bluuebunny/crossref_metadata_2025_split huggingface dataset:
ShRNA Targeting p190BCR-ABL Successfully Eliminates Ph-ALL Cells with or without ABL Kinase Domain Mutation.
Authors: Futami Muneyoshi , Hatano Toshiyuki , Soda Yasushi , Kobayashi Seiichiro , Miyagishi Makoto , Tojo Arinobu 
Published: October 2019

Abstract
In the majority of Philadelphia chromosome (Ph)-positive acute lymphoblastic leukemia (Ph-ALL) cases, the resulting BCR-ABL gene generates 190 kD active tyrosine kinase (p190) which is responsible for leukemogenesis and can be a molecular target for therapy. Although a series of ABL kinase inhibitors including imatinib, nilotinib and dasatinib reveal potent activities against Ph-ALL, acquired resistance caused by point mutations in the kinase domain such as T315I still remains to be overcome. That is why a novel strategy is desired in the treatment of Ph-ALL. We previously reported that lentiviral delivery of maxizyme targeting p190 specifically induced apoptosis of Ph-ALL cells (Blood 104:356, 2004). Since RNA interference proved to be a more powerful tool in selective gene silencing, we applied this technology to test whether specific and efficient killing of Ph-ALL cells could be achieved by down-regulation of p190. We designed a series of 21-mer and 27-mer small hairpin RNA (shRNA) targeting p190 mRNA and constructed plasmid vectors expressing these shRNA, which were screened by transfection of 293T/p190 cells to determine optimal target sites. As a result, three candidate sequences were identified; junctional 27-mer, junctional 21-mer and ABL 21-mer. Then, we inserted each of the shRNA expression cassettes into the lentiviral vector (HIV-U6/shRNA) and prepared high titer virus stock for infection of leukemia cells. shBCR-ABL/21, but not shBCR-ABL/27, induced significant and specific cell death of p190+ Ph-ALL cells in a time-dependent manner. shABL was more potent than shBCR-ABL/21 and also active against p210+ CML cells as well as 293 cells, but did not substantially affect Ph-negative leukemia cells. Both shABL and shBCR-ABL/21 completely inhibited growth of Ba/F3 cells harboring either wild-type or mutant p190 which renders those resistant to imatinib. Furthermore, both shRNA at low multiplicity of infection additively cooperated with imatinib in growth inhibition of Ba/F3-p190 cells. These data suggest that shRNA targeting p190 may become a therapeutic option in Ph-ALL by improvement of its delivery system like liposome.
Growth of BA/F3-p190BCR-ABL Cells transduced with shRNA Targeting p190BCR-ABL Growth of BA/F3-p190BCR-ABL Cells transduced with shRNA Targeting p190BCR-ABL
Inhibition of p38 MAPK Signalling Impairs the Homing of B Lineage ALL Cells to the Bone Marrow of NOD/SCID Mice.
Authors: Juarez Julius , Pena Aileen Dela , Baraz Rana , Bradstock Kenneth Francis , Bendall Linda J. 
