ALBERT

Description: Abstract The accumulation of soil carbon (C) is regulated by a complex interplay between abiotic and biotic factors. Our study aimed to identify the main drivers of soil C accumulation in the boreal forest of eastern North America. Ecosystem C pools were measured in 72 sites of fire origin that burned 2–314 years ago over a vast region with a range of ∆ mean annual temperature of 3°C and one of ∆ 500 mm total precipitation. We used a set of multivariate a priori causal hypotheses to test the influence of time since fire (TSF), climate, soil physico‐chemistry and bryophyte dominance on forest soil organic C accumulation. Integrating the direct and indirect effects among abiotic and biotic variables explained as much as 50% of the full model variability. The main direct drivers of soil C stocks were: TSF 〉bryophyte dominance of the FH layer and metal oxide content 〉pH of the mineral soil. Only climate parameters related to water availability contributed significantly to explaining soil C stock variation. Importantly, climate was found to affect FH layer and mineral soil C stocks indirectly through its effects on bryophyte dominance and organo‐metal complexation, respectively. Soil texture had no influence on soil C stocks. Soil C stocks increased both in the FH layer and mineral soil with TSF and this effect was linked to a decrease in pH with TSF in mineral soil. TSF thus appears to be an important factor of soil development and of C sequestration in mineral soil through its influence on soil chemistry. Overall, this work highlights that integrating the complex interplay between the main drivers of soil C stocks into mechanistic models of C dynamics could improve our ability to assess C stocks and better anticipate the response of the boreal forest to global change.

Description: Introduction Myeloproliferative Neoplasm (MPN) unclassifiable (MPN-U), is a heterogeneous disease that presents with an MPN-type clinical/ histological phenotype yet fails to meet diagnostic criteria for other MPN entities. Incidence is

Description: Ex vivo expansion of hematopoietic stem/progenitor cells may result in defective engraftment. Human cord blood CD34+ progenitor cells were synchronized and assayed for adhesion and migration onto fibronectin (Fn) and vascular cell adhesion molecule-1 (VCAM-1) at different stages of a first cell cycle executed ex vivo. During S phase transit, adhesion to Fn was transiently increased while binding to VCAM-1 was reversibly decreased, after which adhesion to both ligands returned to baseline levels with cell cycle completion. Transmigration across Fn and VCAM-1 decreased irreversibly during S phase progression. The function of α4 and α5 integrins was assessed with specific neutralizing antibodies. In uncultured CD34+ cells and long-term culture-initiating cells (LTC-ICs), both adhesion and migration on Fn were inhibited by anti-α4 but not by anti-α5 antibodies. In mitotically activated CD34+ cells and LTC-ICs, adhesion and migration on Fn were mainly dependent on α5 integrin and to a lesser extent on α4 integrin. Changes in integrin function were not dependent on parallel modulation of integrin expression. In conclusion, Fn and VCAM-1 binding of progenitor cells fluctuates reversibly during cell cycle transit ex vivo. In addition, our data show that mitogenic activation induces a shift from a dominant α4 to a preferential α5 integrin–dependent interaction with Fn.

Description: Poor graft function (PGF) is a rare but serious complication of hematopoietic cell transplantation (HCT). Boost of CD34+stem cells could be a therapeutic option but is not always feasible nor successfull. Due to their hematopoietic support properties and immune regulatory effects, multipotent mesenchymal stromal cells (MSC) could be considered a good candidate to help restore bone marrow (BM) niches homeostasis and facilitate hematopoiesis after HCT. Here, we conducted a multicenter prospective study to evaluate the efficacy of MSC perfusion to improve PGF after HCT. Thirty patients (median age 51y, range 11 to 70y) received a single IV administration of third-party donor BM-derived MSC (1-2 x 10 exp6/kg body weight) for PGF after allogeneic HCT (median 151 days after HCT, range 62- 430). PGF was defined as (1) at least one cytopenia (Hb 〈 80 g/L, absolute neutrophil count [ANC] 0.5 x 10exp9/L and transfusion independence). Corresponding response rates increased at day 180, with 69.2% (95% CI, 51.5 - 86.9%) OR and 61.5% (95% CI, 42.8 - 80.2%) CR, respectively. Overall survival at 1 year post-MSC was 70% (95% CI, 53.6 - 86.3%), with all but one of the d90-responders being alive (1 death to relapse of hematological malignancy). No severe adverse event related to MSC administration was reported during the 1-year follow-up. In conclusion , perfusion of BM-derived MSC from a third party donor appeared to be safe and to improve PGF after allogeneic HCT, although spontaneous amelioration cannot be excluded. Further (ideally comparative) studies are warranted to confirme our results. Disclosures Selleslag: Celyad: Other: Clinical trial research (no honoraria recieved); Novartis: Consultancy, Honoraria, Speakers Bureau.

