bluuebunny/crossref_metadata_2025_split huggingface dataset:
Unexpected Evidence That Dimethylsulphoxide (DMSO) Upregulates Expression of CXCR4 on Hematopoietic Stem/Progenitor Cells (HSPC), Increases Their Responsiveness to an SDF-1 Gradient and Enhances Homing to Bone Marrow.
Authors: Majka Marcin , Jarocha Danuta , Wysoczynski Marcin , Sag Duygu , Zuba-Surma Ewa , Kijowski Jacek , Gozdzik Jolanta , Ratajczak Mariusz Z. 
Published: October 2019

Abstract
Cryopreservation of bone marrow (BM), mobilized peripheral blood (mPB) and cord blood (CB) cells is a routine procedure to store hematopoietic stem/progenitor cells (HSPC) for transplantation. Dimethylsulphoxide (DMSO), the most commonly used cryoprotectant, is toxic to cells at higher concentrations (>10%); moreover, the freezing-thawing procedure itself is inevitably connected with the loss of HSPC. However, by chance we observed that short exposure of HSPC to DMSO enhances the responsiveness of these cells to an SDF-1 gradient and since SDF-1 is a major chemoattractant that navigates homing of HSPC to BM we became interested in elucidating this phenomenon. We found that short incubation (5–10 min) of human CB mononuclear cells (MNC) with DMSO at concentrations employed for cryopreservation (5–10%) significantly upregulates the expression of both CXCR4 (x 2–3) and CD34 (x 1.5) on CB MNC (as measured by FACS). Furthermore, DMSO significantly increased the chemotactic responsiveness (x 2–4) of CB MNC, BM MNC and selected CXCR4+ human hematopoietic cell lines (Jurkat, THP-1 cells) when the cells were exposed to 5–10% DMSO before chemotaxis assay. These responses to an SDF-1 gradient correlated with enhanced chemotaxis also of human CD34+, CD34+ CD38+, CD34+ CD38−, and CD34+ CXCR4+ clonogeneic progenitor cells, suggesting that DMSO directly enhances the responsiveness of human early progenitors (p<0.0001). At the molecular level, 5–10% DMSO strongly stimulated and prolonged SDF-1-dependent AKT phosphorylation. However, at the same time DMSO inhibited phosphorylation of MAPKp42/44. Similar observations were made for Sca-1+ BM-derived murine cells. In parallel experiments we found that murine Sca-1+ cells when preincubated with DMSO formed more 12 day-CFU-S colonies in spleens after transplantation into irradiated syngeneic recipients. Accordingly, x 2 more CFU-S were formed when Sca-1+ cells were exposed before transplantation to 5% DMSO and about x 4 more after exposure to 10% DMSO. Finally we employed a Ly5.1/Ly5.2 congeneic transplant model and showed that transplantation of Ly5.1 Sca-1+ cells exposed to 10% DMSO before transplantation resulted in higher chimerism in transplanted Ly5.2 mice as compared to untreated cells (control) (p<0.0001). In conclusion, we show for the first time an unexpected beneficial role of DMSO (5–10%) in regulation of homing of HSPC after transplantation and suggest that a short priming of HSPC with DMSO, even of non-cryopreserved cells, before transplantation may become a new strategy to enhance engraftment
Bcr-Abl Resistance Screening Predicts a Limited Spectrum of Point Mutations To Be Associated with Clinical Resistance to the Abl Kinase Inhibitor AMN107.
