bluuebunny/crossref_metadata_2025_split huggingface dataset:
Defective Adhesion, Migration and Homing Are Associated with Altered Rho GTPase Activity in Cells from Fanconi Anemia Patients.
Authors: Zhang Xiaoling , Shang Xun , Wang Lei , Guo Fukun , Zheng Yi , Pang Qishen 
Published: October 2019

Abstract
Fanconi anemia (FA) is a genetic disorder characterized by bone marrow failure and predisposition to malignancy. One of the potential therapeutic options for patients with FA is collection of autologous multipotent hematopoietic progenitors prior to the development of severe pancytopenia for autologous transplantation and gene therapy. However, poor engraftment of FA hematopoietic cells represents a major obstacle for effective transplantation. Our current study attempted to investigate the mechanism underlying defective engraftment of FA bone marrow (BM) cells. Using BM cells from patients carrying mutations in the FA complementation group A (FA-A), we demonstrate that SDF (Stromal cell-derived factor)-1alpha – and integrin-mediated migration and adhesion, respectively, is defective in FA primary BM cells compared to those from normal donors (more than 2-fold decrease in both migration and adhesion compared to normal BM cells). Complementation of the FA-A defect by retrovirus gene transfer of FANCA gene almost completely restores the ability of the BM cells to migrate towards the chemokine SDF-1alpha. Similar results are obtained with primary fibroblast cells derived from a FA-A patient, which show 3-fold and 35% decrease in adhesion and migration, respectively, compared to FANCA-corrected cells. Furthermore, when transplanted into immunodeficient Nod/scid recipient mice, the FA-A BM cells exhibited significantly impaired homing function whereas normal cells were efficiently homed in the bone marrow. A significant decrease in the activity of the Rho GTPase Cdc42 in FA-A cells is found associated with the patient cell defective functions. Taken together, these data suggest that the FA proteins play a role in the regulation of cell adhesion and migration in addition to maintaining genomic stability, influencing homing and engraftment, possibly via the small GTPase signaling pathway. These findings may have implications in development of strategies for restoring engraftment function of FA hematopoietic cells.
Multicenter Phase II Trial of Patients with Refractory or Recurrent Multiple Myeloma with Oral Treatment of Thalidomide Combined with Oral Cyclophosphamide, Idarubicin and Dexamethasone.
Authors: Schmidt-Wolf Ingo G.H. , Moehler Thomas , Goldschmidt Hartmut , Geer Thomas , Kleinschmidt Rolf , Gaska Tobias , Janssens Uwe , Schubert Joerg , Brueck Patrick , Clemens Michael R. , Bair Wolfgang , Seipelt Gernot , Schleuning Michael , Hahn-Ast Corinna , Herz Susanne , von Lilienfeld-Toal Marie , Naumann Ralph , Gorschlueter Marcus , Glasmacher Axel 
Published: October 2019

Abstract
Introduction and rationale: The aim of our study is to evaluate the role of thalidomide in combination with oral CID therapy in order to replace the central venous line necessary for continuous infusion of chemotherapy and to avoid hospitalization.
Patients and methods: Patients with refractory or recurrent myeloma were included into this trial. Concerning inclusion criteria, 9 patients had progressive disease after standard chemotherapy, 12 no change after standard chemotherapy, 6 recurrent disease after standard therapy and 21 recurrence after high dose chemotherapy. At start of the treatment patients were in stages IIA (15 patients), IIB (1) and IIIA (34). Idarubicin was given in capsules at 8–10 mg/m2/day for days 1–4, 40 mg dexamethasone were given on days 1–4 and 15–18, cyclophosphamide at 200 mg/m2/d d1–4 and thalidomide 100mg daily with scheduled increase to 400mg. Treatment cycles were repeated every 28 days. 3–8 cycles were applied. Supportive therapy consisted of PEG-filgrastim, cotrimoxazol, dalteparin and ibandronate. In addition, patients were randomized between thalidomide and thalidomide / idarubicin maintenance treatment.
