bluuebunny/crossref_metadata_2025_split huggingface dataset:
IRF-4/MUM-1 Expression Is a Critical Switch in the Generation of Plasma Cells Versus Memory B-Cells.
Authors: Klein Ulf , Casola Stefano , Cattoretti Giorgio , Shen Qiong , Lia Marie , Mo Tongwei , Ludwig Thomas , Rajewsky Klaus , Dalla-Favera Riccardo 
Published: April 2020

Abstract
Most types of human B-cell lymphomas harbor somatically mutated Ig variable region genes, reflecting their origin from cells that have undergone the germinal center (GC) reaction of T-dependent immune responses. The lymphomas exhibit diverse phenotypes and clinical behaviors, likely as a consequence of differences both in the mechanisms of transformation and in the specific target cell. We have recently identified two distinct subsets of GC B-cells that may represent late stages of the GC-reaction and may be the precursors of plasma cells and memory B-cells. The corresponding subsets are characterized by downregulation of the GC-marker BCL6 and the alternative expression of IRF-4/MUM-1 or nuclear c-Rel. These two subsets seem to reflect distinct cellular programs which are altered during B-lymphomagenesis in various tumor subtypes which co-express BCL6, IRF-4 and nuclear c-Rel, an event never observed in normal B-cells. In order to gain insights into the physiologic role of IRF-4/MUM-1 and nuclear c-Rel in GC-development, we are ablating their expression specifically in mouse GC B-cells. Transgenic mice were generated that carry an IRF-4/MUM-1 null allele and a conditional IRF-4/MUM-1 allele which, following Cre-mediated deletion of the loxP-flanked promotor region and exons 1 and 2, expresses eGFP, thus allowing to track the development of the IRF-4/MUM-1 deficient cells at the single cell level. IRF-4/MUM-1fl/- mice were crossed with transgenic mice that express the Cre-recombinase specifically in B-cells undergoing class-switch to IgG1, an event occurring in a large fraction of GC B-cells. Upon immunization with sheep red blood cells or nitrophenyl-(NP)-KLH, mice unable to express IRF-4/MUM-1 in late GC B-cells (IRF-4/MUM-1fl/-/Cγ 1Cre/+), in contrast to control mice (IRF-4/MUM-1fl/+/Cγ 1Cre/+), did not develop plasma cells (IgG1+CD138+) in the peripheral lymphoid organs, blood, and bone marrow. On the other hand, the generation of memory B-cells appears normal since antigen-specific B-cells were present in the spleen (eGFP+B220+PNA-IgG1+) and blood (eGFP+B220+CD38+IgG1+). These results suggest that the IRF-4/MUM-1 gene product is required for the development of antigen-selected GC B-cells into plasma cells. We suggest that the expression of IRF-4/MUM-1 in a GC centrocyte is the critical event in the commitment of B-cells to differentiate into a plasma cell versus a memory B-cell, and are currently testing the role of nuclear c-Rel in the same process.
Zoledronate Exerts Direct Antiproliferative and Apoptotic Effects on Human Myeloma Cells In Vitro and In Vivo.
Authors: Guenther Andreas , Gordon Sharon , Bakker Frank , Burger Renate , Tiemann Markus , Green Jonathan R. , Rogers Michael , Gramatzki Martin 
Published: April 2020

Abstract
Zoledronate (ZOL) is the most potent nitrogen-containing bisphosphonate and is effective at preventing osteolytic bone disease in patients with multiple myeloma (MM) and solid tumors. ZOL inhibits the enzyme farnesylpyrophosphate synthase and thus blocks the prenylation of small GTPases. In vitro studies have demonstrated that ZOL can also directly affect the growth and viability of myeloma cells, however, the molecular mechanisms underlying this activity have not been fully elucidated. The goal of our study was to investigate direct antimyeloma effects of ZOL in vitro and in vivo. In five myeloma cell lines (RPMI8226, L363, U266, JK-6L, and the IL-6 dependent INA-6), growth was inhibited and apoptosis induced by ZOL in a dose-dependent manner (IC50’s between 30 μM and 285 μM). Similar results were obtained in the presence of bone marrow stromal cells, IL-6 (20 ng/mL), IGF-1 (200 ng/mL), or a combination of both cytokines. The potential antitumor effect of ZOL on myeloma cells in vivo was studied in the INA-6 SCID model, in which mice are injected intraperitoneally with INA-6 cells and subsequently develop plasmacytomas. Mice treated with ZOL had reduced tumor burden and a significant survival benefit compared to the control group (p=0.002). Histological examination of plasmacytomas explanted 72 hours after a single injection of 8 μg ZOL revealed extensive apoptotic/necrotic areas while no such areas were found in tumors of untreated animals. Induction of apoptosis was confirmed by Western blot analysis of tumor lysates, which revealed increased levels of cleaved poly (ADP-ribose) polymerase (PARP) in tumors of ZOL treated vs. untreated animals. This correlated with an accumulation of the unprenylated form of the small GTPase Rap1A, which was virtually absent in tumors of untreated mice. Our findings demonstrate a direct and specific effect of ZOL in plasmacytomas in vitro and in vivo and point to a therapeutic potential in MM beyond the prevention of osteolytic lesions.