Published: October 2019

Abstract
The mechanisms regulating the migration of B lineage acute lymphoblastic leukemia (ALL) cells between the blood and bone marrow compartments remain obscure. The chemokine SDF-1 regulates the motility and proliferation of ALL cells in vitro. We have previously demonstrated that the chemotaxic response of these cells to SDF-1 requires signalling through p38 MAPK and that signalling through PI-3K and MEK/ERK pathways plays only a minor role in a subset of cases. This contrasts with normal CD34+ hematopoietic progenitors and CD34+CD19+ B cell progenitors where signalling through PI-3K is the predominant pathway required for chemotaxis. In this study we examined the requirement for SDF-1 and p38 MAPK signalling in the homing of the B lineage ALL cells to the bone marrow of NOD/SCID mice. CFSE-labelled ALL cells were pre-treated with either the specific CXCR4 antagonists AMD3100 (100μM) or TC14012 (100μM), the p38 MAPK inhibitor SB203580 (10μM), the AKT inhibitor LY294002 (6.65μM) or the MEK inhibitor PD98059 (20μM) for 0.5–1 h at 37°C. Control or inhibitor treated cells were then injected intravenously into irradiated NOD/SCID mice which were either pre-treated with control (vehicle) or the inhibitors AMD3100 (200μg/mouse), TC14012 (200μg/mouse), SB203580 (0.1 mg/mouse), the AKT inhibitor LY294002 (2 mg/mouse) or the ERK inhibitor PD98059 (0.2 mg/mouse). Homing was assessed 4 hours after the injection of cells before significant proliferation of homed cells could occur. AMD3100 and TC14012 significantly inhibited the homing of ALL cells to the bone marrow by up to 80% and 67% respectively. Inhibition of p38 MAPK also decreased the homing of ALL cells to the bone marrow of NOD/SCID mice by 40% compared with the DMSO controls (p<0.05). No inhibition of homing was observed with the inhibitors of PI-3K or MEK in ALL cells. These results support our previous in vitro findings with regards to the importance of the p38 MAPK pathway in SDF-1 induced motility of ALL cells and demonstrate that the same pathway plays an essential role in the homing of these cells to the bone marrow of NOD/SCID mice.
Pediatric T-ALL but Not cALL Blasts Express Ligands for Siglec-7 Inhibiting NK Cell Mediated Lysis.
Authors: Viebahn Susanne , Gieseke Friederike , Pfeiffer Matthias , Handgretinger Rupert , Müller Ingo 
Published: October 2019

Abstract
Changes in the glycosylation pattern of cell surface proteins occur during early stages of malignant transformation. These alterations in the glycan repertoire may contribute to numerous processes of tumor cell survival such as adhesion, migration and immune evasion. Sialic acid is the most common terminal carbohydrate moiety of glycan structures in humans binding to a family of eleven receptors termed sialic acid binding Ig-like lectins (siglecs). Siglecs are expressed on virtually all effector cells of the immune system and several members function as inhibitory receptors. Siglec-7 is known as an inhibitory receptor expressed by NK cells and T cells. Therefore, siglec-7 ligands may contribute to immune evasion of malignant cells.
In order to analyze the expression of siglec-7 ligands by malignant cells, we cloned the extracellular domain of human siglec-7. The recombinant protein was expressed in 293 cells to allow glycosylation, which is supposed to be important for specificity. As the interaction of a single glycan and lectin is of low affinity, we tetramerized siglec-7 in analogy to MHC-tetramers to increase avidity. This strategy allowed for reliable use of siglec-7 fusion protein in flow cytometry. Analysis of PBMC from healthy donors revealed that siglec-7 ligands are expressed on all subpopulations of PBMC: expression was detected on CD4+ and CD8+ T-cells as well as on monocytes and NK cells. B cells could be subdivided into a positive and negative population after staining with siglec-7 tetramers.
Subsequently, we analyzed primary lymphatic blasts from childhood leukemias. Interestingly, T-ALL blasts expressed ligands for siglec-7, whereas these ligands were absent from cALL blasts. As cALL blasts are known to be better targets for NK cell-mediated cytotoxicity than T-ALL blasts, we hypothesized that siglec-7 ligands on the surface of leukemic blasts inhibit NK cells. To test the functional relevance of siglec-7 ligand expression for NK cell-mediated cytotoxicity, we used soluble monomeric siglec-7 as competitive inhibitor in cytotoxicity assays. In these competition assays, NK cell-mediated specific lysis of leukemic cells increased roughly twofold in the presence of soluble siglec-7.
These results show that engagement of inhibitory siglecs on the surface of effector cells might contribute to immune escape mechanisms of malignant cells. It underlines the function of siglec-7 as a non-MHC-specific inhibitory receptor on NK cells and the important role of altered glycans expression on malignant cells in tumor immunology.
Factors Influencing Survival after Relapse from Childhood ALL: A Children’s Oncology Group Study.
Authors: Nguyen Kim , Cheng Su-Chun , Raetz Elizabeth , Sather Harland , Carroll William L. , Winick Naomi , Hunger Stephen P. , Gaynon Paul , Loh Mignon L. 