Results: At this interim analysis, 64 patients have been entered into the ongoing trial; 8 patients had to be excluded. Of the 56 included patients, 50 patients have been documented so far and are evaluable for toxicity. In total, 182 cycles have been applied. Median cycle number was 3 (1–8). There was one death during treatment due to progressive myeloma at cycle 1. The median age was 62 years (interquartile range 57–67). The following toxicities were observed: FUO (2 patients), pneumonia (4), pulmonary embolism (1), exsiccosis after diarrhea (1), constipation (1), sepsis (1), severe nausea (2), syncope (1), collaps after bleeding (1) and mucositis grade 4 (1). Hematologic toxicity was reported in 34% of the patients, mainly in cycles 1 and 2. Concerning response to treatment 39 patients are evaluable so far. Of these 30 (77%) achieved a PR or MR according to EBMT criteria, 4 patients a stable disease and 2 patients a PD. Three patients were not evaluable and scorded as failures.
Conclusions: T-CID is feasible and seems to be highly effective in relapsed or refractory myeloma patients. Toxicity seems to be acceptable and mainly hematologic and infectiologic. The remission rate of 77% is encouraging. The trial is still ongoing. At the meeting a follow-up of the trial will be presented. Further information can be obtained at www.myelom.net.
Efficient Expansion of Lymphocytes in a Culture System with Solid Phase Anti-CD3 and Anti-CD28 Monoclonal Antibodies.
Authors: Soeda Atsuko , Kondo Atsushi , Makiyama Hiroaki , Morita Yuriko , Takahashi Noriko , Yagita Hideo , Takaue Yoichi , Heike Yuji 
Published: October 2019

Abstract
[Background] In vitro cell expansion in culture with solid phase anti-CD3-mab (aCD3mab) is a standard method for lymphocyte proliferation and activation. However, clinical trials using activated lymphocytes with this system provided only minimal efficacy. Recently, aCD3mab/aCD28mab-coated micro-bead (aAPC) was developed as a novel tool to efficiently expand lymphocytes, and has been used in several clinical trials. This study was designed to evaluate the effects of solid phase aCD3mab/aCD28mab on the proliferation and activation of lymphocytes to compare the results with culture using aAPC.
[Methods and Results] Parameters to evaluate the efficacy of culture system included magnitude of cell proliferation, killing activity against K562 and various surface markers of cultured cells. Peripheral blood was obtained from healthy volunteers after an informed consent was obtained, and mononuclear cells (PBMNc) were separated by lymphosepal system. Firstly, we cultured PBMNc in system with solid phase aCD3mab/aCD28mab coated to a culture plate (Culture A) or with solid phase aCD3mab coated to a culture plate plus soluble form aCD28mab (Culture B) to seek an optimal condition in the solid phase for lymphocyte proliferation (Figures 1). The results showed that the optimal ratio of aCD3mab:aCD28mab for Culture A is 3:7 to yield fold-expansion of lymphocytes of 12–13, while this in Culture B with the same amount of monoclonal antibodies was 10 (n=6). There was no difference in the expression CD3, CD4 or CD8, and killing activity between expanded cells in Culture A and Culture B. Then, we compared optimized solid phase Culture A with liquid phase culture with aAPC (Culture C). As a result, cell proliferation in Culture A was 1.3–5.6 times higher and the killing activity was 1.1–1.8 times higher than Culture C (Figure 2). In Culture A, the cell expression of CD45RO and GITR was higher and that of CD161 was lower compared to Culture C (Figures 3). Solid phase culture system with aCD3mab/aCD28mab is efficient and easily applicable for the expansion of lymphocytes, however more detail characterization of the expanded cells, especially Treg function and in vivo survival, are necessary.
Figure 1. Figure 1. Figure 2. Figure 2. Figure 3. Figure 3.
Human Mesenchymal Stem Cell Therapy: Platelet Lysate Supports Efficient Preclinical Expansion.
Authors: Schallmoser Katharina , Malischnik Christina , Rohde Eva , Reinisch Andreas , Guelly Christian , Lanzer Gerhard , Linkesch Werner , Strunk Dirk 
Published: October 2019

Abstract
Coinfusion of hematopoietic stem cells with multipotent mesenchymal stromal/stem cells (MSC) to facilitate engraftment and immunomodulatory treatment of graft versus host disease (GvHD) are the two foremost applications of MSC therapy in hematology and oncology. Translation of promising experimental results into clinical applications has been mainly hampered by the strict dependence of MSC propagation from fetal bovine serum (FBS)-derived factors.
We analyzed the capacity of different preparations of human platelet lysates (HPL) to replace FBS for clinical scale MSC production with cytomic, proteomic and genomic technology.