Macrofocal Multiple Myeloma in Young Patients: A Distinct Entity with Favorable Prognosis.
Authors: Dimopoulos Meletios A. , Pouli Anastasia , Angnostopoulos Athanasios , Repoussis Panagiotis , Symeonidis Argyris , Delimbasi Souzana , Tsolakis Fotis , Zervas Constantinos 
Published: April 2020

Abstract
Introduction: Most patients with multiple myeloma (MM) present with multiple lytic bone lesions, extensive marrow plasmacytosis and anemia. Furthermore in several of them hypercalcemia and renal impairment are evident at diagnosis. Over the years, we have seen occasional young patients with MM who presented with multiple lytic bone lesions but without intervening infiltration of bone marrow i.e. a pattern consisting of macrofocal MM.
Patients and methods: In order to assess the clinical and laboratory features and the outcome of patients with macrofocal MM we performed a retrospective analysis of symptomatic patients with MM ≤40 years of age at diagnosis who received primary treatment over a 20 year period (from January 1, 1985 to December 31, 2004). The diagnosis of macrofocal MM required the presence of lytic bone lesions and the absence of obvious bone marrow plasmacytosis (BMPC&lt;10%) on bone marrow aspirate and/or biopsy.
Results: Among 51 patients ≤40 years at the time of initial treatment, 10 patients fulfilled the criteria of macrofocal MM. Patients’ median age, gender, myeloma heavy and light chain were similar among patients of the 2 groups. When compared with patients with typical MM, patients with macrofocal pattern were less anemic (p=0.018), none had hypercalcemia (p=0.1), renal impairment (p=0.17), elevated serum LDH (p=0.14), or stage 3 according to the International Staging System (ISS) (p=0.13). Four of the 10 patients with macrofocal MM versus 0 of 41 patients with typical MM (p=&lt;0.01) had a prior diagnosis of solitary bone plasmacytoma. Approximately 80% of patients in both groups received primary treatment based on high-dose dexamethasone regimens (VAD or VAD-like regimens). An objective response was noted in 55% of patients with macrofocal MM and in 50% of patients without this pattern. The median survival of patients with typical MM was 57 months and has not been reached in patients with macrofocal MM (p=0.087). In the latter group of patients, median survival is projected to exceed 8 years. The 10 year survival rate is 20% and 5% in patients with or without macrofocal myeloma respecrively.
Conclusion: Our analysis indicates that macrofocal MM is a distinct entity in young patients with MM. Despite multiple lytic bone lesions, such patients have features of low tumor burden and an improved survival when compared with young patients with typical MM. Future studies including gene profiling may reveal potential biological differences among the two groups of patients.
Serum Pleiotrophin Is a New Multiple Myeloma Tumor Marker That Also Predicts Clinical Status.
Authors: Yeh Howard S. , Chen Haiming , Manyak Steven J. , Swift Regina A. , Campbell Richard A. , Wang Cathy , Li Mingjie , Jin Hee , Gordon Melinda S. , Bonavida Benjamin , Berenson James R. 
