bluuebunny/crossref_metadata_2025_split huggingface dataset:
Ex Vivo Gene Trasfer for Hemophilia A That Enhances Safety and Results in Sustained In Vivo Factor VIII Expression from Lentivirally-Engineered Endothelial Progenitors.
Authors: Matsui Hideto , Shibata Masaru , Brown Brian , Labelle Andrea , Hegadorn Carol , Andrews Chandler , Hebbel Robert P. , Galipeau Jacques , Hough Christine , Lillicrap David 
Published: October 2019

Abstract
Hemophilia is an excellent candidate disorder for the use of gene therapy as a treatment modality. However, significant obstacles have been encountered with systemic delivery of viral vectors that have prevented sustained expression of the therapeutic protein. Investigation of alternative gene therapy strategies for hemophilia that enhance safety and facilitate long-term, therapeutic levels of the transgene product is imperative. In this study, we evaluated an ex vivo gene therapy strategy for hemophilia A. Circulating endothelial cell progenitors (blood outgrowth endothelial cells - BOECs) were isolated from canine and mouse blood and transduced with a third generation self-inactivating lentiviral vector encoding the canine FVIII transgene under the transcriptional control of either the CMV promoter or an endothelial cell-specific regulatory element. Transduced BOECs were injected either intravenously (IV) or subcutaneously mixed with Matrigel (SC+Matrigel) into NOD/SCID mice. Canine FVIII antigen levels were assayed at weekly intervals using an Asserachrom VIII:Ag ELISA that detects canine FVIII against a background of normal murine FVIII levels in the NOD/SCID mice. The mean FVIII antigen levels in mice injected with BOECs at 3 weeks following treatment were 37.5 mU/mL and 105.8mU/mL, for IV and SC+Matrigel administration, respectively. These FVIII antigen levels were sustained up to 12 weeks at therapeutic levels (21.3mU/mL and 21.7mU/mL, for IV and SC+Matrigel administration respectively). To evaluate if the observed loss of FVIII expression by 12 weeks post-treatment resulted from transcriptional silencing of the viral promoter, the CMV promoter was replaced with the endothelial cell-specific thrombomodulin (TM) promoter and transduced BOECs were implanted SC with Matrigel. In contrast to results from the CMV-regulated transgene, sustained therapeutic levels of FVIII have been documented for the duration of the study with the TM-regulated construct (34.3 mU/mL at 3 weeks and 22.5 mU/mL at 20 weeks) Immunostaining at 18 weeks after SC implantation of the transduced BOECs, shows that these cells still express FVIII and von Willebrand Factor. Biodistribution analysis by flow cytometry and quantitative PCR demonstrated that SC-implanted BOECs were retained inside the scaffold and were not detected at any other anatomic site. These results indicate that genetically-modified endothelial progenitors implanted in a SC scaffold can provide sustained therapeutic levels of FVIII and are a promising safe delivery vehicle for gene therapy of hemophilia. Currently, these engineered cells have been implanted into immunocompetant mice and FVIII levels are being assessed.
Chimeric Immunoreceptor (T-Body) Targeting of Acute B-Cell Lymphoblastic Leukemia (B-ALL) through Lentivirus Engineering of Primary Human T Cells.
Authors: Milone Michael C. , Fish Jonathan , Carpenito Carmine , Lakal Mehdi , Ofori Ella , Danet-Desnoyers Gwenn , Riley James L. , Grupp Stephan A. , June Carl H. 
