bluuebunny/crossref_metadata_2025_split huggingface dataset:
Philadelphia Negative BCR/ABL1 Positive CML: FISH Mapping and aCGH Reveal Breakpoint Hotspots.
Authors: Virgili Anna , Brazma Diana , Grace Colin , Chanalaris Anastasios , Reid Alistair , Apperley Jane F. , Nacheva Elisabeth P. 
Published: October 2019

Abstract
Chronic myeloid leukaemia (CML) is a pluripotent haematopoietic stem cell disorder defined by expression of the BCR/ABL1 fusion gene, a constitutively activated tyrosine kinase. The fusion gene is a result of t(9;22)(q34;q11) or related variant rearrangements leading to formation of the Philadelphia chromosome (Ph). In less than 2% of the CML patients the bone marrow cells appear to have a normal karyogram although the fusion gene can be identified by molecular means and located by FISH on 22q11 and less frequently on 9q34 or on a third chromosome. The biology and clinical significance of the genetic rearrangements in Ph negative BCR/ABL1 positive disease are controversial and widely discussed. Two alternative mechanisms have been postulated but not confirmed: simple insertion or multiple step model. Although sequencing analysis as early as 1990 (Morris and Heisterkamp) showed that a large part of the 9q34 region is present in the masked Ph, little has been done to clarify the underlying rearrangements. Aided by genome array CGH and FISH, we constructed a comprehensive map of the regions flanking ABL1 and BCR genes in 9 patients and one cell line with this form of CML. The main results show:
the formation of the BCR/ABL1 fusion is associated with relocation of sequences well in excess of the ABL1 and BCR gene in all 10 samples, the involvement of downstream 3′ ABL1 is diversely associated with distal breakpoint clusters in the regions of the RAP-GEF1 and RXRA genes, the disease progression can be accompanied by sequential breakages/reunions within the regions flanking the BCR/ABL1 fusion and the presence of multiple cryptic rearrangements of both 9q34 and 22q11 segments identifies a sub-group of patients with shorter survival thus demonstrating the clinical significance of genome instability in Ph negative, BCR/ABL1 positive CML.
MicroRNAs as the Target of Deletions on der(9) in Chronic Myeloid Leukemia.
Authors: Albano Francesco , Anelli Luisa , Zagaria Antonella , Pannunzio Alessandra , Rossi Antonella Russo , Manodoro Floriana , Liso Vincenzo , Rocchi Mariano , Specchia Giorgina 
Published: October 2019

Abstract
Deletions on der(9) are associated with chronic myeloid leukemia(CML) in 15–18% of cases. To date, the biological significance of this genomic loss in the pathogenesis of CML is unknown. The most plausible hypothesis is that the loss of a tumor suppressor gene may confer a proliferative advantage to the Philadelphia-positive clone. On the other hand, it has now become evident that microRNAs (miRNAs) play an important regulatory role in some hematological malignancies. To investigate the presence of miRNAs within the genomic regions lost on der(9) we analyzed 60 CML patients with der(9) deletions. Methods. Genomic characterization of the deleted sequences was performed by fluorescence in situ hybridization (FISH) using a contig of DNA clones; the miRBase (http://microrna.sanger.ac.uk/) was queried to assess the presence of miRNAs in the der(9) deleted genomic regions. FISH experiments showed that the genomic loss on der(9) of the 9 (centromeric to ABL) and 22 (telomeric to BCR) chromosome sequences ranged from 260 Kb to 54 Mb and from 230 Kb to 12.9 Mb, respectively. Consultation of the miRBase revealed that in 16 (27%) patients there was loss of miRNAs mapping on chromosome 9 whereas no known miRNAs were mapped on the deleted genomic sequences belonging to chromosome 22. Moreover, 4 cases with a complex t(9;22) rearrangement and der(9) deletions showed loss of the miRNAs sequence also on the third derivative chromosome (4p16, 7p14, 13q14, and 11q13, respectively); among them, only in one case the loss of miRNAs on the third derivative was not associated with the miRNAs deletion mapped on chromosome 9. The most recurrent miRNAs deleted on der(9) were mir-219–2 (deleted in 100% of cases) and mir-199-b (lost in 67% of cases). It is noteworthy that mir-219–2 neighbors and overlaps CpG-islands, suggesting a potential role of this miRNA in CpG-island methylation. Experimental studies indicate that miRNAs can function as tumor suppressor genes or as oncogenes. In fact, in chronic lymphocytic leukemia associated with del(13)(q14) it has been demonstrated that the miRNAs loss can induce downregulation of the antiapoptotic BCL-2 protein. The novel evidence that deletions on der(9) in CML are associated with miRNAs loss may shed new light on the significance of genomic sequences loss. Further studies are needed since it is known that some microRNAs may have as many as a few thousand targets, so prediction algorithms and strategies allowing large-scale screening of multiple target genes are required.