Published: October 2019

Abstract
Despite improvements in front-line therapy for acute lymphoblastic leukemia (ALL), treatment after relapse remains a critical challenge. Examination of outcome after relapse in a large, recently treated, and unselected population provides a useful benchmark for evaluating new approaches.
Survival after relapse was tracked on Children’s Cancer Group (CCG) ALL trials. Relapses were categorized by site (isolated or combined bone marrow relapse (MR); isolated central nervous system relapse (CNSR); isolated testicular relapse (TR)), and time from diagnosis to relapse (early < 18 months; intermediate between 18 and 36 months; late > 36 months). We examined the impact of age, sex, ethnicity, initial risk assignment (standard risk (SR) or higher risk (HR)), immunophenotype, length of first remission (CR1), and site of relapse on outcome in multivariate analyses.
A total of 9,585 infants, children, and adolescents with newly diagnosed acute lymphoblastic leukemia were enrolled on CCG trials conducted between 1988 and 2002. We examined survival after relapse for 1,874 children with MR, CNSR, or TR.
Survival after relapse by site and time to relapse Site MR CNSR TR Time to Relapse n *survival(%) n *survival(%) n *survival(%) *3-year survival Early 498 10 175 49 15 50 Intermediate 378 23 179 73 35 57 Late 488 53 52 77 54 70
Overall survival for any bone marrow relapse was best for late MR and worst for early MR. Early MR was linked to HR presenting features and T-cell disease. Survival after intermediate or late MR was superior for SR patients (p <.05). Survival was better for CNSR or TR than for MR. Older age at diagnosis, male sex, T cell immunopheno-type, and CNS disease at presentation were significant adverse factors for survival after adjustment for site and time to relapse in multivariate analyses (p<.05).
Importantly, in the same multivariate analysis, patients treated on more contemporary trials (CCG1952CCG1953CCG1961, and 1962) had inferior survival rates after relapse when compared to patients treated on earlier trials (CCG 1881CCG 1882CCG 1883CCG 1891CCG 1901, and 1922) (p<.001). Importantly, these data from the analysis of a large, unselected group of patients indicates that features at initial presentation and prior therapy influences outcome following relapse. The inferior survival rates of those relapsing on more intensive protocols likely reflects particularly aggressive and chemoresistant disease leading to poor salvage. Increasing the intensity of retrieval therapy is unlikely to improve after relapse and novel therapeutic approaches that target these resistant leukemias are urgently needed for successful treatment of relapsed ALL.
The Role of Cranial Irradiation (CRT) or Extended Triple Intrathecal Therapy (T.I.T.) in Childhood T-Cell ALL; the ALL Israel National Studies (INS) 89 and 98 (Modified ALL-BFM 86 and 95) Report.
Authors: Stark Batia , Attias Dina , Avrahami Gali , Ballin Ami , Bielorai Bela , Burstein Yoav , Elhasid Ronit , Gavriel Herzel , Kapelushnik Josef , Miskin Hagit , Sthoeger Dalia , Weintraub Michael , Steinberg David , Izraeli Shai 
Published: October 2019

Abstract
Patients with T-cell ALL are at increased risk of CNS relapse and require more intensive CNS directed therapy. CRT is considered to be the most effective therapy, but is associated with serious adverse late effects and secondary brain tumors. We evaluated retrospectively, in the context of the ALL-BFM-based protocols, the impact of CRT or extended T.I.T. on outcome within High Risk (HR: Prednisone poor responders) and non-HR (Prednisone good responders) subgroups.
In INS 89, modified ALL-BFM 86/90, all Non-HR patients received extended T.I.T (×18) as CNS preventive treatment, and VP-16 was added systemically in between the HD-MTX (5g/m2). In INS 98, based on ALL-BFM 95, VP-16 was omitted and pulses of VCR+DEXA were added in maintenance. Patients with T-ALL Non-HR and WBC >100000, were assigned to receive CRT 12 Gy, following the results of the AIEOP-ALL 91 (Conter J.C.O. 15:2786–2790, 1997) reporting inferior outcome in this group when comparing T.I.T only to BFM with irradiation. T-ALL HR patients were treated without modification and were all assigned to CRT 18 or 12Gy.