Published: April 2020

Abstract
Pleiotrophin (PTN), an 18 kD heparin-binding protein, has been found to be expressed in some solid tumors whereas it has not been previously evaluated in hematologic malignancies. Constitutive expression of PTN promotes tumor expansion and may increase metastatic potential. This protein binds to syndecan-1 (CD138) and stimulates angiogenesis. Its role in multiple myeloma (MM), thus far, has not been evaluated. Recently, we showed that PTN was strongly expressed in MM cell lines and malignant plasma cells from fresh bone marrow samples but not in normal control bone marrow specimens. Since PTN has been shown to be elevated in the serum of some solid tumor patients, we determined whether PTN may be elevated in the serum of MM patients, and correlated these levels with the patients’ disease status. By using a highly sensitive and specific enzyme-liked immunosorbent assay (ELISA) for PTN, we tested 270 different serum samples [MM (n=194), MGUS (n=18) and age-matched healthy control subjects (n=58)]. Serum samples from MM patients were obtained at the time of diagnosis as well as during the course of their disease. Serum PTN levels were higher in MM patients than in the control subjects (median=1.44 ng/ml vs 0.42 ng/ml, p &lt;0.0001). In addition, among patients with MGUS and smoldering myeloma, serum PTN was also higher than in the controls (median=1.14 ng/ml vs 0.42 ng/ml, p &lt;0.0001) but lower than among patients with active MM (p &lt;0.0001). Patients with untreated MM showed serum PTN levels similar to those patients who received prior treatment. Among previously treated patients, the group with progressive disease at the time of evaluating their PTN level had higher serum concentrations of this protein than patients with responsive or stable disease (median=1.73 ng/ml vs 1.16 ng/ml, p &lt;0.0001). Evaluation of patients who had a change in disease status showed that the serum PTN level increased at the time of progression when compared with their baseline level (median=1.76 ng/ml vs 1.01 ng/ml, p=0.009). Conversely, patients who responded to anti-MM therapy exhibited significant decreases of PTN as compared with their pre-treatment PTN values (median=0.95 ng/ml vs 1.90 ng/ml, p=0.007). These results suggest that serum PTN may be a new tumor marker for MM, and monitoring of this protein may be useful to follow the responsiveness to therapy and disease status of MM patients. We are now undertaking studies to evaluate PTN as a prognostic marker in a large cohort of newly diagnosed MM patients.
Mutations of Myeloid Transcription Factors AML1, CEBPα and PU.1 in Patients with De Novo Myelodysplastic Syndrome: An Analysis of Paired Bone Marrow Samples at Diagnosis and at AML Transformation.
Authors: Shih Lee-Yung , Liang Der-Cherng , Huang Chein-Fuang , Wu Jin-Hou , Wang Po-Nan , Kuo Ming-Chung , Dunn Po , Tang Tzung-Chih , Lin Tung-Liang , Chang Hung 
Published: April 2020

Abstract
Transcription factors AML1, PU.1 and CEBPα are essential for normal hematopoiesis. Recently, somatic mutations of genes encoding these transcription factors have been described in myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). The role of mutations of these transcription factors in the progression of MDS into AML remained to be determined, as no matched paired samples of both phases have been analyzed in the previous studies. We analyzed mutations of AML1, CEBPα and PU.1 on paired bone marrow samples at initial diagnosis of de novo MDS (8 RCMD, 12 RAEB-1, and 17 RAEB-2) and at AML transformation. Mutation analysis was performed by direct sequencing of all RT-PCR or DNA PCR products using different primer pairs which cover the entire coding sequences of AML1, PU.1 and CEBPα genes. Eight of 37 patients (21.6%) had AML1 mutations and one of them had 2 mutations; 3 mutations were located in Runt homology domain (RHD) (exons 3–5), whereas 6 mutations were located in the C-terminal region (exons 6–8) at initial presentation of MDS. Another 2 acquired C-terminal mutations at AML transformation. The 11 AML1 mutations detected in 10 patients during their disease course, included 4 missense mutations, 3 nonsense mutations, 1 silent mutation, and 3 frameshift mutations. Taken together, 30% (3/10) of AML1 mutations detected in MDS patients were located in RHD and were exclusively missense compared with 70% at C-terminal region with frequent nonsense or frameshift mutations (86%) (P=0.033). Patients with AML1 mutations were older than those without mutations (P=0.019). No differences were observed between patients with and without AML1 mutations with respect to sex, blood counts, percentage of blasts in bone marrow or peripheral blood, morphologic subtype, and cytogenetic risk group. There was no difference in the time to AML transformation and overall survival with respect to the status of AML1 mutation. Only one out of 34 MDS patients had CEBPα mutation at initial diagnosis, she lost the mutation after chemotherapy and no mutation was detected at AML transformation. Another one did not have CEBPα mutation at diagnosis but acquired CEBPα mutation at AML transformation, the patient also had AML1 mutation at both phases. PU.1 mutation was not detected in 32 patients examined at both MDS and AML phases. The present study showed that AML1 mutations were common in patients with MDS, especially at C-terminal region and frequently with frameshift or nonsense mutations, and might be acquired during AML transformation; whereas CEBPα mutations were infrequent and PU.1 mutation was absent.