NM23-H2 Is a Tumor Associated Antigen (TAA) in Chronic Myeloid Leukemia (CML).
Authors: Tschiedel Sabine , Jilo Annette , Niederwieser Dietger W. 
Published: October 2019

Abstract
Introduction: The human Nm23 family consists of 8 genes, termed Nm23-H1 through -H8. H1 and H2 are 88% identical and localized to chromosome 17. Both Nm23-H1 and -H2 display nucleotide-diphosphate kinase (NDPk) activity responsible for balancing NTP levels by transferring gamma-phosphates between nucleotides, and individually demonstrate a variety of other activities including DNA binding, transcriptional regulation and apoptotic cleavage of DNA. A variety of studies imply a role for Nm23 proteins in development and in a range of cancers, in which level of expression rather than mutation correlates to prognosis. Specifically, a mutual transcriptional activation between the nm23-H2 and c-myc genes has been implicated in transformation. Our group described the identification of Nm23-H2 as a candidate tumor-associated antigen in a case of CML and demonstrated the presence of Nm23-H2 reactive T cells 5 years after transplantation. Here, we compare the expression of Nm23-H2 and H1 proteins in leukaemic patients and healthy donors.
Materials and Methods: PBMC were collected from 10 CML patients at initial diagnosis, from 4 Glivec-resistant CML patients, from 12 AML patients at diagnosis and from 5 healthy donors. PBMC, CD3+, CD14+, CD33+ and CD34+ populations were stained, fixed and sorted by FACS onto microscope slides. Immunocytochemical analysis was performed with Nm23-H1 (Novacastra), Nm23-H2 (Santa Cruz) and β-Actin (Sigma) specific antibodies. Evaluation was carried out using a confocal laser scanning microscope LSM 510 (Zeiss).
Results: PBMC from 9 out of 10 untreated CML patients (90%) showed a strong Nm23-H2 staining. Fractionation of the PBMC population revealed that CD14+, CD33+ and CD34+ but not CD3+ cells are Nm23-H2 positive in the majority of these patients. Nm23-H2 was also detected in the PBMC of 3 out of 4 Glivec-resistant CML patients. In most cases, Nm23-H2 co-localized with β-actin. Under the same staining conditions, PBMC and sorted subpopulations from 11 of 12 AML patients (92%) and from all healthy donors were Nm23-H2 negative, while very weak staining was detected in PBMC of one AML patient. Nm23-H1 protein was clearly detectable in a number of cell lines (HL-60, U-937, K562 and T47D), but was very weak or absent in PBMC from CML patients and healthy donors.
Conclusion: Taken together, our results show that those myeloid populations functionally affected by the bcr/abl mutation specifically over express Nm23-H2 but not Nm23-H1 protein in 9 out of 10 cases of CML. While the molecular basis of this expression pattern and the possible involvement of c-Myc remains to be resolved, these results support the proposal that immune responses to Nm23-H2 may mediate GvL effects and that peptides derived from Nm23-H2 might therefore be attractive targets for specific immunotherapy in CML patients.