Published: October 2019

Abstract
T-bodies that combine the antigen recognition capability of single chain antibody molecules (scFv) with the signal transduction domains of activating receptors can be used to generate T cells with novel, engineered specificities independent of their endogenous TCR. We have developed a series of T-bodies with specificity towards CD19, a molecule with expression that appears restricted to normal and malignant B cells. These T-bodies contain the activating signals from the costimulatory T cell receptors, CD28 and/or 4-1BB, in tandem with the TCR-ζ signal transduction domain. Using lentiviral gene transfer and T cells expansion via bead-immobilized anti-CD3 and anti-CD28 antibodies, we can generate T cells 95% of which express the T-bodies. Lentiviral transduction combined with this efficient T cell expansion system has the potential to produce >1011 antigen-specific T cells without the need for a cell selection step. Interestingly, we have noted significant constitutive signaling effects of T-body overexpression on transduced T cells that may been overlooked previously due to lower transduction efficiency. Despite the observed constitutive effects, we show that lentiviral-engineered T cells expressing anti-CD19 T bodies efficiently kill CD19 expressing cell lines and primary B-ALL cells in an antigen-specific manner. T-body engineered T cells also produce cytokines in response to antigen triggering with different patterns of cytokine production by T cells engineered with different T-bodies. In particular, a T-body receptor containing the 4-1BB signal transduction domain combined with the TCR-ζ domain stimulates a >6-fold increase in IL-6 production compared with other receptor combinations in CD4+ T cells. In contrast, a CD28 and TCR-ζ domain containing T body uniquely enhances IL-2 (>3-fold) and TNF-α (> 7-fold) secretion by CD8+ T cells. These results indicate that the functional characteristics of our tested T-body signaling constructs are distinct and depend upon the particular receptor and T cell subset combination. We are also comparing the anti-tumor response generated by adoptively transferred T-body engineered T cells in leukemia bearing immunodeficient mice as a pre-clinical, therapeutic model. In addition to elucidating important functional differences of our lentiviral constructs, these studies indicate that an important variable in clinical trials will be optimizing the cytosolic signaling domain for the desired T cell subset. The present results suggest that distinct combinatorial signaling domains may be required for optimal function in CD4 and CD8 engineered T cells.
The Use of Nitrous Oxide To Treat Pain of Lumbar Punctures in Ambulatory Pediatric Leukemia and Lymphoma Patients.
Authors: Hargens Liesl , Slagerman Kathleen , Moertel Christopher , Moore Caron , Slomiany David , Messinger Yoav 
Published: October 2019

Abstract
Nitrous oxide is an anesthetic gas that has been used extensively to decrease pain during dental procedures. Recent publications report that nitrous oxide administration to pediatric patients is safe and effective in reducing pain and anxiety associated with a variety of medical procedures. We completed a randomized, crossover, pilot study in our outpatient clinic to compare the efficacy of nitrous oxide (N2O) to standard conscious sedation (SCS) in providing analgesia during lumbar puncture (LP) in children with leukemia and lymphoma.
Two nurses evaluated 12 subjects during a N2O LP and 10 subjects during a SCS LP (midazolam and either fentanyl or morphine). Inter-nurse reliability was not different for any measure. The mean time for clinic visit was statistically shorter for the N2O group, 34 minutes, compared to 57 minutes for SCS (p=0.010). The mean length of procedure was 9 minutes using N2O or SCS. The type of sedation did not affect the difficulty performing the LP. The incidence of side effects (nausea, vomiting, combative behavior, and unsteadiness when leaving for home) was similar in the N2O and SCS groups. Pain control was equivalent. The nurse, parent, and subject reported pain during the procedure as zero for 5/12 subjects with N2O and 3/10 subjects with SCS. Parents and subjects were very satisfied with N2O (parent=1.8, child=2.8) and SCS (parent=1.5, child=2.2) sedation (10-point Likert scale with 1=very satisfied, 10=very dissatisfied). After completion of both procedures, 8/12 subjects preferred N2O whereas 2/10 preferred SCS. All twelve subjects who received N2O returned to normal activity within an hour of leaving the clinic and 3/10 subjects who received SCS required longer than one hour.
Nitrous oxide is effective in reducing procedural pain with few side effects while decreasing time required for clinic visit and return to normal activity. Patient and parent satisfaction was high with more patients and families preferring N2O sedation to SCS.