Lenalidomide Effects on NK Function in Patients with Myelodysplastic Syndrome (MDS).
Authors: Epling-Burnette P.K. , Bai Fanqi , Painter Jeffrey S. , Djeu Julie Y. , List Alan F. 
Published: April 2020

Abstract
Lenalidomide, which is a 4-amino-glutarimide analogue of thalidomide, has significant erythropoietic activity in patients with lower-risk MDS (List et al, NEJM, 351:26,2004). Although its precise target of action in MDS is not known, lenalidomide modulates cellular response to varied stimuli including inhibition of angiogenic response and endotoxin induction of inflammatory cytokines and enhancement of antigen-induced immunologic response and erythropoietin receptor signaling. Clinical investigations in multiple myeloma indicate that the immunomodulatory effects of thalidomide and lenalidomide extends to the expansion of natural killer (NK) cells. We recently found that MDS patients have defective NK function, araising in part to reduced expression of activating NK receptors (NKRs) NKp30, CD244 (2B4), and NKG2D. To determine if lenalidomide may restore NK function in MDS, we investigated the effects of in vitro treatment with lenalidomide on NK function and phenotype. Lytic function was studied using peripheral blood mononuclear cells (PBMCs) as effector cells and the leukemia cell line, K562, as a target in standard 4-hr 51Cr-release assays at 12:1 and 25:1 effector:target (E:T) ratios. Among eight MDS patient’s specimens evaluated, five patients had significant increase in tumor lysis after treatment with 1 μM lenalidomide for 72 hours (p ≤ 0.01, T-test). In similar experiments using PBMCs from normal donors, we found that NK lysis of K562 was 42% ± 15 (25:1 E:T ratio) pre-treatment which increased to 71% ± 17 (25:1 E:T ratio) after treatment, which was statistically significant (p ≤ 0.01, T-test). We also examined the in vitro effects of lenalidomide on lytic activity by the NK cell lines, NK92 and NKL, which was significantly increased after drug treatment. To discern the mechanisms of lenalidomide action in NK cell lines and normal NK cells, we evaluated NKR display by flow cytometry, and NKR function by antibody redirected cytotoxicity using the FcγR+ murine mastocytoma (P815) target cell line. Using NK92 and NKL cells, treatment with lenalidomide 1 μM for 72 hrs increased lysis by anti-NKG2D and anti-CD244 activating antibodies. We also found that NKG2D surface expression was increased on normal NK cells after lenalidomide treatment in vitro. These results suggest that some MDS patients may have improved NK function through the immunomodulatory effects of lenalidomide. The relationship between in vitro NK responsiveness to lenalidomide and in vivo hematological response warrants investigation in patients with MDS.
Comprehensive Promoter Methylation Analysis of Genes on 5q31.1 in MDS and AML.
Authors: Oki Yasuhiro , Jelinek Jaroslav , Shen Lanlan , Issa Jean-Pierre 
Published: April 2020

Abstract
Interstitial deletion or loss of chromosome 5 is a frequent chromosomal abnormality in myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). Previous studies uncovered a 1.5–2Mb region on 5q31.1 to be a common deleted region (CDR) in those diseases. It is believed that there exists one or more of critical tumor suppressor gene(s) in this CDR. Mutation analysis, however, failed to identify target gene(s). With a hypothesis that epigenetic silencing may play a significant role as a “second hit” in MDS/AML, we performed a comprehensive methylation analysis of genes in this CDR on 5q31.1. Among 25 genes that are located in this region (SPOCK-AF116688, covering CDR determined by Horrigan et al. Blood 2000 and Fang et al. Genomics 2000), 14 genes had promoter regions in CpG islands that were therefore susceptible to methylation dependent silencing. Methylation status of multiple CpG sites in promoter regions of these those 14 genes (SPOCK, HNRPA1, PKD2L2, C5orf5, KIF20A, GFRA3, CDC25C, C5orf6, JMJD1B, C5orf19, EGR1, ETF1, HSPA9B, CTNNA1, SIL1) was analyzed using bisulfite-PCR followed by pyrosequencing. Analyzed samples were 13 leukemic cell lines as well as peripheral blood mononuclear cells from 5 patients with AML with 5q deletion (5qAML), 9 AML without 5q deletion (non-5qAML), 17 MDS with 5q deletion (5qMDS), 9 MDS without 5q deletion (non-5qMDS) and 10 individuals without leukemia. Significant hypermethylation (&gt;15% by pyrosequencing) was observed in 5 genes in patient samples (SPOCK, PKD2L2, GFRA3, HSPA9B, and ETF1, see Table 1) and 3 other genes in cell lines only (CDC25C, C5orf19, CTNNA1). In 5qMDS, the most frequently methylated genes were ETF1 (5/17), GFRA3 (4/17), and HSPA9B (11/17), but HSPA9B also showed methylation in normal blood. We then evaluated the expression levels of 5 genes that showed significant methylation in patients’ samples using quantitative RT-PCR. SPOCK, PKD2L2, GFRA3 were not expressed in normal PBMC. HSPA9B expression levels did not correlate with the degree of methylation. On the other hand, ETF1 expression was inversely correlated with the degree of methylation. ETF1 is a eukaryotic translation termination factor, which plays a role in recognizing stop codon in mRNA and terminates translation. Based on specific hypermethylation and silencing in 5qMDS, we propose ETF1 as a candidate tumor suppressor gene in the 5q31.1 region. Functional analysis of ETF1 in leukemic cells is underway.