Immune Effects of Imatinib in Chronic Myelogenous Leukemia In Vitro.
Authors: Bund Dagmar , Buhmann Raymund , Kolb Hans-Jochem 
Published: October 2019

Abstract
Imatinib mesylate, a potent and selective inhibitor of the BCR-ABL tyrosine kinase, has been shown to induce durable haematological and major cytogenetic responses in a high percentage of CML patients. However in most patients the disease recurs, when imatinib is discontinued. In contrast, allogeneic stem cell transplantation (ASCT) is considered to be curative by the immune effect of donor T cells against CML progenitor cells. In this context, the role of imatinib is controversial; it may improve the results of ASCT by reducing the tumour load, it may also reduce the effect of donor lymphocyte transfusions (DLT) by impairing the function of T cells and the capacity of myeloid cells to present antigen. Patient derived CML-cells were studied for the stimulation of allogeneic HLA-matched and mismatched T-cells in the presence and absence of imatinib. In a 5 day culture the proliferative response of HLA-mismatched T cells was evaluated in presence of different concentrations of imatinib (0, 1, 2, or 5 micro M) and various responder-to-stimulator ratios. Thereby, proliferation was detected via a CFDA based assays and the activation profile (CD25, CD69) of the T-cells was determined by FACS. Cr51-release assays were performed after a 7 day culture of CML cells with HLA-matched T cells to test cytotoxicity of CD8+ T-cells. In addition, we characterized the antigen-presenting profile (CD14, CD33, HLA-DR, CD40, CD80, CD86, CD54, CD58) of the CML cells over a 5 day culture with and without imatinib. The presence of imatinib inhibited the proliferative capacity of allogeneic T-cells in a dose-dependent manner. Also, the expression of T cell activation markers was reduced in the presence of the different imatinib concentrations. Preincubation of CML cells with imatinib for 48 hours strengthened the effect on proliferation and activation of T cells. Moreover, imatinib impaired the cytotoxic function of T-cell (HLA-matched setting; CR51-release assay) also in a dose-dependent manner. Finally, the antigen-presenting profile of the myeloid leukemia cells was down regulated by increasing concentrations of imatinib. In summary, imatinib may interfere with the T cellular immune response and the antigen presenting profile on the CML cells in vitro. These results may have an impact on new strategies of treatment of CML with immunotherapy.
Efficacy of Imatinib Mesylate Dose-Escalation in CML Patients Showing Suboptimal Response to Standard Dose.
Authors: Kim D.Y. , Kim H.J. , Jeong J.S. , Kim S.J. , Joo Y.D. , Sohn S.K. , Kim I. , Kim J.S. , Song H.S. , Yoon S.S. , Kim C.S. , Jung C.W. , Yi K.H. , Min Y.H. , Hyun M.S. , Lee N.R. , Chang D.Y. , Jo D.Y. , Kim J.A. , Park J.W. , Kim B.K. , Park S. 
Published: October 2019

Abstract
Introduction: We report the results of our prospective single arm study performed to evaluate the efficacy of Imatinib mesylate (IMT) dose-escalation therapy and the influence of BCR/ABL mutational status on the treatment outcome.
Methods: Patients with chronic phase(CP) chronic myelogenous leukemia (CML) who had not achieved a complete hematologic response (CHR) after 3 months, a major cytogenetic response (MCyR) after 6 months, nor a complete cytogenetic response (CCyR) after 6-12 months of standard IMT therapy (400mg per day) were included. Patients in accelerated phase (AP) or blast crisis (BC) who had failed to achieve CHR after 3 months of IMT (400 or 600mg per day) were also eligible. 600mg of IMT had been administered for patients with CP CML per day, and 600mg or 800mg for AP or BC CML for at least 12 months. For each patient, BCR/ABL gene mutation test was performed on 6 loci. Cytogenetic response (CyR) was evaluated with the percentage of bcr/abl positive cells in bone marrow aspirate by means of FISH per 6 months. Molecular responses (MR) were assessed per 3 months with a logarithmic decrease in BCR-ABL/ABL gene ratio (BA/A) of peripheral blood or bone marrow aspirate from a standardized median value of BA/A in 56 patients with newly diagnosed CML.