Using a previously established two-step good manufacturing practice procedure for MSC propagation, we found that HPL can fully replace FBS as a growth factor supplement for clinical scale MSC preparation. HPL was reproducibly more efficient than FBS and supported the outgrowth of >400 million MSC from <0.4 million primary MSC (derived from <2mL bone marrow aspiration within 10 days ex vivo) within only 2 additional weeks of culture. Although morphologically distinct, HPL-MSC and FBS-MSC did not differ regarding their CD13+/29+/73+/90+/105+/146+/34−/45−/133−/HLA-AB+ immunophenotype. Multiplex analyses allowed delineating a distinct cytokine and growth factor profile. Whole transcriptome microarray analyses revealed a differential regulation of >300 genes after only one cycle of human as compared to bovine growth factor stimulation. Allogeneic HPL-MSC infusion in an intent to treat refractory GvHD was feasible and without infusion-related side effects in one patient. This new FBS-free two-step procedure for clinical scale MSC propagation will largely facilitate rational clinical testing of MSC based therapies. Replacing FBS with HPL excludes bovine prion, viral and zoonose contamination of the stem cell product. Based on this protocol a clinical trial for the treatment of steroid-refractory GvHD with HPL-MSC has been initiated.
New Cytogenetic Abnormalities Are Frequent in AML and MDS Relapsing after Allogeneic Hematopoietic Cell Transplantation (HCT).
Authors: Pollyea Daniel A. , Artz Andrew S. , Stock Wendy , Daugherty Christopher , Godley Lucy , Odenike Olatoyosi M. , Rich Elizabeth , Smith Sonali M. , Zimmerman Todd , Zhang Yanming , Huo Dezheng , Larson Richard , Van Besien Koen 
Published: October 2019

Abstract
There are limited data regarding the incidence or prognostic value of cytogenetic abnormalities in pts with leukemic relapse after allogeneic hematopoietic cell transplantation (HCT). Between 2002 and 2005, 70 consecutive pts with high risk AML or MDS were transplanted with a reduced intensity preparative regimen of fludarabine 30 mg/m2/day IV (150 mg/m2 total), alemtuzumab SC 20 mg/day IV (100 mg total) D-7 to D-3, and melphalan 140 mg/m2 IV D-2, with tacrolimus given for post-transplantation immunosuppression. Twenty-five pts relapsed or progressed; 21 had AML, 3 had MDS and 1 had mast cell leukemia.
Twenty-two pts had cytogenetic analysis available prior to HCT and at relapse. Cytogenetic abnormalities were present in 12/22 (55%) pts prior to HCT. The median OS was 184 days (95% CI: 81 – 300) after relapse. Four pts with cytogenetic abnormalities prior to HCT reverted to a normal karyotype at relapse. Ten pts had no changes in their cytogenetics from HCT to relapse; they either remained normal or retained the same abnormality. Eight pts developed a new clonal abnormality at relapse, and had a median OS of 106 days (95% CI: 30 – 322). There was a non-significant trend toward inferior OS among pts with new abnormalities compared to the other groups (HR = 1.74, 95% CI 0.69 – 4.44, P = 0.24). The higher than previously reported rate of clonal evolution (8/22, 36%) may be due to the high prevalence of refractory disease at HCT in this cohort, more refined cytogenetic analysis, or regimen related factors (e.g. reduced intensity conditioning). The same clonal abnormality with or without new changes occurred in 7/22 pts. Thus, minimal residual disease monitoring in the subset of pts harboring pre-HCT karyotypic derangements may be a viable strategy for early detection and intervention. Our data suggest that clonal evolution at relapse of AML and MDS after HCT is relatively frequent, and in this small series, a trend toward worse outcomes exists for pts who develop new cytogenetic abnormalities. Larger studies are warranted to more completely characterize the prognostic value of cytogenetics and karyotypic evolution at relapse after HCT.
Cytogenetic abnormalities for AML/MDS relapsing after HCT (N = 22) Pre HCT* Relapse *History of cytogenetic abnormality any time before HCT **Clonal evolution in 8/22 (36%) No 10 (45%) No 7 (32%) Yes (New)** 3 (14%) Yes 12 (55%) No 4 (18%) Yes (Same) 3 (14%) Yes (New)** 5 (27%)
Expression of Proteinase Inhibitor-9 in Primary AML Blasts and Its Regulation by Interferon-γ: A Potential Immune Escape Mechanism after Allogeneic Stem Cell Transplantation.
