ALBERT

Beschreibung: Introduction: The collection of hematopoietic stem cells from the peripheral blood in healthy donors has been established as a highly efficient method in clinical practice. Nevertheless, there are some donors who mobilize poorly despite adequate mobilization regimes. Several factors influencing the process of stem cell mobilization have previously been discussed in the literature. Methods: In total, the data of 7.216 unrelated donors who underwent GCSF-induced stem cell mobilization and collection in two German apheresis centers between July 1997 and August 2012 were retrospectively reviewed. We systematically analyzed more than 30 factors with potential influence on the mobilization process and established a statistically stable model in order to predict the mobilization efficacy and the harvest success in unrelated stem cell donors. Based on empirical data and standard values, we created three model donors of each gender with different donor profiles (favorable/average/unfavorable). In those model donors we calculated the corresponding likelihood of a successful stem cell harvest dependent on the recipient’s weight. Results: Overall, ten variables with high statistical significance were included in the prediction model. Body mass index, platelet count, absolute lymphocyte count and the relative monocyte count correlated positively with the CD34+ count in the peripheral blood after five days of GCSF use (p 〈 0.0001 in a multivariate analysis). In contrast, female sex, age, smoking, lactate dehydrogenase (ldh), relative monocyte count, and the relative large unstained cell count were associated negatively with the CD34+ count on day five (p 〈 0.0001 in a multivariate analysis). In order to predict the harvest success (collection of 〉 2 x 10^6 CD34+ cells/kg recipient weight after first apheresis) three different models were compared in a ROC-analysis. The first model was purely based on female sex and recipient weight. The second model additionally contained the predicted CD34+ counts. Finally, in the third model the actual observed CD34+ counts were included. By adding the predicted CD34+ counts to the simple model, a significant improvement of the predictability of a harvest success could be viewed although a considerable difference comparing these results to the model with the observed CD34+ counts remained (figure 1). Furthermore, the prediction of harvest success in model donors (figure 2) revealed that donors with a favorable donor profile (covariates set to the most favorable values within the normal range) showed a particular high likelihood for a successful harvest (100 % likelihood, irrespective to donor’s gender or recipient’s weight). The likelihood for a successful harvest in male donors with an average distribution of covariates (covariates set to the empirical mean value) was nearly 100% as well even in donations for heavy recipients. Contrary, the likelihood for an average female donor was high in normal weight recipients (97 %; 60 kg recipient) but decreased with rising recipient weight (78 %; 140 kg recipient). In donors with an unfavorable profile (covariates set to the worst values within the normal range), especially in females, the chance for a successful stem cell collection was poor even when donating for light recipients (54 % in males, 10 % in females; 60 kg recipient). Conclusions: In conclusion, multiple factors with influence on the CD34+ count after GCSF mobilization in healthy donors have been identified. With the prediction model a significant gain in the predictability of a harvest success could be achieved though a certain amount of unexplained variance still remains. Model donors with a favorable or average donor profile had a high likelihood for a successful stem cell collection. In donors with an unfavorable profile, especially in females, the chance for a harvest success was very low. Figure 1: Prediction of harvest success (〉 2 x 10^6 CD34+ cells/kg recipient weight after 1st apheresis) Figure 1:. Prediction of harvest success (〉 2 x 10^6 CD34+ cells/kg recipient weight after 1st apheresis) Abb.: AUC – area under the curve Figure 2: Probability of harvest success in model donors (〉 2 x 10^6 CD34+ cells/kg recipient weight after 1st apheresis) Figure 2:. Probability of harvest success in model donors (〉 2 x 10^6 CD34+ cells/kg recipient weight after 1st apheresis) Disclosures Schmidt: Cellex GmbH: Employment. Ehninger:Cellex GmbH: Equity Ownership. Off Label Use: G-CSF.

Beschreibung: Key Points CD107a protects cytotoxic lymphocytes from damage during degranulation. Interference with CD107a expression can cause the death of cytotoxic lymphocytes during degranulation.

Beschreibung: In patients with multiple myeloma (MM), risk stratification by chromosomal abnormalities may enable a more rational selection of therapeutic approaches. In the present study, we analyzed the prognostic value of 12 chromosomal abnormalities in a series of 354 MM patients treated within the HOVON-65/GMMG-HD4 trial. Because of the 2-arm design of the study, we were able to analyze the effect of a bortezomib-based treatment before and after autologous stem cell transplantation (arm B) compared with standard treatment without bortezomib (arm A). For allanalyzed chromosomal aberrations, progression-free survival (PFS) and overall survival (OS) were at least equal or superior in the bortezomib arm compared with the standard arm. Strikingly, patients with del(17p13) benefited the most from the bortezomib-containing treatment: the median PFS in arm A was 12.0 months and in arm B it was 26.2 months (P = .024); the 3 year-OS for arm A was 17% and for arm B it was 69% (P = .028). After multivariate analysis, del(17p13) was an independent predictor for PFS (P 〈 .0001) and OS (P 〈 .0001) in arm A, whereas no statistically significant effect on PFS (P = .28) or OS (P = .12) was seen in arm B. In conclusion, the adverse impact of del(17p13) on PFS and OS could be significantly reduced by bortezomib-based treatment, suggesting that long-term administration of bortezomib should be recommended for patients carrying del(17p13). This trial is registered at the International Standard Randomised Controlled Trial Number Register as ISRCTN64455289.

