bluuebunny/crossref_metadata_2025_split huggingface dataset:
Immunoglobulin Heavy Chain (IgH) Gene Somatic Hypermutation Status Using Homoduplex-Separated RT-PCR Products.
Authors: Melan Melissa A. , Foon Kenneth A. , Marks Stanley M. , Kant Jeffrey A. 
Published: April 2020

Abstract
Assessment of IgH somatic hypermutation status has been shown to be a valuable indicator for judging the prognosis of patients with chronic lymphocytic leukemia (CLL). Our laboratory has developed a streamlined method to improve the rate of successful evaluation of IgH mutation status. Six individual PCR reactions are first performed using random hexamer-generated cDNA as template. These reactions have identical reaction parameters, use a common JH reverse primer and one of six VH class-specific forward primers within Framework 1. PCR products are separated on acrylamide or MetaPhor® agarose gels following formamide denaturation. In almost all cases, a single homoduplex band is resolved indicating a class-specific clonal product. The homoduplex band is excised from the gel, eluted and directly sequenced. Mutation analysis is performed using the NCBI Ig BLAST program with percent identity determined for the region from the beginning of CDR1 to the end of Framework 3. To date, less than 10% of cases analyzed have not yielded a clearcut clonal PCR product using this approach
The Effect of Subsequent Therapies in the Patients with Chronic Lymphocytic Leukemia Previously Treated with Cladribine and Prednisone or Chlorambucil and Prednisone in Randomized Studies.
Authors: Robak Tadeusz , Blonski Jerzy Z. , Kasznicki Marek , Gora-Tybor Joanna , Dmoszynska Anna , Wojtaszko Maria , Skotnicki Aleksander , Nowak Wieslaw , Hellmann Andrzej , Lewandowski Krzysztof , Zdziarska Barbara , Konopka Lech , Ceglarek Bernadeta , Dwilewicz-Trojaczek Jadwiga , Boguradzki Piotr , Kuliczkowski Kazimierz , Sulek Kazimierz , Warzocha Krzysztof 
Published: April 2020

Abstract
Previously, we reported a study on efficacy and toxicity of cladribine (2-CdA) + prednisone (P) compared with chlorambucil (Chl) + P in previously untreated patients with progressive or symptomatic chronic lymphocytic leukemia (CLL) in a randomized multicenter trial (Blood2000, 96, 2723). Of 229 patients enrolled, 126 received 2-CdA+P and 103 received Chl+P. Here, we present the long-term follow-up and the results of subsequent treatment in refractory or relapsed patients. Patients with non responsive (NR) disease or with progression after 3 courses of 2-CdA + P or Chl+P or who relapsed earlier then 12 months after completing the treatment were switched to an alternative arm. Patients who relapsed later than 12 months from first remission (late relapse) were retreated with the same schedule that induced previous response.
In 50 patients, non-responsive or with early relapse, treated originally with Chl+P who received 2-CdA+P as second line, complete response (CR) was achieved in 12 (24%) and overall response (OR) in 32 (64%). Twenty eight patients originally treated with 2-CdA+P received Chl+P and CR and OR was obtained in 1 (3,6%) and in 6 (21.4%), respectively (p=0.002 for both arms).
Thirty three patients with late relapse were retreated with 2-CdA+P and 19 patients were retreated with Chl+P. OR (CR) was 54.6% (6.1%) and 47.4% (15.8%), respectively (p=0.34).
The third line treatment was CHOP in both arms. Twenty seven patients from 2-CdA+P group and 37 patients from Chl+P group were treated with CHOP and only one patient responded.
Median survival was 3.3 and 3.75 years, respectively (p=0.63). Secondary neoplasms were observed in 8 (6.4%) patients treated with 2-CdA+P and in 4 (3.9%) treated with Chl+P as first line treatment (p=0.6). Death rates have been similar in patients treated originally with 2-CdA+P 82 (64.6%) and Chl+P 69 (66.4%) (p=0.8).
In conclusion, cladribine is significantly more effective as a second line treatment than chlorambucil. Both agents are similarly effective in retreatment of late relapsed patients. CHOP is of no value in patients previously treated with 2-CdA + P and with Chl + P. There is no significant difference in median survival of patients treated initially with Chl + P and 2-CdA + P.
Structural and Functional Characterization of the NHR2 and Runt Domains of AML1/ETO.
Authors: Cheney Matthew D. , Liu Yizhou , Zhou Yunpeng , Chruszcz Maksymilian , Laue Thomas M. , Minor Wladek , Bushweller John H. , Speck Nancy A. 
Published: April 2020