Authors: Hoves Sabine , Kolbeck Alexandra , Mondal Krishna , Andreesen Reinhard , Mackensen Andreas 
Published: October 2019

Abstract
It is well established, that the curative potential of allogeneic peripheral blood stem cell transplantation (allo PBSCT) is due to immunocompetent donor T cells inducing potent anti-neoplastic effects against host tumor cells. This reaction, which is termed graft-versus-leukemia (GVL) effect, is clinically effective against a number of different hematologic malignancies such as myeloid and lymphoid leukemias. Despite great efforts of allo PBSCT in treatment of CML, the 5-year survival rate of AML patients after allo PBSCT is only about 30% due to relapsing disease. The recurrent disease is inefficiently controlled by the immune system, due most likely to the various immune escape mechanisms described for AML blasts including upregulation of anti-apoptotic molecules. Since cytotoxic T lymphocytes (CTL) and natural killer cells are the cells responsible for eliminating leukemic blasts, the most important effector molecule is Granzyme B (GrB). Misdirected GrB is quenched by its specific physiological inhibitor Protease Inhibitor-9 (PI-9) leading to inactivation of GrB. PI-9 expression by tumour cells can be used to escape immune surveillance and its presence has been shown for different tumors e.g. melanoma, colon carcinoma and lymphoma. Despite other regulators, interferon-γ (IFN-γ) has been shown to upregulate PI-9 expression in hepatocytes. Here, we wanted to investigate the expression of PI-9 in primary AML blasts and its regulation by IFN-γ. Using CD34+ positive magnetic selection, we isolated primary blasts with a purity of >90% from 20 AML patients with different FAB subtypes. For detection of PI-9 expression by Western Blotting, whole cell lysates were made from freshly purified blasts and after 24 h +/− 200 IU/ml IFN-γ. In some patients, PI-9 expression was confirmed by FACS analysis with an anti- PI-9 specific monoclonal antibody. Here we describe for the first time, that PI-9 is constitutively expressed in 16/20 (80%) of AML blasts. Treatment of AML blasts with IFN-γ could upregulate PI-9 expression in a dose-dependent manner (2–2,000 IU/ml) and strong expression of PI-9 was detectable in 6/18 patients within 4–5 h after IFN-γ exposure. Of note, a mild upregulation of PI-9 upon 24 h incubation w/o IFN-γ could be detected in 4/18 (22%) patients. We conclude, that cytokines such as IFN-γ which are secreted during the cytokine storm of acute graft-versus-host disease can contribute to the development of immune escape mechanisms in AML blasts.
Vaccine-Induced Anti-Neuroblastoma Immunity Early after HSCT Arises in the Presence of Foxp3+CD25+CD4+ T Cells.
Authors: Jing Weiqing , Orentas Rimas J. , Johnson Bryon D. 
Published: October 2019

Abstract
Effective anti-neuroblastoma immunity can be induced in mice as early as 1 week after syngeneic HSCT by vaccination with neuroblastoma cells transfected to express CD80, CD86, CD54, and CD137L. However, effective vaccination at this early time point also requires adoptive transfer of T cells (2×107 splenocytes). Previous results in non-transplanted mice showed that T-reg depletion/blockade with anti-CD25 mAb (clone PC61) increased tumor vaccine efficacy. To determine if PC61 treatment might be effective early after HSCT, we first determined if CD4+CD25+Foxp3+ T-reg cells were present, derived either from host (i.e., they survived lethal TBI) or from the transferred spleen cells. Even though transferred splenocytes accelerated T cell reconstitution, HSCT recipient mice remained severely lymphopenic until 3 weeks after transplant. Furthermore, the percentage of Foxp3+ cells in the splenic CD4 fraction was significantly increased in transplanted mice, being as much as 3 times higher than that in non-transplanted mice (35% vs. 12%). During this time, the majority of CD25+Foxp3+CD4+ cells were derived from the adoptively-transferred T cells. Despite the presence of CD25+Foxp3+CD4+ T cells, protective anti-tumor immunity could still be efficiently induced. To examine whether depletion/inhibition of CD25+ T-reg cells could further enhance vaccine-induced immunity in transplanted mice, two strategies were tested: (a) physical depletion of CD25+ regulatory T cells from the adoptively-transferred T cells, and (b) anti-CD25 mAb treatment of the transplanted mice 4 days after HSCT to target both transferred and residual endogenous CD25+ cells. Immunomagnetic depletion eliminated more then 95% of the CD25+ T cells in the transferred population, but only 70% of Foxp3+CD4+ T cells. Adoptive transfer of CD25-depleted splenocytes at the time of HSCT resulted in modest increased tumor vaccine efficacy as compared to mice given non-depleted splenocytes. Two weeks after HSCT, both groups of mice contained similar percentages of Foxp3+CD4+ T cells in the blood and lymph nodes, but a lower frequency of Foxp3+CD4+ T cells was detected in the spleens of mice given CD25-depleted splenocytes (35% vs. 43%, p<0.01). Our second strategy, treatment of HSCT recipients with anti-CD25 mAb prior to tumor vaccination, negatively influenced the vaccine-induced anti-tumor response and resulted in decreased survival. This suggested that the antibody treatment inhibited effector cell function in the early post-transplant setting. IFN-γ ELISpot analysis demonstrated that vaccinated transplant recipients given anti-CD25 mAb had significantly decreased numbers of tumor-specific CD4 T cells in spleen. Collectively, these results suggest that anti-neuroblastoma immunity could be induced in the present of CD25+Foxp3+CD4+ T-reg cells early after HSCT; strategies that targeted at CD25 did not significantly increase the efficacy of the tumor vaccine.