Results: 77 patients were enrolled and 61 patients were assessable for the evaluation of MR. The median age was 50 years old (range: 20-70). Stages at the time of the enrollment were CP for 56, AP for 4, BC for 1 patient(s). The median time from the initiation of IMT therapy to dose-up was 20 months (range: 0-62). BCR/ABL gene mutational status could be obtained in 24 patients (wild:mutant=20:4). The median follow-up was 19.9 months (range: 6.5-24.3). For 34 patients whose 6th-month CyR is available, MCyR rate (MCyRR) was 26.5% and CCyR rate (CCyRR) was 23.5%. The 12th-month MCyRR was 61.1% and CCyRR was 50.0%. Patients with wild type showed a MCyRR of 20% and a CCyRR of 15% on the 12th month after dose-up, whereas no patient with mutant type showed a CCyR or a MCyR. CMR rate and MMR rate on the 12th-month were 16.7% and 26.7%. Among 20 patients with wild type, one patient achieved a 6th-month CMR, 2 patients MMR’s, whereas no patient with mutant type could achieve a MMR during the entire treatment period. Hematologic and non-hematologic toxicities were tolerable.
Conclusion: IMT dose-escalation could induce quite a number of durable CyR’s and MR’s without any serious toxicity in patients who had failed to achieve either a CHR or a CyR with a standard IMT therapy.
Human Bone Marrow as a Source of Multifunctional CMV-Specific CD4+ T Cells for Adoptive Cell Therapy.
Authors: Na Il-Kang , Letsch Anne , Noack Ines , Bauer Sandra , Geginat Jens , Reinke Petra , Volk Hans Dieter , Thiel Eckhard , Scheibenbogen Carmen 
Published: October 2019

AbstractIntroduction Adoptive cell transfer of ex vivo primed and expanded human cytotoxic T lymphocytes (CTLs) has emerged as a promising approach to treat both infectious and malignant diseases in humans. First clinical studies have shown that transfer of Cytomegalovirus (CMV)-specific CD8+ T cells is safe and effective in reconstitution of cellular immunity against CMV disease. Efficacy of adoptive T cell therapy is limited by the numbers of CTLs in vitro and the survival and function after infusion. CD4+ T cells may enhance activity via direct or indirect effector functions (Matloubian 1994 [1]). In this study we have analysed whether bone marrow is superior to peripheral blood for expansion of CMV-specific T cells. Experimental design Paired peripheral blood and bone marrow samples were obtained from patients who underwent total hip arthroplasty. By using two different protocols T cells were expanded in the presence of IL-2 and IL-7 either from bulk culture with exposure of two different peptide pools (IE1 and pp65) or after selection via IFN-γ secretion by stimulation with pp65. CMV specific immune responses were assessed by using multiparameter flow cytometry staining cells for CD3, CD4, CD8, CCR7 and CD45RA and for the cytokines IFN-γ IL-2 and TNF at day 0 and after 10 days of in vitro expansion.Results Similar frequencies of cytokine-producing pp65– and IE1-specific CD4+ and CD8+ T cells were found in unmanipulated paired PB and BM samples. Expansion of CMV-specific T cells from BM resulted in significantly higher frequencies of specific CD4+ T cells than from PB, whereas no difference in frequencies of CMV-specific CD8+ T cells was observed. Interestingly, significantly higher frequencies of BM pp65 and IE1-specific CD4+ T cells were multifunctional, characterized by producing simultaneously IFN-γ, TNF and IL-2 (IE1: BM mean 0.44% ± 0.16; PB mean 0.09% ± 0.05, p=0.031; pp65: BM mean 3.87% ± 2.46; PB mean 1.24% ± 0.90, p=0.031). Expansion of multi-functional CD4+ T cells from BM was observed with both the bulk and selection assay protocol. Both PB and BM CMV-specific CD4+ and CD8+ T cell lines had a predominant CD45RA-CCR7- effector memory phenotype.Conclusions This study implicates the use of human bone marrow as a source for expansion of multifunctional CMV-specific CD4+ T cells. Recent studies in HIV and Leishmania support the crucial role of multifunctional T cells in disease control (Darrah 2007 [2]).