Beschreibung: Key Points EGFL7 promotes angiogenesis via its interaction with integrin αvβ3. EGFL7 is involved in physiological and pathological angiogenesis.

Beschreibung: Abstract 3986 Background: Multiple Myeloma (MM) is an incurable plasma cell neoplasm. Although current lenalidomide (R) and bortezomib containing up-front regimens can now achieve overall response rates approaching 100%, patients eventually relapse with progressively refractory disease. Histone deacetylase inhibitors (HDACi), in Phase I clinical trials in patients with multiple myeloma, have shown promising activity when combined with other agents such as bortezomib. Vorinostat, (suberoylanilide hydroxamic acid; SAHA) is an oral HDACi, currently FDA approved in the United States for the treatment of cutaneous T-cell lymphoma. Here we report the findings of the combination of vorinostat (Zolinza®), lenalidomide and dexamethasone (ZRD) in multiple myeloma patients who were refractory to RD. Methods: Patients received oral vorinostat 300 mg or 400 mg once daily (days 1–7 and days 15–21), lenalidomide 10–25 mg (days 1–21) and dexamethasone 20–40 mg weekly (days 1, 8, 15, 22) in a 28-day cycle Subjects: Twenty-nine patients were treated and all were refractory to RD; 76% were refractory to at least one bortezomib containing regimen and 48% were refractory to the combination of VRD. Twenty-six patients (90%) had undergone prior high dose therapy with autologous stem cell transplant. The median number of prior therapies was 4 (range 2–13). Results: The overall response rate (ORR) was 24 % with 1 VGPR and 6 PR. The clinical benefit rate (ORR + MR) was 51% including 8 MR. Nine patients (31%) had stable disease. The median duration of response (DOR) was 4 months (range, 0–36). The median overall survival (OS) was 11 months (range, 4–36). Common toxicities including diarrhea and fatigue (all grades) were 41% and 34% respectively. The incidence of grade 3/4 neutropenia was 45 % and grade 3/4 thrombocytopenia was 34%. Conclusion: The combination of ZRD showed significant activity in patients with RD relapsed/refractory multiple myeloma. ZRD was well tolerated and is a viable option for patients who do not respond to lenalidomide-based therapy. Further, since all 3 agents are available in oral formulations, ZRD provides an additional option for those patients wishing to avoid intravenous therapy. Formal phase II studies of this combination are in preparation. Disclosures: Off Label Use: Vorinostat is an oral HDAC inhibitor and is being evaluated in the treatment of Multiple Myeloma. Bilotti:Celgene: Consultancy, Speakers Bureau; Millenium: Consultancy, Speakers Bureau. McNeill:Celgene: Consultancy, Speakers Bureau; Millenium: Consultancy, Speakers Bureau. Graef:Merck: Employment. Vesole:Celgene: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Millennium: Speakers Bureau. Siegel:Celgene: Consultancy, Honoraria, Research Funding, Speakers Bureau; Millenium: Consultancy, Honoraria, Research Funding, Speakers Bureau; Merck: Consultancy.