Parent and Nurse Mean Pain Scores Before, During, and After Lumbar Puncture Using N2O and SCS (10-point Likert Scale) Pre-Procedure Pain During Procedure Pain Post-Procedure Pain Parent Nurse Parent Nurse Parent Nurse N2O 0.3 0 1.4 0.9 0.5 0.3 SCS 0.3 0.1 2.5 1.9 1.5 0.7
Cancer/Testis Genes in Multiple Myeloma: Expression Patterns and Vaccination Strategies Determined by Microarray Analysis.
Authors: Condomines Maud , Hose Dirk , Reme Thierry , Hundemer Michael , De Vos John , Baudard Marion , Moehler Thomas , Pantesco Veronique , Schved Jean-Francois , Rossi Jean-Francois , Goldschmidt Hartmut , Klein Bernard 
Published: October 2019

Abstract
Cancer-testis (CT) antigens are expressed in testis and malignant tumors, but rarely in non-gametogenic tissues. Due to this pattern, they represent attractive targets for cancer vaccination approaches. The aims of the present study were (1) to assess for the first time the expression of CT genes on a pangenomic basis in multiple myeloma (MM), (2) to provide selection strategies of CT antigens for clinical vaccination trials and (3) to assess the impact of CT gene expression on event-free survival. We report here the expression pattern of CT genes in purified MM cells (MMC) of 64 patients with newly-diagnosed MM, 12 patients with monoclonal gammopathy of unknown significance (MGUS), in normal plasma cell and B cell samples and in 20 MMC lines, using gene expression profiling (GEP). Out of 46 CT genes interrogated by the Affymetrix HG U133 Set arrays, 35 were expressed in MMC of at least one patient, according to the Affymetrix “present” call (frequency range: 2% – 66%). Of these, 24 CT genes were expressed in more than 5% of the MMC samples and 25 are located on chromosome X. MMC of 98% of the patients expressed at least one CT gene, 86% at least two, and 70% at least three CT genes. By using a set of 10 CT genes including KM-HN-1, MAGE-C1, MAGE-A3/6/12, MAGE-A5, MORC, DDX43, SPACA3, SSX-4, GAGE-1–8 and MAGE-C2, a combination of at least three CT genes - desirable to circumvent tumor escape mechanisms and immune tolerance - could be obtained in MMC of 67% of the patients. Thus, gene expression profiling can be used to select CT antigens as vaccination targets in individual patients. In a series of MMC from 111 patients treated with the same high-dose chemotherapy and autologous peripheral blood stem cell transplantation protocol and having a median two-year follow-up, we found that the expression of six CT genes, i.e. CTAG-1B, CTAG-2, MAGE-A1, MAGE-A2, MAGE-A3 and MAGE-A6 was associated with a shorter event-free survival (EFS). Furthermore, considering only the 25 CT genes encoded by chromosome X, a CT-Xhigh cluster comprising MMC of one third of the patients (35 of 111) could be defined using a binary hierarchical clustering based on Affymetrix call. Patients in the CT-Xhigh cluster had a shorter EFS (median 13 months) compared to patients in the CT-Xlow cluster (median 18 months, P = .003). The CT-Xhigh clsuter included more patients with a stage III disease (P = .004). These results confirm data from previous studies indicating that patients expressing some CT genes located on chromosome X have a poor outcome.
JNK Regulates DKK1 Expression in Multiple Myeloma Cells.
Authors: Colla Simona , Zhan Fenghuang , Yaccoby Shmuel , Epstein Joshua , Barlogie Bart , Shaughnessy John D. 