Chronic Hemolytic Anemia in Two Patients Double Heterozygotes for Pyruvate Kinase and Another RBC Glycolitic Pathway Deficiency.
Authors: Pissard Serge , Saddedine Nafa , Vasson Aurelie , Ballayguier Marie-Odette , Galacteros Frederic , Wajcman Henri 
Published: October 2019

Abstract
Homozygous or compound heterozygous for Pyruvate Kinase (PK) deficiency are classical etiology for chronic non spherocytic hemolytic anemias while heterozygous carriers are free from disease. We report here 2 patients heterozygous for PK deficiency which displayed an unexpected marked chronic anemia. Enzymatic and molecular studies were performed to unravel the mechanism causing this phenotype. The first patient, a 60-year-old woman from Mali presented with Hb 10g/dL, MCV 98fL, and was free from any Hb abnormality. The second one was the first child of a healthy French Caucasian couple, and suffered since birth from a marked hemolytic anemia (Hb 7g/dL). We found that the first patient carried together an Arg569Met/Leu PK-R mutation (Pissard et al. Brit J Haematol, 2006, 133, 683-9) and the rare G6PD Santa Maria mutation (nt c.542 A>T, Asp181Val). The second patient had a PK splice site mutation (IVS4 + 10 G>T) and a new hexokinase mutation [c.1793_c.1836 +7(del 50)] which starts in exon 12 and ends in intron 12. It results in a protein troncated inside the glucose binding site. In this case, family study showed that the PK deficiency was inherited from the father and the HK deficiency from the mother. Enzymatic data are shown in the table. None of these enzymatic defects could alone, in the heterozygous state, be responsible for an hemolytic anemia. To explain why, in these two cases the combination of two defects resulted in a hemolytic disease, we hypothesized that the increase of the intra-erythrocytic 2,3-DPG level resulting from the PK-R deficiency might cause these disorders. It is well known that increase of the 2,3-DPG level dramatically change several properties of the RBCs such as a decrease in oxygen affinity and an inhibition of the G6PD activity (Tomoda A. Brit J Haematol, 1983, 54, 475 – 84). It has been shown that, when associated to a sickle cell trait, PK-R deficiency by increasing 2,3-DPG leads to sickle cell anemia (Cohen.Solal M. et al, Brit J Haematol, 1998, 103, 950-6). We propose that, in these two patients, the global mechanism leading to the disease results from the increased 2,3-DPG which cause a failure in the anti oxidant pathway. In the first case G6PD inhibition occurs along with a mutated enzyme and in the second one inhibition take place in a under supplied pentose phosphate pathway due to the hexokinase deficiency. Together with the diminished ATP supply of the cell, this decreased anti-oxidant activity might cause the hemolysis. Thus in any anemic heterozygous PK deficient patient, another RBC abnormality needs to be searched for.
enzymatic data patients Pk activity (5.9–8.1) g6pd activity (5.3–7.9) hexokinase activity (0.74–1.14) 2.3 DPG (11.7–15.3) nd : not determined 1 3.2 UI/g Hb 4.5 UI/g Hb nd 21.2 μM /g Hb 2 6.8 UI /g Hb 9.2 UI /g Hb 0.3 UI / g Hb 36.8 μM /g Hb