Beschreibung: BACKGROUND: Suicide gene therapy applied to haploidentical hematopoietic stem cell transplantation (haplo-HSCT) is one of the widest clinical applications of gene therapy. By the infusion of donor lymphocytes transduced to express the Herpes Simplex Virus Thymidine Kinase (TK) suicide gene, patients achieve a rapid immune reconstitution and substantial protection against tumor recurrence. TK-cells are promptly eliminated in case of graft versus host disease (GvHD), with complete resolution of the adverse reaction. In previous studies, we showed that TK-cell infusions are necessary and sufficient to promote the generation of a fast, polyclonal and full competent T cell repertoire. In the present work we characterize the immunological profile of a cohort of long-term survivors after suicide gene therapy and we studied the long-term fate of TK-cells to shed light on memory T cell dynamics after transplantation. RESULTS: We studied 9 adult patients who underwent haplo-HSCT and infusion of purified suicide-gene modified donor T cells (median dose: 1.9x107 cells/kg, range:0.9x106-39.5x106) for high-risk hematologic malignancies between 1995 and 2010 (TK patients). At a median follow-up of 7,4 years (range 3.2-12.3), all patients are in complete remission. Two out of 9 patients (22%) experienced GvHD in the early phase post immune reconstitution; in all cases, ganciclovir (GCV) administration proved effective in abrogating the adverse reaction. No symptoms or complications related to GvHD were observed during the long-term follow up, and none of the patient is receiving immunosuppressive drugs. A complete recovery of NK cells, B lymphocytes and αβ or γδ T cells was observed. The CD8+ and CD4+ T cell compartment of TK patients were characterized by level of naïve and memory cell comparable to age and sex matched healthy controls. The quantification of CD4+ CD31+ CD62L+ CD45RA+ CD95- recent thymic emigrants and measure of single joint T-cell receptor excision circles demonstrated that the normalization of the T cell compartment was supported by a completely recovered thymic output. TK-cells were detected in all patients (100%), at low levels (median=4cells/uL). Ex vivo selection of pure TK-cells after polyclonal stimulation and LNGFR-purification confirmed the presence of functional transduced cells, thus directly demonstrating the ability of memory T cells to persist for years. Of notice TK-cells could be retrieved also in patients successfully treated with GCV for GvHD, thus confirming the selective action of GCV only on proliferating TK-cells. Accordingly, GCV sensitivity was preserved in long-term persisting TK-cells, independently from their differentiation phenotype. TK-cells circulating in patients displayed a memory phenotype comprising effector memory (TEM), central memory (TCM) and stem memory (TSCM) T cells and exhibited a low level of Ki-67 positivity, thus suggesting the maintenance of a pool of gene modified memory cells through homeostatic proliferation. The number of TK-cells circulating at the longest follow-up did not correlate with the number of infused cells, nor patients or donors’ age, but instead with the peak of TK-cells observed within the first months after infusion, suggesting that antigen recognition is dominant in driving in vivo expansion and persistence of memory T cells. We evaluated whether the phenotype of infused TK-cells was able to affect the long-term fate of gene-modified memory T cells. We observed that the number of infused TSCM cells positively correlated with early TK-cell expansion and with their long-term persistence, suggesting that TSCMmight play a privileged role in the generation of a long-lasting immunological memory. CONCLUSION: These data show that a complete and physiological donor-derived immune system is restored in adult surviving long-term after suicide gene therapy. After infusion, gene modified cells persist for up to 12 years in treated patients. This setting can be exploited to investigate the requirements at the basis of the generation of a long-lasting immunological memory in vivo. Further studies on TK-cell TCR repertoire and vector integrations are currently being performed to elucidate the in vivo dynamics of infused memory T cells. Disclosures Lambiase: MolMed S.p.A: Employment. Traversari:MolMed S.p.A: Employment. Bordignon:MolMed S.p.A: Chairman and CEO Other. Bonini:MolMed S.p.A: Consultancy.

Beschreibung: Introduction The JAK2 V617F mutation is present in almost all Polycythemia Vera (PV) and in 60-70% of Essential Thrombocythemia (ET) cases. There are experimental evidences that this acquired clonal driving mutation is involved in disease phenotype in murine models and in humans. However, it is still unclear if the %V617F changes over time and if it is necessary to systematically quantify %V617F at diagnosis and during follow-up. Patients and Methods We are conducted a prospective multicentre (n=5) study cohort in newly diagnosed PV and ET patients, harvested before any cytotoxic therapy and at 3 years, with the aim to correlate the %V617F at diagnosis, the %V617F 3 years later, and the variation of the %V617F between y0 and y3 with clinical evolution. Primary or secondary myelofibrosis, and secondary leukemia were not included. The primary objective is to determine whether an increase of %V617F between diagnosis and after 3 years could be related to a poor evolution of the disease. A secondary objective is to determine whether the %V617F at diagnosis is associated with the evolution at 3y. The primary endpoint is the worsening (Y / N) at y3 defined by the presence of at least one of the following criteria: leukocytosis 〉12G / L or immature granulocytes 〉2% or erythroblasts 〉1%; anemia or thrombocytopenia not related to treatment toxicity; development or progressive splenomegaly; thrombocytosis on cytoreductive therapy; patients inadequately controlled with treatment (defined by at least one treatment change for reason other than an adverse event). Results To date, 201 patients have been included. Clinical datas are currently available for the 176 first included (91 PV and 85 ET; 90 females and 86 males) and %V617F measurement for 167 cases. At diagnosis, overall median age was 65, identical for PV and ET patients, but curiously women were a little older than men (72 vs 65, p