Published: October 2019

Abstract
Multiple myeloma (MM), a tumor of terminally differentiated plasma cells, depends on the bone marrow for growth and survival. MM is characterized by extensive bone loss and osteolytic lesions located at the sites of medullary plasmacytomas, suggesting that myeloma cells alter the biology of bone remodeling. Wnt signaling in osteoblasts is essential for the maintenance of osteoblast and osteoclast homeostasis and thus coupled bone turnover. MM plasma cells, but not those from healthy donors or MGUS, secrete Wnt signaling inhibitors DKK1, sFRP-2 and sFRP-3. The production of these factors contributes to osteolytic lesions by inhibiting Wnt signaling in the bone marrow milieu. It is currently not known how DKK1 expression is activated and regulated in MM. Here we report that DKK-1 expression is significantly lower in 45 myeloma cell lines (MMCL) and 5 secondary plasma cell leukemias/ pleural effusions compared to primary myeloma cells and down-regulated in all 9 primary myeloma cells with high DKK1 expression levels (Affymetrix Signal > 1,000) following co-culture with osteoclasts. This inhibition of DKK1 is associated with the simultaneous down-regulation of JUN, FOS, and FOSB, of which JUN has been shown to regulate DKK1 (Grotewald and Ruther, 2002). Ectopic expression of c-Jun in Jun negative MMCLs did not induce DKK1 expression, indicating that DKK1 transcription in this cell type is not solely dependent on c-Jun. DKK1 transcription increased 2- to 20-fold in myeloma cells from 21 of 42 primary MM following 48 hour of in-vivo exposure to thalidomide and 19 of 24 exposed to lenolidomide. The JNK-specific inhibitor SP600125 could significantly inhibit the up-regulation of DKK1 in 7 of 9 primary MM cells following in-vitro exposure to thalidomide or lenolidomide. Finally, treatment of MMCL and primary MM with the JNK activator Anisomycin resulted in up-regulation of DKK1 2- to 5-fold, but not in the presence of SP600125. These data indicate that DKK1 expression in MM cells is, in part, dependent on JNK signaling and suggests that JNK inhibitors may be an effective means of inhibiting both DKK1 expression and MM bone disease.
Upregulation of c-Jun Induces Cell Death Via Caspase-Triggered c-Abl Cleavage in Human Multiple Myeloma.
Authors: Podar Klaus , Raab Marc S. , Tonon Giovanni , Sattler Martin , Barila Daniela , Tai Yu-Tzu , Yasui Hiroshi , Raje Noopur , Depinho Ronald A. , Hideshima Teru , Chauhan Dharminder , Anderson Kenneth C. 
Published: October 2019

Abstract
The tyrophostin adaphostin (NSC680410) achieves remarkeable responses in patients with chronic myelocytic leukemia (CML), including Bcr/Abl-positive, Bcr/Abl-negative, and Bcr-Abl T315I mutant tumor cells resistant to both imatinib mesylate and second- generation BMS354825 and AMN107. In addition it also demonstrates cytotoxicity against chronic lymphcytic leukemia (CLL) and acute myelocytic leukemia (AML) cells. Several mechanisms have been proposed as a basis for its robust anti-tumor activity including generation and release of reactive oxygen species (ROS), cytochrome-c and apoptosis- inhibiting factor (AIF), caspase- cleavage, JNK activation, as well as inactivation of Raf-1, Stat3, and Stat5. As these adaphostin- targeted pathways are relevant to MM pathogenesis, we here sought to determine the potential molecular sequelae and therapeutic promise of adaphostin in MM. After demonstrating the anti- myeloma cytotoxicity of adaphostin, we carried out expression profiling of adaphostin- treated MM cells to identify its molecular targets. Surprisingly, c-Jun was the most upregulated gene even at the earliest point of analysis (2 hours). We also observed adaphostin- induced c-Abl cleavage in immunoblot analysis. Proteasome inhibitor bortezomib, but not melphalan or dexamethasone, induced similar effects, indicating unique agent- dependent mechanisms. Using caspase inhibitors as well as caspase- resistant mutants of c-Abl (TM-c-Abl and D565A-Abl), we then showed that c-Abl cleavage in MM cells requires caspase activity. Importantly, knockdown of c-Jun and c-Abl expression by siRNA confirms that adaphostin- induced c-Jun upregulation triggers downstream caspase- mediated c-Abl cleavage, inhibition of MM cell growth, and induction of apoptosis. Finally, our data suggest that this mechanism may not be restricted to MM, but may also be important in a broad range of malignancies including erythroleukemia and solid tumors.