Description: Serum erythropoietin (sEpo) concentration is primarily related to the rate of renal production and, under the stimulus of hypoxia, increases exponentially as hemoglobin (Hb) decreases. Additional factors, however, appear to influence sEpo, and in this work, we performed studies to evaluate the role of the red blood cell precursor mass. We first compared the relationship of sEpo with Hb in patients with low versus high erythroid activity. The first group included 27 patients with erythroid aplasia or hypoplasia having serum transferrin receptor (sTfR) levels 〈 3 mg/L (erythroid activity 〈 0.6 times normal), while the second one included 28 patients with β-thalassemia intermedia having sTfR levels 〉 10 mg/L (erythroid activity 〉 2 times normal). There was no difference between the two groups with respect to Hb (8.3 ± 1.6 v 8.0 ± 1.3 g/dL, P 〉 .05), but sEpo levels were notably higher in patients with low erythroid activity (1,601 ± 1,542 v 235 ± 143 mU/mL,P 〈 .001). In fact, multivariate analysis of variance (ANOVA) showed that, at any given Hb level, sEpo was higher in patients with low erythroid activity (P 〈 .0001). Twenty patients undergoing allogeneic or autologous bone marrow transplantation (BMT) were then investigated. A marked increase in sEpo was seen in all cases at the time of marrow aplasia, disproportionately high when compared with the small decrease in Hb level. Sequential studies were also performed in five patients with iron deficiency anemia undergoing intravenous (IV) iron therapy. Within 24 to 72 hours after starting iron treatment, marked decreases in sEpo (up to one log magnitude) were found before any change in Hb level. Similar observations were made in patients with megaloblastic anemia and in a case of pure red blood cell aplasia. These findings point to an inverse relationship between red blood cell precursor mass and sEpo: at any given Hb level, the higher the number of red blood cell precursors, the lower the sEpo concentration. The most likely explanation for this is that sEpo levels are regulated not only by the rate of renal production, but also by the rate of utilization by erythroid cells.

Description: Relapse remains the major cause of failure after high-dose chemotherapy and autologous peripheral blood stem cell transplantation (PBSCT). As tumor contamination of the graft may contribute to this, CD34 selection has been used as an attempt to decrease the relapse rate. However, immune reconstitution may be delayed with CD34 selection due to the removal of T cells, natural killer (NK) cells and monocytes. The result could be an increased incidence of infectious complications. Some small series of patients have reported a higher incidence of infections, and in particular of viral infections, but others have not. Therefore, we compared the incidence of infection in 25 recipients of CD34-selected PBSC (CD34 group) and 75 case-matched recipients of unmanipulated PBSC (PBSC group) transplants. There were 52 males and 48 females with an average age of 52±11 yrs. They included 32 patients with non-Hodgkin’s lymphoma, 8 with Hodgkin’s disease, 40 with multiple myeloma and 20 with breast cancer, with 42 in first-line therapy and 58 in relapse, receiving their 1st (N=63) or 2nd (N=37) transplant. The number of CD34 cells infused as well as speed of engraftment were comparable in the two groups. There were no significant increase in the CD34 compared to the PBSC group for: actuarial incidence of infection (56 vs 49% at day 30, 70 vs 64% at 1 yr); proportion of patients with at least 1 (68 vs 68%), 2 (24 vs 17%) or 3 (0 vs 3%) infections; number of infectious events per patient before (1.32 vs 1.04) or after (0.59 vs 0.71) disease progression, before day 30 (0.84 vs 0.64), within days 31–100 (0.00 vs 0.09, p