Beschreibung: Abstract 165FN2 CB and KB contributed equally and should be considered aequo loco. Wiskott-Aldrich-Syndrome (WAS) is a rare and life-threatening immune-disorder characterized by autoimmunity, microthrombocytopenia, immunodeficiency and susceptibility to lymphoma. WAS is caused by mutations in the WAS gene which encodes WASP, a key regulator of actin polymerization exclusively expressed in hematopoietic cells. WASP deficiency causes defects in lymphocytes, myeloid cells, and platelets. We here report a comprehensive analysis of ten patients treated by hematopoietic stem cell gene therapy between 2006 and 2009 (median follow up time 29.6 months, range 15 to 58 months). Patients were mobilized with G-CSF alone (3/10) or G-CSF combined with anti-CXCR4/AMD3100 (7/10), conditioned with busulfan (8mg/kg body weight) and received between 2.8×106 and 24.9×106 cells/kg bw, with a median transduction efficacy of 52%. Upon transplantation of retrovirus-transduced WASP-expressing progenitor cells, the proportion of corrected platelets and lymphocytes increased steadily over time reaching 85–90% and 80–85%, respectively. Interestingly, also myeloid cells showed a continuously increasing percentage of WASP-expressing fractions (20 to 70%). Due to transplantation of insufficient numbers of WASP-transduced HSC, one patient failed to engraft. He had no evidence of corrected myeloid/hematopoietic progenitor cells and continued to suffer from life-threatening infections and autoimmunity. He was successfully treated by haploidentical HSCT. All other patients had marked improvement of their clinical status. Bleeding diatheses, susceptibility to infections, and autoimmunity resolved. Patients had evidence of significant and sustained increase in platelet counts (p=0.01) together with a reconstituted WASP expression and normalization of thrombocyte size (p

Beschreibung: Key Points Tmprss6 siRNA induces hepcidin and diminishes iron in hemochromatosis or thalassemia mice, improving the anemia seen in the latter model. Manipulation of TMPRSS6 with RNAi therapeutics may be an approach to treating iron overload diseases associated with low hepcidin levels.

Beschreibung: Background Arsenic trioxide (ATO) is regarded as the treatment of choice for relapsed PML-RARA+ acute promyelocytic leukemia (rAPL). In 2008, a European online registry of rAPL based on uniform CRFs was established under the auspices of the European LeukemiaNet (ELN) to gain insights in the clinical and biological characteristics of rAPL treated with ATO and to allow an assessment of the different options for postconsolidation therapy. Methods Eligibility criteria for prospective or retrospective registration were PML-RARA+ 1st or successive molecular or clinical relapse of APL from the year 2003 onwards. The treatment should be based on the European recommendations for treatment of rAPL (www.leukemia-net.org/content/), which offer induction and consolidation therapy with ATO and several options for post-consolidation therapy including autologous or allogeneic stem cell transplantation or chemotherapy consolidation followed by various modifications of maintenance therapy ± ATO - to be selected depending on several variables including patient age, performance status, PCR status after consolidation, type of frontline therapy, first CR duration and donor availability. Results By 30 June 2013, of 220 registered cases, 198 were evaluable (172 in 1st, 26 in ≥2nd relapse). Of these, 149 patients (pts) in 1st relapse received ATO-based salvage therapy after standard frontline therapy based on all-trans retinoic acid (ATRA) and anthracyclines (98 hematological (hematol) relapses of bone marrow combined with CNS relapse in 5 pts, 40 molecular (mol), 11 isolated extramedullary, mainly CNS). Clinical characteristics: median age at relapse 44 years (y) (4 to 81), 67% males, Sanz Risk Score at 1st diagnosis: low 23%, intermediate 48% and high 29% of pts. Median duration of first remission was 565 d (105 d to 7.0 y). The median treatment duration of ATO (0,15 mg/kg/day) plus ATRA (44% of pts) for remission induction (ind) was 30 days (d) and for consolidation (cons) 25 d. CNS relapses received ATO and additional intrathecal chemotherapy ± irradiation. WBC white blood cell count; Leuko.: leukocytosis requiring hydroxyurea; ADS: APL differentiation syndrome; 1 median; 2 hematological; 3 RT-PCR of PML-RARA negative. In non-hematological relapses, no early deaths occurred and no major side effects of ATO were seen. Median follow up of the 149 pts was 2.8 y (6 d to 10 y). Three-year overall survival (OS) was 70%, 95%CI [61;79] and 6-year OS 56% [42;70]. Three-year OS of hematological, molecular and extramedullary relapse was 69% [58;80], 66% [48;84] and 90% [81;99], respectively (p=0.2). Concerning outcome after transplantation, 3-year OS after autologous was 82% [70;94] (n=55), after allogeneic 75% [58;92] (n=32) and without transplantation 66% [48;84] (n=55) (p=0.4). Conclusions With ATO-based salvage therapy over 50% of patients in 1st relapse of APL can probably be cured. Pts in molecular relapse have a lower rate of early complications and death. Long-term survival is possible with transplantation-free approaches, but transplantation seems to improve the outcome. Disclosures: Lengfelder: TEVA/ Cephalon: Research Funding. Lo-Coco:TEVA: Speakers Bureau.