Authors: von Bubnoff Nikolas , Saenger Jana , Manley Paul W. , Mestan Juergen , Peschel Christian , Duyster Justus 
Published: October 2019

Abstract
In advanced-phase CML, resistance to imatinib mesylate is frequently associated with point mutations in the Bcr-Abl kinase domain. New, highly potent Abl kinase inhibitors such as AMN107 and BMS-354824, have recently entered clinical trials. Data from analyses of resistant patients will be available not before a large number of resistant patients will have been treated within clinical trials. Therefore, it will be important to generate specific resistance profiles for each compound prior to its therapeutic application. Using a cell-based screening method for resistance of Bcr-Abl positive leukemia to Abl kinase inhibitors, we generated a resistance profile for AMN107 and compared it to the resistance profile of imatinib mesylate. In contrast to imatinib, resistance to AMN107 was associated with a very limited spectrum of Bcr-Abl kinase mutations. While 26 exchanges at 21 positions occured with imatinib, the AMN107 screen revealed eight different exchanges at seven amino acid positions, with four exchanges affecting the P-loop. Novel mutations which have never been observed with imatinib, either in vitro or in resistant patients, emerged in the presence of AMN107 including an F359 exchange to isoleucine and a Q252H/S349L double mutant. In contrast to imatinib, the frequency of resistant colonies dramatically decreased with increasing AMN107 concentrations. Rarely emerging resistant colonies at a concentration of 400 nM AMN107 exclusively contained T315I. With the exception of T315I, all mutations that were identified were effectively suppressed when AMN107 was increased to 2000 nM, a concentration which is achieved in plasma in treated patients. Thus, in this system, increasing the AMN107 concentration to 400 nM prevented the emergence of resistant colonies, with the exception of T315I. Our findings suggest that AMN107 might be superior to imatinib in terms of the development of resistance. Also, AMN107 at clinically relevant concentrations may overcome imatinib resistant disease, including cases with expression of P-loop mutations. However, our study indicates that clinical resistance to AMN107 may be associated with the predominant emergence of T315I. Using this or similar approaches, it will be possible to provide information that translates into combinatorial and sequential treatment strategies and to determine critical plasma concentrations for mutations that might occur during treatment.
SPTBN1-FLT3 in Atypical Chronic Myeloid Leukemia Transforms Ba/F3 Cells to IL-3 Independence and Is Sensitive to Both Tyrosine Kinase Inhibitors and Immunotherapy.
Authors: Grand Francis H. , Iqbal Sameena , Zhang Lingyan , Russell Nigel H. , Chase Andrew , Cross Nicholas C.P. 
Published: October 2019

Abstract
We have identified a patient who presented with BCR-ABL negative chronic myeloid leukemia (CML) and an acquired 46XX, t(2;13;2;21) (p13;q12;q33;q11.2) in all bone marrow metaphases examined. Fluorescence in situ hybridization (FISH) using probes flanking the FLT3 gene at 13q12 suggested that this gene was disrupted. 5′-RACE PCR using primers to the region of FLT3 encoding the tyrosine kinase domain identified a novel in-frame mRNA fusion between exon 3 of SPTBN1 (spectrin, beta, non-erythrocytic 1 isoform 2, NM 178313) on chromosome 2p16 and exon 13 of FLT3 (NM 004119). Juxtaposition of SPTBN1 and FLT3 was confirmed by two color FISH and amplification of the genomic DNA breakpoint confirmed a fusion between intron 3 of SPTBN1 and intron 12 of FLT3. The SPTBN1-FLT3 fusion gene is predicted to be translated into a 570 amino acid chimeric protein that retains two coiled-coil domains from SPTBN1 and 424 amino acids from FLT3, including the entire tyrosine kinase domain. Since the t(2;13) is readily visible by cytogenetic analysis but has not been reported previously it seems likely that SPTBN1-FLT3 is uncommon. However to test if FLT3 might be involved more widely in BCR-ABL negative CML we analysed 40 cases by RT-PCR. Two cases were positive for the FLT3 internal tandem duplication (ITD) but mutation of residue D835 was not observed. Expression of the SPTBN1-FLT3 fusion transformed the interleukin 3 (IL-3)-dependent cell line Ba/F3 to growth factor independence and was accompanied by constitutive phosphorylation of the fusion protein and the downstream substrate ERK1/2. The growth of transformed cells was inhibited in a dose-dependent fashion by SU11567 and PKC142, but not by imatinib mesylate. The patient was initially treated with hydroxyurea and subsequently underwent an unrelated donor bone marrow transplant. She relapsed cytogenetically at 4 years but responded to donor lymphocyte infusion (DLI), achieving sustained cytogenetic and molecular (nested RT-PCR) remission. We conclude that SPTBN1-FLT3 is a rare abnormality in BCR-ABL negative CML that is responsive to both targeted signal transduction therapy and immunotherapy by DLI.
Early Bortezomib Following Allogeneic Stem Cell Transplantation (SCT) for Multiple Myeloma To Enhance or Maintain Remission Status.
Authors: Kroeger Nicolaus , Zabelina Tatjana , Ayuk Francis , Atanackovic Djordje , Schieder Heike , Renges Helmut , Zander Axel R. 