Iron Overload Adversely Affects Survival and Increases Risk of Graft-Versus-Host Disease and Blood Stream Infections after Allogeneic Hematopoietic Stem Cell Transplantation.
Authors: Pullarkat Vinod , Blanchard Suzette , Tegtmeier Bernard , Dagis Andrew , Patane Kathryn , Ito James , Forman Stephen J. 
Published: October 2019

Abstract
Background: Iron overload, mainly from multiple red cell transfusions is common in patients undergoing allogeneic hematopoietic stem cell transplantation (HSCT) for hematologic disorders. Although the adverse impact of iron overload on survival of patients undergoing HSCT for thalassemia are well established, the impact of iron overload on the transplant outcome and complications in non-thalassemic patients is less clear.
Methods: Serum ferritin level, a marker of tissue iron overload was measured immediately prior to initiation of transplant conditioning in adult patients undergoing myeloablative HSCT from matched sibling or unrelated donors. The effect of elevated pretransplant ferritin (defined as ferritin over 1000ng/ml) on day 100 mortality, acute graft-versus-host disease (GVHD), infectious complications were assessed.
Results: Data on 190 patients were analyzed for day 100 mortality, overall survival, the combined endpoint of incidence of acute GVHD (Grade II or above) or death within 100 days and the combined endpoint of blood stream infection or death within 100 days. In univariate analysis, the high ferritin group had increased day 100 mortality (20% vs 9%, P=0.04), decreased overall survival (log-rank test: P-value=0.004), increased acute GVHD/death (63% vs 43%, P=0.009) and increased incidence of blood stream infections/death (60% vs 44%, P=0.04). In a multivariate analysis, high ferritin was associated with increased risk of death (Cox Model: hazard ratio 2.28, P-value=0.004), increased day 100 mortality (generalized linear models (GLM) odds ratio 3.82, P-value=0.013), increased incidence of acute GVHD/death (GLM odds ratio 3.11, P-value=0.001) and increased risk of blood stream infections/death (GLM odds ratio 1.99, P-value=0.032). The results remained similar when serum ferritin was considered a continuous variable.
Conclusions: Elevated serum ferritin adversely impacts day 100 mortality and overall survival after allogeneic HSCT. In addition, high serum ferritin also increases the likelihood of acute GVHD and blood stream infections after allogeneic HSCT.
The Mask Mutation Identifies TMPRSS6 as an Essential Suppressor of Hepcidin Gene Expression, Required for Normal Uptake of Dietary Iron.