Results: Between 1989 and 2002, 145 T-ALL children aged <18 years were enrolled in the INS 89 (84 patients, 1989–97) and INS 98 (61 patients, 1998–2002). With a median follow-up of 12 and 5 years in INS 89 and INS 98 respectively, the 5 year event-free-survival (EFS) was 61.9±5.3% and 75.4±5.5%, respectively (p=0.14), and CNS relapse (isolated and combined) rates were 4.1% and 1.8%, respectively (p=0.4). Overall 5 yrs EFS, DFS, and CNS relapse rate of all the T-ALL patients were 67.4%, 78.8%, and 3.1% respectively. Outcome of Non-HR patients with WBC <100000 (all were non irradiated 72 pts) was; EFS 74.7%, and no CNS relapse. In the Non-HR with WBC >100000 (without CNS involvement) 9 patients were irradiated and 18 patients received the Extended T.I.T. These groups had 5 yr EFS of 78%±14% and 76%±10% respectively, and CNS relapse rate 11.1% and 5.6% respectively (p=0.59). In the HR group (34 pts) the 8 patients who refused CRT fared significantly worse than the irradiated patients, with EFS of 38%±17% Vs 73%±11% (p=0.08), and CNS relapse rate of 17% Vs 0 (p=0.14). T-cell ALL Non-HR children (prednisone good responders), in the context of the ALL-BFM intensive protocols and extended T.I.T may not require preventive CRT regardless of their WBC at presentation.
ABCA3 Expression in AML Blast Cells: Association with Poor Disease Outcome and Intracellular Drug Sequestration.
Authors: Chapuy Bjoern , Radunski Ulf , Corsham Sabrina , Koch Rafael , Cheong Naeun , Inagaki Nobuya , Griesinger Frank , Reinhardt Dirk , Truemper Lorenz , Wulf Gerald G. 
Published: October 2019

Abstract
Analysing the intrinsic drug efflux capacity of leukemic stem cell with the side population (SP) phenotype, we recently found the intracellular ABC transporter ABCA3 expressed in blast cells from patients with acute myeloid leukaemia (AML). Here we extend our observations to a larger cohort of AML samples and addressed the role of ABCA3 for cytostatic drug resistance. Applying quantitative RT/PCR (Taqman), we detected significant levels of ABCA3 in 59 of 92 blast cell samples from 58 pediatric and 34 adult patients with AML. The expression of ABCA3 correlated positively (coefficient 0.83) with the expression of CD34, while there was no association with the acquisition of markers for myeloid and monocytic maturation. As for disease outcome analysed by univariate analysis, we found a significantly worse outcome in both event free survival (p=0.048) and overall survival (p=0.034) for the patients with high-level ABCA3 expression. Functionally, the expression of ABCA3 was restricted to the cytoplasm in the AML blast cells, and co-localisation studies revealed an association with endosomal markers. Ectopic expression of ABCA3 augmented the lysosomal compartment in HEK 293 cells, and enhanced intracellular anthracyclin sequestration into vesicles of the endosomal system. Correspondingly, we observed a significant reduction of cytostatic efficacy from anthracyclins in stably ABCA3 expressing HEK293 cells compared to the parental cell line. In conclusion, we found the intracellular transporter ABCA3 expressed in the blast cells of most patients with AML, and elevated ABCA3 levels associated with poor disease outcome. Functional data revealed intracellular anthracyclin sequestration by ABCA3, suggesting a role of ABCA3 for drug resistance in patients with AML.
Differential Expression of SLAM Family Receptor Markers in Normal Human Hematopoietic Stem Cells and Their Malignant Counterpart in MDS and AML.
Authors: Tiu Ramon V. , Powers Jennifer J. , Haddad Abdo , Jiang Ying , Maciejewski Jaroslaw P. 