Published: October 2019

Abstract
We investigated the toxicity and efficacy of early administration of bortezomib in 14 patients with residual disease (n=9) or complete remission (immunofixation-negative) (n=5) after a median of 383 days (range, 47–1317) after melphalan/fludarabine-based dose-reduced allogeneic SCT for multiple myeloma. The median age was 49 years (range, 32–62), and stem cell graft was from HLA-identical siblings (n=2) or unrelated donors (n=12). Three patients had received prior Thalidomide and donor lymphocyte infusion, and 2 of them still received low dose thalidomide (50 mg) during bortezomib treatment. The study consisted of two cycles bortezomib (1.3 mg/m2) given intravenously on day 1, 4, 8 and 14 (q21d). In case of response and low toxicity, two further cycles were applied. Eleven patients completed two cycles, and 4 patients completed four cycles. Two patients stopped after three cycles due to side-effects; 3 patients could not complete the proposed two cycles and stopped either after the first injection (n=1) of the first cycle, or after the second (n=1) or third (n=1) injection of the second cycle due to neurotoxicity. Major toxicity was thrombocytopenia grade IV (> 75 % reduction) which was seen in 6 patients (43 %), but none required platelet support. Neutropenia grade IV was seen in 2 patients (14 %), neurotoxicity grade III and IV was observed in 3 patients (21 %) while mild grade I-II neuropathy was seen in 10 patients (71 %). Other toxicity was fatigue grade I (85 %), nausea grade I (50 %), diarrhea grade I-II (93 %). Two patients with acute or chronic GvHD of the skin experienced deterioration of the manifestation, which could be treated succesfully with corticosteroids. A significant decrease during bortezomib-therapy was observed for CD3+ cells (p=0.02), while no effect was seen for B-cells (CD19+) (p=0.4). Two patients who received bortezomib on day 125 and 201 after allogeneic SCT experienced Herpes zoster infection during and 2 weeks after bortezomib treatment, respectively. All 9 patients with persistent disease showed further regression of the paraprotein including negative immunofixation in 5 patients. We conclude that early bortezomib after allogeneic SCT for patients with multiple myeloma is active and can induce further regression of the paraprotein. The toxicity is considerable especially if given early after allo SCT.
High Frequency and Early Onset of Bone Mineral Density Loss Following Allogeneic Stem Cell Transplantation.
Authors: Dunford Lauren M. , Roy David M. , Hahn Theresa , Brown Karen , Paplham Pamela , Lamonica Dominick M. , Battiwalla Minoo , Padmanabhan Swaminathan , McCarthy Philip L. 
Published: October 2019

Abstract
Bone mineral density (BMD) loss has been recently associated with allogeneic stem cell transplantation (alloSCT). It is not certain whether BMD loss in alloSCT patients (pts) occurs primarily within the first six months, since some reports cite continuing BMD loss up to six years post-transplant. Furthermore, it has been suggested that alloSCT-BMD loss is more likely to manifest in the femoral neck (FN), as opposed to the lumbar spine (LS). There are no studies that have detailed the incidence of alloSCT-BMD loss. We retrospectively reviewed 90 alloSCT pts who underwent a single alloSCT between 7/1/2001 and 2/26/2005 to assess the frequency of DEXA scan use and alloSCT-BMD loss. 7/1/2001 was chosen as the start date since this was when DEXA scans were first available at RPCI. 50 patients did not have DEXA scans following BMT: 15/50 pts died before day 130, 21/50 pts died after day 130 post BMT, and 14/50 pts are alive post day 130 without DEXA scans (median 1.1 yrs, range 0.34–3.0 yrs). Clinical criteria for obtaining DEXA scans were a medically stable condition with a good KPS and alive at day 130. 37 pts met this criteria. The median survival of the non-DEXA scan pts was 180 (7–1296) days and 571 (110–1282) days for the DEXA scan pts. DEXA scan pt characteristics in this sample included: Median age 39 years (range 7–66); AML (n=12), Lymphoma (n=12), CML (n=5), MDS/MPD (n=5), ALL (n=2), or Aplastic Anemia (n=1); Flu+Mel (n=14), Bu+Cy (n=8), Cy+TBI (n=7), non-myeloablative regimen (n=6), or other regimens (n=2); FK+Mt (n=28), FK+/− other (n=4), Mt (n=2), Cs+Mt (n=2), or Cs+FK+Mt (n=1). Approximately half (n=18/37) of alloSCT pts were found to have BMD loss at day 130 post-transplant. Results were stratified by year, and no trends regarding the frequency of DEXA scans in the population or the frequency of abnormal results over time were noted. There were no differences in terms of conditioning regimens or GVHD prophylaxis between the normal and low BMD groups. Contrary to previously published reports, we did not find a marked difference between the LS and FN T scores of the 18 pts with BMD loss at baseline (LS median T score = −1.4, FN median T score = −1.55). Also, BMD improvement following oral bisphosphonates was seen in six pts with available follow-up scans (LS median T score improved from −1.15 to −0.55, FN median T score improved from −1.45 to −1.1). The day 130 incidence of BMD loss in our alloSCT population was higher than we expected. Therefore, we recommend that all future alloSCT pts undergo DEXA scans as part of the day 100 workup. Further study may elucidate risk factors for BDM loss post BMT. Bisphosphonate use may reverse alloSCT BDM loss.