PU.1 Is Downregulated in a Subset of Multiple Myeloma by the Methylation of Its -17 kb Upstream Regulatory Region and the Promoter.
Authors: Okuno Yutaka , Tatetsu Hiro , Ueno Shikiko , Hata Hiroyuki , Yamada Yasuhiro , Takeya Motohiro , Iino Tadafumi , Akashi Koichi , Tenen Daniel G. , Mitsuya Hiroaki 
Published: October 2019

Abstract
It has been reported that disruption of transcription factors critical for hematopoiesis, such as C/EBPa and AML1, is involved in leukemogenesis. PU.1 is a transcription factor important for both myeloid and lymphoid development. We reported that mice in which the levels of PU.1 were 20% of that of wild-type developed acute myeloid leukemia, T cell lymphoma, and a CLL-like disease. These findings strongly suggest that PU.1 has tumor suppressive activity in multiple hematopoietic lineages. Last year, we reported that PU.1 is downregulated in a majority of multiple myeloma cell lines and and freshly isolated CD138 positive myeloma cells from certain number of myeloma patients, and that tet-off inducible exogenous expression of PU.1 in PU.1 negative myeloma cell lines induced cell growth arrest and apoptosis. Based on their PU.1 expression levels, we divided the myeloma patients into two groups, namely PU.1 high and PU.1 low-to-negative, (cutoff index of 25th percentile of the PU.1 expression level distribution among all patients). The PU.1 low-to-negative patients had a significantly poorer prognosis than the PU.1 high patients. To elucidate the mechanisms of downregulation of PU.1, we performed sequence and epigenetic analysis of the promoter region and the -17 kb upstream region that is conserved among mammalians and important for proper expression of PU.1. There are no mutations in these regions of all five myeloma cell lines. In contrast, the -17 kb upstream region was highly methylated in 3 of 4 PU.1 negative myeloma cell lines, while the promoter region was also methylated to various levels in all five myeloma cell lines including one PU.1 positive cell line. These data suggested that the downregulation of PU.1 in myeloma cell lines might be dependent on the methylation of both regulatory regions of PU.1 gene, especially the -17 kb upstream region. We also evaluated the mechanisms of cell growth arrest and apoptosis of myeloma cell lines induced by PU.1. Among apoptosis-related genes, we identified that TRAIL was upregulated after PU.1 induction. To evaluate the effect of upregulation of TRAIL, we stably introduced siRNA for TRAIL into myeloma cell lines expressing PU.1, and we found that apoptosis of these cells was partially suppressed by siRNA for TRAIL, suggesting that apoptosis of myeloma cells induced by PU.1 might be at least partially due to TRAIL upregulation. We are currently performing DNA microarray analysis to compare the expression levels of genes between before and after PU.1 induction, in order to further elucidate the mechanisms of cell growth arrest and apoptosis.
Toward Defining the Functional Relevance of Wnt Signaling in Osteoclasts in Multiple Myeloma.
Authors: Qiang Ya-Wei , Chen Yu , Brown Nathan , Ojha Rohit P. , Rudikoff Stuart , Barlogie Bart , Shaughnessy John D. 