Published: October 2019

Abstract
Members of the signaling lymphocytic activation molecule (SLAM) family, including CD150, CD48 and CD244, were shown to precisely distinguish more committed lineage restricted progenitor cells from pluripotent and multipotent murine hematopoietic stem cells (HSC; Kiel et al; 2005 Cell). Similar SLAM profiles may also be present on HSC subsets in humans. We hypothesized that these SLAM markers may be indicators not only of stem cell potential in normal hematopoiesis but also distinguish a subset of the most immature malignant precursors of leukemia. In agreement with the concept of a “cancer stem cell,” the presence of leukemic stem cell population may be an indicator of important clinical and biological properties. We first tested the distribution of CD150, CD48 and CD244 antigens on human CD34+ cells derived from 7 control individuals using 4-color flow cytometry. CD34+ cells were measured in the blast gate based on side scatter and CD45 expression. Within CD34+ blasts, expression of CD48, CD150, and CD244 was detected on 16.71%±9.69, 6.53%±2.93, and 26.92%±6.95 of cells respectively. Subsequently, we investigated SLAM expression in 9 immature leukemic cell lines, including KG-1, K562, U937, HEL, HL60, MKN-95, NB-4, Kasumi and UT7, and found increased expression of SLAM markers in KG-1 (CD48+, CD150+, CD244+) and Kasumi (CD48−, CD150−, CD244+). Consequently, none of the leukemic cells showed pluripotent/multipotent SLAM profiles. We then compared the SLAM marker expression on blasts from patients with AML and MDS with that of CD34+ cells from normal controls. We studied a total of 28 patients: 11 MDS (2 low grade, 5 advanced MDS, 3 MDS/MPD overlap) and 10 AML (FAB: 3 M1, 2 M2, 1 M3, 2 M4/M4E0 and 2 M6). In our cohort, 8/10 AML patients expressed one of the three SLAM markers; 6/10 were CD150−CD48−CD244+ (63.57%±6.96) and 2/10 were CD150+CD48−CD244−(46%±10.96) suggestive of the presence of either pluripotent or multipotent leukemic stem cell phenotype. In the MDS cohort, 8/11 patients expressed one of three SLAM markers, 7/11 were CD150−CD48−CD244+ (41.21% ± 8.9) and 1/11 were CD150+CD48−CD244− (1.26%±0.59) again consistent with a profile derived from either pluripotent or multipotent stem cells. None of the MDS and AML patients had either co-expression of CD244 and CD48 or increased expression of CD48 alone. Two of the M1 type AML patients with CD150−CD48−CD244+ phenotype received prior chemotherapy and achieved complete remission on bone marrow biopsy and flow cytometry using traditional blast markers. In some, we conclude that the SLAM receptor markers may be associated with certain types of leukemic blasts and may be useful in the identification of leukemic stem cell population in both MDS and AML.
TAT-Apoptin Mediated Induction of Apoptosis in Leukaemic Cells.
Authors: Gaken Joop , Pericleous Louisa , Farzaneh Farzin , Mufti Ghulam J. , Tavassoli Mahvash 
Published: October 2019

Abstract
We have studied the specific targeting of leukaemic cells using the Chicken Anaemia Virus (CAV)-derived protein VP3 (Apoptin) linked to the protein transduction domain (PTD) from HIV TAT with the aim of using this strategy for in vitro purging. Apoptin is a 13.6 kDa protein which induces apoptosis specifically in cancer cells whilst leaving normal cells unaffected. Expression of Apoptin in normal cells results in its cytoplasmic localisation. In tumour cells Apoptin resides initially in the cytoplasm and subsequently translocates to the nucleus and induces apoptosis. Apoptin is phosphorylated both in vitro and in vivo in tumour cells but negligibly in normal cells at threonine 108. A gain-of-function point mutation (T108E) results in accumulation of Apoptin in the nucleus and the killing of normal cells, implying that phosphorylation is a key factor of the tumour-specific properties of Apoptin.