PI3K/Akt/FOXO3a Pathway Contributes to Thrombopoietin-Induced Proliferation of Primary Megakaryocytes In Vitro and In Vivo Via Modulation of p27Kip1.
Authors: Nakao Takafumi , Geddis Amy E , Fox Norma E. , Kaushansky Kenneth 
Published: October 2019

Abstract
Thrombopoietin (TPO), the primary regulator of megakaryocyte (MK) and platelet formation, modulates the activity of multiple signal transduction molecules, including those in the Jak/STAT, p42/p44 MAPK, and phosphatidylinositol 3-kinase (PI3K)/Akt pathways. In the previous study, we reported that PI3K and Akt are necessary for TPO-induced cell cycle progression of primary MK progenitors. The absence of PI3K activity results in a block of transition from G1 to S phase in these cells (Geddis AE et al. JBC2001276:34473–34479). However, the molecular events secondary to the activation of PI3K/Akt responsible for MK proliferation remain unclear. In this study we show that FOXO3a and its downstream target p27Kip1 play an important role in TPO-induced proliferation of MK progenitors. TPO induces phosphorylation of Akt and FOXO3a in both UT-7/TPO, a megakaryocytic cell line, and primary murine MKs in a PI3K dependent fashion. Cell cycle progression of UT-7/TPO cells is blocked in G1 phase by inhibition of PI3K. We found that TPO down-modulates p27Kip1 expression at both the mRNA and protein levels in UT-7/TPO cells and primary MKs in a PI3K dependent fashion. UT-7/TPO stably expressing constitutively active Akt or a dominant-negative form of FOXO3a failed to induce p27Kip1 expression after TPO withdrawal. Induced expression of an active form of FOXO3a resulted in increased p27Kip1 expression in this cell line. In an attempt to assess whether FOXO3a has an effect of MK proliferation in vivo, we compared the number of MKs in Foxo3a-deficient mice and in wild type controls. Although peripheral blood cell counts of erythrocytes, neutrophils, monocytes and platelets were normal in the Foxo3a-deficient mice, total nucleated marrow cell count of Foxo3a-deficient mice were 60% increased compared with wild type controls. In addition, the increase of MKs was more profound than that of total nucleated marrow cells; CD41+ MKs from Foxo3a-deficient mice increased 2.1-fold, and mature MKs with 8N and greater ploidy increased 2.5-fold, compared with wild type controls. Taken together with the previous observation that p27Kip1-deficient mice also display increased numbers of MK progenitors, our findings strongly suggest that the effect of TPO on MK proliferation is mediated by PI3K/Akt-induced FOXO3a inactivation and subsequent p27Kip1 down-regulation in vitro and in vivo.
Hematocrit and Oxygen Dependence of Nitric Oxide Scavenging by Red Blood Cells.
Authors: Kim-Shapiro Daniel B. , Azarov Ivan , Huang Kris T. , Hogg Neil , Gladwin Mark T. 