Authors: Beutler Ernest , Lee Pauline , Gelbart Terri , Du Xin , Beutler Bruce 
Published: October 2019

Abstract
Hepcidin, the central negative regulator of iron absorption and iron release from macrophages, is upregulated by iron. Mutations in hemojuvelin, Hfe, transferrin receptor 2, and SMAD4 are known to prevent upregulation. Additionally, the bone morphogenetic proteins (BMPs), and the inflammatory cytokines IL-1 and IL-6 stimulate hepcidin gene activation. Downregulation of hepcidin is effected by anemia and hypoxia, but nothing is known of the mechanism through which this occurs. Here we describe the recessive ENU-induced phenotype Mask, so called because affected homozygotes developed regional alopecia in which truncal hair was shed while facial hair was retained. The Mask phenotype was found to be a manifestation of iron deficiency, and was eliminated by correcting the iron deficiency. When fed an iron deficient diet, mutant mice absorbed less iron than controls, as measured by total body 59Fe counting. After reaching a plateau total body counts stabilized, indicating that blood loss did not play a role in the iron deficiency. The level of liver hepcidin mRNA of iron deficient mice is normally greatly decreased; in contrast, the Mask mouse had high liver hepcidin mRNA levels. By positional cloning, we were able to ascribe the Mask phenotype to a splicing error in the Tmprss6 gene, which encodes a membrane-bound serine protease of previously unknown function. The mutation truncates the protein, eliminating the serine protease domain. Transfecting HepG2 cells to express the wildtype TMPRSS6 protein decreased baseline hepcidin reporter activity and almost entirely blunted the hepcidin inducing effect of IL-6, IL-1, hemojuvelin, and the BMPs. A construct encoding the Mask truncation mutant had diminished activity. Thus, TMPRSS6 powerfully down-regulates hepcidin gene transcription in the baseline state and prevents its upregulation by all known stimulators. TMPRSS6 is a non-redundant component of a hepcidin suppression pathway that exerts dominant effect over all known hepcidin inducing pathways, and is required for normal absorption of dietary iron.
Hematopoietic Stem Cell Transplantation (HSCT) in Waldenström Macroglobulinemia (Wm), Update of the French Experience in 54 Cases.
Authors: Dhedin N. , Tabrizi R. , Bulabois P.E. , Le Gouill S. , Coiteux V. , Dartigeas C. , Robin M. , Huynh A. , Larosa F. , Cacheux V. , Garnier A. , Morel P. , Kuentz M. , Dreyfus B. , Desablens B. , Guillerm G. , Pignon B. , Rio B. , Bilger K. , Cabrespine A. , Uzunov M. , Bay Jacques-Olivier , Leblond V. , Tournihac O. 
Published: October 2019

Abstract
Introduction: HSCT has been developed in few Wm cases and is nowadays challenged by other innovative approaches. However, high dose therapy followed by autologous HSCT (HD-auto) produces high response rate and some long term responses while allogeneic HSCT performed after either myeloablative (MA-allo) or reduced intensity conditioning (RIC-allo) regimens may be cure of Wm (Dreger 1998, Tournilhac 2003, Maloney 2006).
Methods: We updated and extended our retrospective experience on 32 HD-auto, 11 MA-allo and 11-RIC-allo performed from 1990 to 2006 in 51 patients from 18 institutions. A MA-allo and a RIC-allo were performed in 1 and 2 cases respectively following relapse after a 1st HD-auto.
Results: Data are presented in the table.
HD-auto MA-allo RIC-allo Nb 32 11 11 Median age at transplant 56 46 56 Interval: diagnosis-transplant 38 50 74 Chemoresistance at transplant 25% 36% 55% Conditioning regimen BEAM (13), TBI/melphalan (9), TBI/endoxan (7), other (3) TBI/endoxan (9), other (2) TBI/fluda (10), other (1) Donor Sibling (9), unrelated (2) Sibling (8), unrelated (2), cord blood (1) Median follow up (m) 45 (3–121) 68 (3–132) 22(2–60) Relapse 18 (56%) 4 (36%) 0 Transplant related mortality 12,5% (one 2th cancer) 36% 27% (one 2th cancer) overall survival (1;3;5y) 87%, 77%, 58% 64%, 54%, 54% 82%, 68%, 68% Event free survival (5y) 25% 48% 68%
Acute GVHD developed following 9 MA-allo [Grade III-IV (n=1)] and 8 RIC-allo [Grade III-IV (n=1)]. Chronic GVHD developed following 7 MA-allo [limited (n=5), extensive (n=2)] and 5 RIC-allo [limited (n=2), extensive (n=3)].
Conclusion: We confirm that autologous HSCT achieves some long term responses even in heavily pretreated patients. Allogeneic HSCT induces very long term disease control and may cure WM. Specially, the RIC-allo gives impressive results on disease control in a set of older patients, with refractory disease, mostly heavily pretreated.