Description: Destructive bone lesions due to osteolytic bone disease are a major cause of morbidity and mortality in multiple myeloma patients, occurring in more than 80% of cases. Underlying osteolytic bone disease is an uncoupling of the bone remodeling process, with an increased activity of osteoclasts and a decreased activity of osteoblasts. Current strategies to treat osteolytic bone disease focus on anti-resorptive agents, which do not rebuild bone loss. Src kinase has been implicated in both osteoclast and osteoblast function. In this study, we assessed the effect of Src inhibition with AZD0530 (saracatinib, Astra Zeneca) on the development of multiple myeloma and its associated osteolytic bone disease. We first determined Src family kinase expression in the multiple myeloma microenvironment and found that patient-derived myeloma cells express Src at low levels but disease stage does not correlate with Src expression levels. In accordance with the literature, Src mRNA expression was found to increase during osteoclast differentiation and decrease during osteoblast differentiation in publicly available microarray datasets. Next, we validated an inhibitory role of AZD0530 on osteoclast differentiation and function. At a pharmacological relevant concentration of 1 micromolar, AZD0530 inhibited the differentiation of RAW264.7 osteoclasts (Oc.N/FOV: 15.5+-1.6 treated vs. 53+-1.5 non-treated). AZD0530 treatment appeared to hamper efficient progenitor cell fusion and osteoclast polarization, reflected by a decrease of CTSK and DC-STAMP mRNA levels and a defective actin ring formation in treated cultures, which culminated in a complete inhibition of bone resorption. When assessing the effect of AZD0530 on osteoblast function we found that AZD0530 inhibits osteoblast differentiation, with a decreased expression of OSX and OCN, and alters osteoblast morphology. In vivo, AZD0530 did not alter myeloma cell bone marrow infiltration in both the 5TGM.1 (37+-6.3% AZD0530 treated vs. 25.2+-6.7% non-treated) and 5T2MM (26.1+-7.7% vs. 29.1+-6.4%) murine multiple myeloma models. However, bone health was significantly improved in both models following treatment with AZD0530. In the 5TGM.1 model multiple trabecular bone parameters were restored to levels observed in healthy control mice following AZD0530 treatment, including BV/TV (11.7+-0.3% treated vs. 6.4+-0.3% non-treated), Tb.N. (2.5+-6x10^-2/mm vs. 1.7+-9x10^-2/mm) and Tb.Th (46.2+-1micron vs. 37+-0.8micron). These results were confirmed in the 5T2MM model, which displays a more severe osteolytic bone disease. In addition, AZD0530 treatment resulted in an increase in cortical thickness (157.8+-0.8micron treated vs. 151.4+-0.7micron non-treated) and a decrease in the number and size of cortical lesions in 5TGM.1 mice. Finally, our findings were corroborated by histomorphometric analyses. In conclusion, we report a potent inhibitory effect of the Src inhibitor AZD0530 on the development of osteolytic bone disease in multiple myeloma. Our results indicate that AZD0530 exerts this effect via the modulation of both osteoclast and osteoblast function. These findings warrant further study of the feasibility and efficacy of AZD0530 to treat osteolytic bone disease in multiple myeloma patients. Disclosures No relevant conflicts of interest to declare.

Description: Abstract 2269 Poster Board II-246 Background. Acute graft-versus-host disease (GVHD) has remained a significant cause of nonrelapse mortality after allogeneic hematopoietic cell transplantation (HCT) with nonmyeloablative conditioning. The role of tumor necrosis factor-alpha (TNF-α) in the biology of acute GVHD following nonmyeloablative conditioning has not been studied thus far. Here, we measured TNF receptor 1 (TNFR1) as a surrogate marker for TNF-α in 106 patients before the start of the conditioning regimen (baseline) and 7 days after allogeneic HCT following nonmyeloablative conditioning. Patients and Methods. The nonmyeloablative regimen consisted of 2 Gy total body irradiation (TBI) alone (n=15), 2 Gy TBI plus fludarabine 90 mg/m2 (n=18). Postgrafting immunosuppression combined mycophenolate mofetil (MMF) with a calcineurin inhibitor for all patients. Blood samples were prospectively collected before the start of the conditioning regimen, then on days 7, 14, 21, 28, 35, 42, 49, 56, 63, 70, 77, 84, 91, and 98 after HCT, and then generally once every 2 weeks up to day 180. The serum component of each blood sample was separated and frozen for later analysis on the day of sample acquisition. TNFR1 serum concentration was retrospectively assessed using a cytokine enzyme-linked immunoabsorbent assay (R&D, Minneapolis, MN) according to the manufacturer's protocol. Results. TNFR1 levels increased significantly from baseline to day 7 after nonmyeloablative HCT (P