Abstract
AML1/ETO is the chimeric fusion protein resulting from the t(8;21) found in AML of the M2 subtype. It contains the N-terminal 177 amino acids of RUNX1 and virtually all (575aa) of ETO. The RUNX1 component includes the Runt domain, which mediates both DNA binding and heterodimerization with CBFβ, but lacks the more C-terminal sequences required for transactivation. AML1/ETO occupies RUNX target genes in vivo and is associated with a repressive chromatin structure characterized by reduced levels of acetylated histone H3. AML1/ETO is thought to repress transcription by recruiting a SMRT (N-CoR)/Sin3A/HDAC complex to chromatin via sequences in ETO. ETO is the human homologue of the Drosophila Nervy protein and shares 4 regions of homology with Nervy called Nervy Homology Regions (NHR) 1–4. Deletion studies have shown that three of the AML1/ETO domains essential for its repressive function are the Runt domain, NHR2, and NHR4. The NHR2 domain is a hydrophobic heptad repeat that mediates oligomerization of AML1/ETO, interaction with ETO family members, and also with mSin3A and HDACs. We recently solved an x-ray structure of the NHR2 domain and found it to be an alpha-helical tetramer. Based on this structure we have introduced amino acid substitutions into the NHR2 domain that disrupt tetramer formation but not AML1/ETO stability. These mutations impair the ability of AML1/ETO to inhibit the differentiation of GR-1+/Mac-1+ cells following retroviral transduction into primary mouse bone marrow cells, and also inhibit the serial replating ability of AML1/ETO expressing bone marrow cells in vitro. We additionally show that mutations reported by Amann et al. (Mol Cell Biol. 21, 6470, 2001) to disrupt mSin3A binding to NHR2 do not affect the biological activity of AML1/ETO in vitro. We also introduced mutations in the Runt domain of AML1/ETO that disrupt CBFβ binding by defined amounts (40-fold, 200-fold, 500-fold), and demonstrated that CBFβ binding by AML1/ETO is essential for its dominant negative activity. The latter results suggest that small molecules designed to selectively impair heterodimerization of AML1/ETO with CBFβ could potentially block AML1/ETO’s dominant negative activity.
Cpg Oligodeoxynucleotide-Mediated Effects on B-Cell Chronic Lymphocytic Leukemia In Vitro Are Influenced by Cytogenetic Status and the Presence of Serum.
Authors: Jahrsdorfer Bernd , Blackwell Sue E. , Wooldridge James E. , Taylor Christiana M. , Weiner George J. 
Published: April 2020

