onset of FN by prolonged model G-CSF use when it had difficulty to contorol infection. Background: Chemotherapy takes an important role in mammary cancer treatment, and to obtain therapy effect surely, RDI (relative dose intensity) is important. To preserve RDI, management of febrile neutropenia (FN) is demanded in a side effect. In this country, prolonged model G-CSF is approved, and primary prevention administration was enabled, but examination to relate to choice of the administration patient is not enough. We investigated a risk of FN in patient performed FEC and we examined whether exhaustion of the case that should have considered prolonged model G-CSF administration was possible. Methods: We intended for patient with breast cancer of 186 examples who took FEC medical treatment during from January, 2009 to December, 2014 in our hospital.We analyzed symptom example number that caused FN, patient background, and RDI. We evaluated the adverse event using CTCAE v.4.0. Results: In 186 examples, 10 examples (5.3%) had FN. Preoperative adjuvant therapy 2 of 88 examples (2.3%), postoperative adjuvant therapy 5 of 76 examples (6.5%), recurrence treatment 1 of 9 examples (11.1%), stage4 treatment 2 of 13 examples (15.4%) had FN (P = 0.1). The case that inflammation of mouth was present before FN showed 4 examples. One example had the post-operative wound infection. After FN, 2 examples canceled FEC, and one example made RDI decrease by 80%, and 7 examples maintained RDI 100% (one example uses prolonged model G-SCF together). Conclusion: FN was seen a lot in metastasis recurrence example. It was thought that there was need of mouth care before the FN onset, but we might prevent the onset of FN by prolonged model G-CSF use when it had difficulty to contorol infection.

Spontaneous splenic rupture following stem cell mobilization with G-CSF and plerixafor in AL amyloidosis

BackgroundAL amyloidosis is a rare plasma cell dyscrasia with multiorgan involvement. Good risk patients are candidate to high dose chemotherapy and autologous stem cell transplantation. However both transplantation and stem cell collection entail significant risk in such patients. Plerixafor is a novel mobilizing agent approved for use in “poor mobilizer” patients with lymphoma and multiple myeloma; experience in systemic amyloidosis patients is limited. Case ReportWe describe a case of spontaneous splenic rupture following administration of G-CSF and plerixafor in a patient with AL amyloidosis who previously underwent heart transplantation due to amyloid heart involvement. Results and ConclusionThis is the first report of spontaneous splenic rupture following stem cell mobilization with G-CSF and plerixafor in AL amyloidosis. The role of plerixafor has to be established. AL amyloidosis patients undergoing stem cell mobilization need careful monitoring of signs and symptoms of spontaneous splenic rupture.

Donor CD4 T Cell Diversity Determines Virus Reactivation in Patients After HLA-Matched Allogeneic Stem Cell Transplantation

Delayed reconstitution of the T cell compartment in recipients of allogeneic stem cell grafts is associated with an increase of reactivation of latent viruses. Thereby, the transplanted T cell repertoire appears to be one of the factors that affect T cell reconstitution. Therefore, we studied the T cell receptor beta (TCRβ) gene rearrangements of flow cytometry–sorted CD4+ and CD8+ T cells from the peripheral blood of 23 allogeneic donors before G-CSF administration and on the day of apheresis. For this purpose, TCRβ rearrangements were amplified by multiplex PCR followed by high-throughput amplicon sequencing. Overall, CD4+ T cells displayed a significantly higher TCRβ diversity compared to CD8+ T cells irrespective of G-CSF administration. In line, no significant impact of G-CSF treatment on the TCR Vβ repertoire usage was found. However, correlation of the donor T cell repertoire with clinical outcomes of the recipient revealed that a higher CD4+ TCRβ diversity after G-CSF treatment is associated with lower reactivation of cytomegalovirus and Epstein–Barr virus. By contrast, no protecting correlation was observed for CD8+ T cells. In essence, our deep TCRβ analysis identifies the importance of the CD4+ T cell compartment for the control of latent viruses after allogeneic stem cell transplantation.

Clinical predictors of failure of granulocyte colony stimulating factor (G-CSF) prophylaxis in patients with breast cancer treated with dose dense epirubicin (E), cyclophosphomide (C) + paclitaxel (T) Adjuvant chemotherapy: Subgroup analysis of the NCIC CTG MA.21 study (NCT00014222).

1033 Background: Administration of prophylactic GCSF is recommended for curative chemotherapy regimens with an estimated incidence of febrile neutropenia of ≥ 20% (Smith et al.,JCO 24:3187-3205, 20...

The use of hematocrit level for predicting the efficiency of peripheral blood CD34(+) cell collection after G-CSF Mobilization in Healthy Donors.

Recently, peripheral blood stem cell (PBSC) has been widely used and replaced bone marrow (BM) as the stem cell source in allogeneic hematopoietic stem cell transplantation (HSCT) because of a more rapid engraftment, easier accessibility, and lower risk of donor complications. We, therefore, report the predicting factors for the high PBSC harvest yields in 50 healthy donors. Among the 50 donors, median collected CD34(+) cell number was 4.6 × 10(6/) kg (1.5-16.3 × 10(6) /kg). Number of circulating CD34+ cells and hematocrit (HCT) level increased parallelly whereas peripheral CD34+ cell numbers were decreased with increasing donor age. In univariate analysis, HCT level≥ 35.5% at the time of PBSC collection was significantly associated with high PBSC number (≥ 5.0 × 10(6) cells/kg) and donor aged <30 years was significantly associated with collected CD34+ cells ≥ 6.0 × 10(6) /kg, P = 0.03. HCT level ≥35.5% was an independent parameter for high WBC count (≥50 × 10(9) /L), P < 0.05. None of donor who had both HCT < 35.5% and WBC < 50 × 10(9) /L had circulating CD34+ cells ≥ 5.0 × 10(6) /kg. Platelet count ≥ 200 × 10(9) /L was found significantly in donors with WBC ≥ 40 × 10(9) /L (P = 0.03) and HCT ≥ 35.5%, P < 0.05. Collected PBSC number tended to be higher in our donors with high levels of HCT, WBC, and platelet. We also found that HCT and platelet levels in our donors decreased after receiving G-CSF administration compared with the initial complete blood counts (CBC) results. We, therefore, concluded that HCT level at the time of initiation leukapheresis was an important predictor for PBSC collection yields.