Description: Abstract 1149 Poster Board I-171 Background Nonmyeloablative conditioning followed by allogeneic hematopoietic cell transplantation (HCT) has been increasingly used as treatment for elderly patients with hematologic malignancies. It has been suspected that T cell reconstitution would be impaired in elderly patients given nonmyeloablative conditioning because of age-related thymic atrophy. Here, we investigated long term lymphocyte reconstitution and thymic function in 80 patients given allogeneic peripheral blood stem cells (PBSC) after nonmyeloablative conditioning. Patients and Methods Median age at transplant was 57 years (range 10-71). Conditioning regimen consisted of 2 Gy total body irradiation (TBI) with (n=46) or without (n=20) added fludarabine, 4 Gy TBI with fludarabine (n=6), or cyclophosphamide plus fludarabine (n=8). Thirty-three of the 80 patients received grafts from HLA-matched related donors, 22 from HLA-matched unrelated donors, and 25 from HLA-mismatched related or unrelated donors. PBSC were unmanipulated in 56 patients, CD8-depleted in 19 others, and CD34-selected in the remaining 5 patients. GVHD prophylaxis consisted of mycophenolate mofetil and cyclosporine or tacrolimus. Immune recovery was assessed between 1 and 8.5 years after HCT by signal-joint T-cell receptor excision circle (sjTREC) quantification (221 samples), and flow cytometry. Further, in order to demonstrate a potential thymic recovery, sjTREC level changes from day 100 to days 365 and 730 were also assessed by using Wilcoxon signed rank tests. Results There was a close correlation between sjTREC levels and naive CD4+ T cells (defined as CD4+CD45RA+) counts (P

Description: Introduction Myelodysplastic syndromes (MDS) are diagnosed at median age of 70 years. Allogeneic stem cell transplantation (HSCT) is the only curative treatment option, but with an increasing age, morbidity escalates. Treatment guidelines suggest HSCT for intermediate-II and high risk constellations up to the age of 65, and reduced intensity conditioning (RIC) regimens are commonly used up to 70 years of age. However, increasing life expectancy, availability of RIC regimens and good Karnofsky performance status (KPS) of MDS patients more than 70 years of age, has led to an increased use of HSCT. We performed a retrospective analysis to investigate results after HSCT for those patients and influence of KPS on outcome. Patients and methods We analyzed data of 345 patients in the EMBT database older than 70 years with MDS/sAML. The disease status at transplantation was available in 233 patients and most of the them were in more advanced stage of the disease: RA/RARS,RCMD (n=25) , RAEB (n=68) and RAEB-T/secondary acute leukemia (sAL, n=140). Donor were: related (n=88) and unrelated (n=257). Cytogenetic data were available only in 73 patients and classified as good (58), intermediate (6), poor (5) and very poor (4). Median follow up was 29.7 months. Median age at transplantation was 72 years (70-79 years) with 249 male and 96 female patients. KPS was defined in 300 cases, being 90-100% in 61% and 80% or less in 39%. Stem cell source was peripheral blood (94%) or bone marrow (6%). The intensity of the conditioning regimen was mainly reduced intensity (78%) rather than myeloablative (22%). Negative or positive CMV sero-status of the patient were seen in 35% and 65%, respectively. Results The number of HSCT for MDS patients of 70 years or more has increased over time. While 2000-2004 only 19 patients received transplantation, the following 3-year periods included 28 (2005-2007), 97 (2008-2010) and 200 (2011-2013) patients, respectively. The estimated 3-year OS was 33% (27-39%). A significant better 3 year OS in the univariate analysis was seen for Karnofsky (90-100%) vs 80% or less (41 vs 23%, p=0.008) and for CMV negative sero-status (46% vs 27%, p〉