We have demonstrated that Apoptin induces apoptosis in a variety of human solid tumour cell lines, but not in normal fibroblast and epithelial cells. Apoptin induced apoptosis in HSC3 head and neck cancer cells acts through the mitochondrial pathway and was blocked (>75%) by shRNA against PUMA, a BH3 Only protein which induces Bax and BAK resulting in loss of mitochondrial membrane potential and release of cytochrome C. Furthermore, activation of the p53 family member, p73, substantially increased (5–10 fold for p73 β and γ) sensitivity of Saos2 tumour cells to Apoptin-induced killing.
For efficient protein delivery, Apoptin was fused to a TAT PTD and addition of this protein to normal and tumour cells resulted in the selective killing of tumour cells. To increase the stability and solubility of TAT-Apoptin we have fused it to the maltose binding protein (MBP), this modification significantly increases both yield and the solubility of Apoptin while retaining its biological function. Apoptin tumour specific toxicity was assessed in a range of leukaemic and solid tumour cell lines. Addition of MBP-TAT-Apoptin protein to HL60, K562 and Jurkat cells resulted in 50%, 55% and 75% cell death by apoptosis as judged by PARP cleavage, respectively, at day 4 as compared to MBP-TAT control whilst normal B cells, fibroblasts and epithelial cells are unaffected. Fluorescent microscopy demonstrated that MBP-TAT-Apoptin was rapidly internalised in almost 100% of cells within 24hrs in all cell types tested. Direct injection of Apoptin expressing Ad vectors also showed clear regression of established tumours in mice.
The cancer specific toxicity of Apoptin has potential value for a range of therapeutic applications such as purging of autologous bone marrow as used for the treatment of multiple myeloma and possibly direct treatment of leukaemias either alone or linked to antibodies for targeting of specific types of leukaemias.
Heme Oxygenase-1 (HO-1)/Heat Shock Protein 32 (Hsp32) as a Novel Survival Factor and Target in AML.
Authors: Kneidinger Michael , Gleixner Karoline V. , Kondo Rudin , Samorapoompichit Puchit , Vales Anja , Krauth Maria T. , Mayerhofer Matthias , Gruze Alexander , Pickl Winfried F. , Greish Khaled , Boehm Alexandra , Sperr Wolfgang R. , Maeda Hiroshi , Valent Peter 
Published: October 2019

Abstract
Heme oxygenase 1 (HO-1), also known as heat shock protein 32 (Hsp32), has recently been identified as a stress-related survival molecule that acts anti-apoptotic and cytoprotective in inflammatory reactions. Recent data suggest that HO-1/Hsp32 is also expressed in neoplastic cells in various malignancies. In the present study, we provide evidence that HO-1 is constitutively expressed in primary leukemic cells in patients with acute myeloid leukemia (AML, n=17) and in various AML cell lines such as HL60, KG1, KG1a, and U937. Expression of HO-1 mRNA was demonstrable by RT-PCR, and the HO-1 protein by immunocytochemistry and Western blotting. In addition, we were able to demonstrate expression of HO-1 mRNA and of HO-1 protein in the CD34+/CD38− progenitor/stem cell fraction in the leukemic clone in patients with AML. The HO-1 inductor hemin (10 μM) was found to promote expression of HO-1 in AML cells. Incubation with the HO-1-targeting drugs pegylated zink protoporphyrin (PEG-ZnPP) or styrene maleic acid-conjugated ZnPP (SMA-ZnPP), resulted in a dose-dependent inhibition of growth of leukemic cells at pharmacologic concentrations (IC50: 5–20 μM for cell lines and primary AML cells). The SMA-ZnPP-induced growth-inhibition of AML cells were found to be associated with induction of apoptosis as evidenced by light microscopy, electron microscopy, and by a Tunel assay. In consecutive experiments, combination experiments were performed using SMA-ZnPP and AML cell lines. In these experiments, SMA-ZnPP was found to synergize with cytarabine in producing growth inhibition in all AML cell lines tested. In summary, these data show that HO-1/Hsp32 is a novel survival factor and interesting target in AML. The clinical significance of this observation remains to be determined in forthcoming trials.