Published: October 2019

Abstract
Human, animal and in vitro studies suggest that reactions involving nitric oxide (NO) and cell-free hemoglobin (Hb) occur up to 1000 times faster than those with red blood cell (RBC) encapsulated hemoglobin. Thus, cell-free Hb resulting from hemolysis or infusion upsets vascular homeostasis and contributes to pathological conditions in hemolytic anemias such as sickle cell disease and paroxysmal nocturnal hemoglobinuria. The slow NO uptake by RBCs results either from internal factors such as a physical barrier set up by the RBC membrane or external factors such as the time required for NO to diffuse to the cell. We have conducted experiments in which both external and internal factors play a role in limiting NO uptake wherein excess NO is rapidly mixed with RBCs and NO uptake is monitored by absorption spectroscopy. In addition, we have measured NO uptake under conditions where the role of the external factors are reduced by having cell-free and RBC encapsulated Hb compete for limited amounts of slowly released NO, measuring the reaction products using electron paramagnetic resonance spectroscopy. We are the first to have conducted these types of measurements at physiologically relevant hematocrits. Our major findings are that (1) the relative rate that deoxygenated red blood cells scavenge nitric oxide is 3–8 times faster than oxygenated red cells; surprisingly, this effect appears to be partially attributable to a physical membrane barrier, and (2) the rate of NO scavenging by oxygenated RBCs increases significantly (about 3 fold) when the hematocrit is raised from 15% to 50%. Our biophysical data support the thesis that cell-free Hb is capable of greatly reducing the bioavailability of NO and thereby negatively effect hemodynamics and endothelial function. The hematocrit dependence of NO scavenging by intact erythrocytes provides a potential pathogenic mechanism for hypertension during erythropoietin therapy, mortality associated with eNOS inhibition in polycythemic mice, and endothelial dysfunction in acquired polycythemic conditions. The observed oxygen dependence of NO uptake rate supports models in which the submembrane red cell protein scaffolding contributes to NO homeostasis, but not to the extent that has been previously proposed by others. Modulation of the effects of the scaffolding on scavenging and export of NO activity by RBCs under normoxic and hypoxic conditions requires further investigation.
Is Antilymphocyte Globulin Useful in Conditioning Regimen of Unrelated Donor Hematopoïetic Stem Cell Transplantation?.
Authors: Esperou Helene , Appert Marie Lorraine , Agha Ibrahim Yacoub , Milpied Noel , Ifrah Norbert , Kuentz Mathieu , Chir Zina , Raffoux Colette 
Published: October 2019

Abstract
Between 1998 and 2004, 2141 allogeneic hematopoietic stem cell transplantations with unrelated donor (excluding cord blood) have been recorded on the SFGM-TC data base. The lack of unique behavior in regard of Anti Lymphocyte Globulin (ALG) use in unrelated donor transplants allowed us to perform a retrospective study to assess the influence of ALG in such grafts. To avoid biaises we excluded T cell depletion or monoclonal antibody use, reduced intensity conditioning and focused only on patients (pts) with acute leukemia (AL) and myelodysplasia (MDS). So we defined a study population of 553 pts. In addition, in order to analyze the actual impact of ALG on wellknown targets (engraftment, acute GvHD, relapse, survival) we choose to keep only the cases in which was available the allelic HLA typing (4 digits) for the recipient and the donor, i.e. 250 pts. There were 106 females and 144 males, allografted in 29 centers. The median age was 23 years (1–62) and there were 77 children under 15. There were 223 AL - myeloid n=120, lymphoid n=103 - and 27 MDS. The source of cells was bone marrow for 190 pts and peripheral mobilized stem cells for 60. One hundred and ninety-three pts received a conditioning regimen including Total Body Irradiation. Ninety-five pts received ALG (mainly Thymoglobulin Mérieux-Genzyme Lyon France at various doses) for a median of 3 days (group 1) and 155 did not (group 2). There were 189 donor/recipient pairs strictly HLA-matched 10/10, 49 with one allelic difference (HLA A n=13; HLA B n=8; HLA C n=13; HLA DRB1 n= 5; HLA DQB1 n=10 ), 11 with 2 allelic differences and 1 with 3 allelic differences. 238 pts have sustained engraftment at a median time of 20 days (9–41) without any influence of ALG use (97% vs 94% NS). Overall survival at 3 years was 45% in group 1 and 52% in group 2 (NS). The incidence of acute GvHD (grade II to IV) was similar in the two groups (59% vs 54%), but the incidence of acute GvHD (grade III to IV) was lower in group 1 (21% vs 32% p=0.03). Use of ALG did not affect rate relapse (18% vs 20% NS). In a multivariable analysis including HLA disparities, patient age, use of ALG, two factors appear as predictive for grade III–IV GvHD: HLA-identity (p=0.05) and use of ALG (p=0.02). These factors are not significant for survival in the same multivariable analysis but in this allelic matched population, the influence of ALG should be analyzed in regard of exact dose of ALG.