Published: October 2019

Abstract
Disruption of Wnt signaling is thought to play a central role in the development of myeloma bone disease. Although Wnt signaling in osteoblasts (OB) is essential for their differentiation and Wnt signaling in OB indirectly regulates osteoclast (OC) development through the regulation of both OPG and RANKL, little is known about the direct role of Wnt signaling in osteoclastogenesis. In this study, we sought to characterize the Wnt signaling pathway and its functional role in OC formation using the macrophage cell line Raw 264.7, which can be induced to differentiate into OC, as well as human primal progenitor osteoclast cells isolated from bone marrow of normal donors and patients with MM. We first analyzed the pattern of Wnt signaling components in these cells by RT-PCR and sequence analysis. High level of expression of Wnt receptors Frizzled (Fz) Fz2, 3, 4, 5, 7, 8 and 9, and co-receptors LRP5, and LRP6, and transcription factors TCF1, TCF3, and LEF1 was observed in 10 of 10 OC cells from MM patients and 4 of 4 OC from normal donors and Raw264.7. Wnt3a conditioned medium and recombinant Wnt3a induced increased levels of total and un-phosphorylated beta-catenin in all cells. Pretreatment of OC cells with sFRP1 decreased Wnt3a induced increased beta-catenin. Pretreatment with DKK1, a specific inhibitor of the canonical Wnt pathway, blocked Wnt3a induced stabilization of beta-catenin. Additionally, the GSK3beta inhibitor lithium chloride induced stabilization of beta-catenin in OC and Raw264.7 cells in dose-dependent fashion. Wnt-3a induced TCF/LEF transcriptional activity in Raw264.7 cells transfected with TOPflash compared with Raw264.7 transfected with FOPflash, which possess mutant sequence for binding beta-catenin. These results suggest that Wnt signaling functions in primal MM OC cells, as well as OC cell lines. Finally, we sought to determine the biological function of Wnt signaling in OC cells. The results showed that Wnt-3a did not affect the proliferation and survival of OC cells, nor increased formation of TRAP positive cells. Wnt3a did not synergize with RANKL and M-CSF in induction of OC differentiation. Additional biological functional assays are underway to determine the direct role of Wnt signaling on OC. Identification of the role of Wnt signaling in osteoclastogenesis may improve our understanding of the bone lytic process in multiple myeloma.
Inhibition of p38α MAPK Reduces Tumor Burden, Prevents the Development of Myeloma Bone Disease, and Increases Survival in the 5T2 and 5T33 Murine Models of Myeloma.
Authors: Vanderkerken Karin , Medicherla Satya , Coulton Les , Van Camp Benjamin , Protter Andy , Higgins Linda , Menu Eline , Croucher Peter 
Published: October 2019

Abstract
The bone microenvironment plays a critical role in supporting the growth and survival of myeloma cells and the development of osteolytic bone disease. Signalling through p38 α MAPK mediates synthesis of myeloma cell survival factors by stromal cells; whereas, inhibiting p38 α MAPK reduces myeloma cell proliferation and inhibits osteoclast formation in vitro. However, it is unclear whether p38 α MAPK inhibition will prevent the growth and survival of myeloma cells and the bone disease in vivo. The aim of this study was to determine whether SCIO-469, a selective p38 α MAPK inhibitor, would inhibit myeloma growth and prevent the development of bone disease in the 5TMM syngeneic models of myeloma.
Treatment of 5TMM cells, in vitro, with SCIO-469 resulted in a clear inhibition of p38 phosphorylation, as assessed by Western blotting and an inhibition up to 35% of stromal cell induced 5T33MM proliferation.
Injection of 5T2MM murine myeloma cells into C57Bl/KaLwRij mice resulted in the growth of myeloma in bone and the development of bone disease characterized by increased osteoclast surface (p<0.05), a reduction in cancellous bone (p<0.01) and the presence of osteolytic bone lesions on x-ray (p<0.01). Treatment of 5T2MM-bearing mice with SCIO-469 (150mg/kg in the diet, therapeutical treatment from paraprotein detection) resulted in a 42% decrease in serum paraprotein and prevented development of osteolytic lesions (p<0.01). Injection of 5T33MM cells into C57Bl/KaLwRij mice also resulted in the development of myeloma but not associated bone disease. Treatment of 5T33MM-bearing mice from the time of tumor cell injection with SCIO-469 resulted in a decrease in serum paraprotein (8.8+/−1.4g/dl to 0.04+/− 0.03g/dl, p<0.001) and a reduction in the proportion of tumor cells in the bone marrow (67 +/− 8.1% to 1.09 +/− 0.58%, p<0.001). Kaplan-Meier analysis demonstrated an increase in disease-free survival (veh=27.5 days vs 96 days, p<0.001) after treatment of the mice with SCIO-469.
These data demonstrate that targeting p38 α MAPK with SCIO-469 is associated with an anti-myeloma effect, which indirectly prevents the development of myeloma bone disease.