Frequency of promoter CpG sites methylation Samples N SPOCK PKD2L2 GFRA3 HSPA9B ETF1 Cell lines 13 62% 46% 77% 38% 0% 5qAML 5 0% 0% 0% 20% 0% Non-5qAML 9 33% 0% 0% 0% 0% 5qMDS 17 6% 6% 24% 65% 29% Non-5qMDS 9 11% 11% 33% 33% 11% Normal 10 0% 0% 0% 90% 0%
Systemic Treatment with the Antiangiogenic Agent ABT-510 Inhibited Myeloma-Induced Microvessels and Tumor Growth in Myelomatous SCID-hu Mice.
Authors: Yaccoby Shmuel , Ling Wen , Saha Rinku , Yata Kenichiro , Tricot Guido 
Published: April 2020

Abstract
Myeloma cells induce bone marrow angiogenesis and increased microvessel density (MVD) is associated with poor prognosis in this disease. The aim of this study was to investigate whether neovascularization is required for myeloma growth by testing the anti-angiogenesis and anti-tumor efficacy of thrombospondin-1 (TSP-1)-derived mimetic peptide, ABT-510 (Abbott Laboratories), in the SCID-hu model for primary myeloma. TSP-1 is a matricellular glycoprotein with multiple biological functions, including antiangiogenic activity. ABT-510 is a capped nonapeptide based on the linear TSP-1 heptapeptide sequence. Like TSP-1, ABT-510 can be targeted to a specific receptor on vascular endothelial cells and efficiently inhibits angiogenesis in vitro and after systemic injection in experimental models of human malignancies. SCID-hu mice were constructed as previously described (Yaccoby et al., Blood 1998; 1999; 2002). Myelomatous SCID-hu mice were prepared by injection of myeloma cells from 6 patients into the implanted human bone in the hosts. Changes in levels of tumor burden were monitored by weekly measurements of human monotypic immunoglobulins (hIg) using ELISA and histologically confirmed by immunohistochemical staining of human bone sections for cIg. MVD, visualized by CD34 staining, were counted in 4 non-overlapping areas. Upon establishment of myeloma growth (401±146 μg/ml pre-treatment levels) mice were intraperitoneally injected with ABT-510 (100 mg/kg, twice a day) or with the vehicle (5% dextrose) for 4–10 weeks. Treatment was well tolerated and no sign of toxicity or reductions in body weight were observed. Whereas all dextrose -treated mice had increased hIg levels during the experimental period, ABT-510 treatment resulted in marked reduction of tumor burden in 2 experiments by 48% and 20%, respectively, retardation of myeloma growth in 3 additional experiments and no response in one experiment. Overall, tumor burden in control- and ABT-510-treated mice was increased by 3696%±3264 and 448%±169 from pre-treatment levels, respectively (p&lt;0.03). MVD in control- and ABT-510-treated mice were 21±6/mm2 and 11±4/mm2, respectively (p&lt;0.02). Intriguingly, MVD in the non-responding mouse was reduced by &gt;35% (11±2 vs. 17±1 microvessels/MM2 in control mouse, p&lt;0.04), suggesting that certain myeloma cells are not dependent on neovascularization. We conclude that myeloma cells from the majority of patients require angiogenesis for growth and that ABT-510, which is undergoing a phase II clinical trial for treating solid tumors, is a potential agent for treating myeloma.