Abstract
Immunostimulatory CpG-containing oligodeoxynucleotides (CpG ODN) are TLR9 agonists that mediate a number of immunologic effects in normal and malignant B cells including upregulation of immunogenic molecules. They are therefore felt to be attractive as potential components of immunotherapy for B cell chronic lymphocytic leukemia (B-CLL). The cytogenetic status of B-CLL is predictive of clinical prognosis, but little is known about how CpG ODN-mediated effects on B-CLL cells correlate with cytogenetic status. The present study was designed to explore the impact cytogenetic status has on in vitro B-CLL cell survival and immunogenicity. Since most in vitro studies to date have been performed under serum-low conditions, we also evaluated how the presence of autologous serum or plasma impacts on CpG ODN-mediated effects on B-CLL cells. B-CLL cytogenetic status by interphase FISH, as well as immunophenotype and cell survival in the absence or presence of CpG ODN was determined in 23 samples. CpG ODN decreased in vitro survival of B-CLL cells with good prognosis cytogenetics, but had little effect on cells with poor prognosis cytogenetics. In contrast, CpG ODN upregulated costimulatory and antigen-presenting molecules and enhanced allogeneic T cell response in samples with good and poor prognosis cytogenetics. The optimal concentration for CpG ODN-mediated effects in the presence of 100% autologous serum or plasma was higher (10–20 micrograms/ml ODN) than for serum-low conditions (2.5–5 micrograms/ml ODN). The observed inhibition of CpG ODN-mediated effects by serum directly correlated with a lower uptake of ODN into B-CLL cells in the presence of serum. In conclusion, CpG ODN induced changes in B-CLL consistent with enhanced immunogenicity in all samples studied, but induced apoptosis most effectively in the subset of B-CLL cells with good prognosis cytogenetics. Approximately 4-fold more CpG ODN was needed to induce these changes when serum was present as compared to low serum conditions due to decreased ODN uptake by the cells in the presence of serum. These studies suggest CpG ODN may be useful as immunotherapeutic agents for B-CLL irrespective of cytogenetic status because of their potential effects on immunogenicity. Higher systemic doses of CpG ODN than previously thought might be necessary to induce these changes because of the effect of serum proteins on ODN uptake.
Optophoresis Accurately Predicts Response to Chemotherapy in CLL.
Authors: McNeeley Patricia , Saven Alan , Kariv Ilona , Nieva Jorge , Marchand Philippe , Diver Jonathan , Riccitelli Samuel , Nerenberg Michael 
Published: April 2020

Abstract
In vitro drug resistance assays have shown efficacy in predicting response to chemotherapeutic drugs in a number of solid tumors. In chronic lymphocytic leukemia (CLL) many assays are limited by the difficulty involved in culturing these cells ex vivo. We have developed a novel, optical-based methodology that is sensitive to broad cellular physical characteristics, such as morphology, size, refractive index, density and surface properties. This measurement, known as Optophoresis, quantifies cell motion induced by exposure to a moving optical gradient, generated from a near-infrared laser beam (Forster AH, et al. Anal Biochem 2004, 327(1):14–22 and Wang MM, et al. Appl Opt 2003, 42(28):5765–73). In Optophoresis small numbers of cells are analyzed intact, in their native state. No labels are required for quantification of functional responses, but cell subpopulations may be identified using fluorescent tags. We have used this assay to predict response to chemotherapeutic agents in patients with CLL.
Methods: We performed Optophoresis with six drugs (fludarabine, chlorambucil, vincristine, cyclophosphamide, cladribine and prednisolone) on 74 CLL patient samples. 68 of these were classic B-CLL; there was one hairy cell variant, two were T-CLL, two evolved to PLL and one evolved to lymphoma. 21 of the samples were from patients for whom clinical data on response to chemotherapy was available for 33 drug treatments. Response to therapy was defined as a decrease in RAI stage.
Results: 21 patients aged 58 to 93 years; 71% Rai stage III or IV were included in this analysis. Six patients were evaluated prospectively, with the assay performed before chemotherapy administration, and 15 patients were evaluated retrospectively with the assay performed after a course of treatment. Optophoresis accurately predicted response to chemotherapy in 88% of the 33 treatments evaluated. In the four instances for which the Optophoresis results did not match the retrospective clinical outcome; the patient was historically sensitive to treatment and Optophoresis results indicated current resistance to the drug. Subsequent resistance to drug therapy is known to occur in a significant percentage of treated CLL patients.
Conclusion: Optophoresis of CLL cells accurately predicts response to chemotherapy in CLL. Further studies using results of Optophoresis to guide CLL treatment are warranted.
Combined Disruption of Both the MEK/ERK and the IL-6R/STAT3 Pathways Is Required To Induce Apoptosis of Multiple Myeloma Cells in the Presence of Bone Marrow Stromal Cells.
Authors: Chatterjee Manik , Stuehmer Thorsten , Herrmann Pia , Bommert Kurt , Dorken Bernd , Bargou Ralf C. 
Published: April 2020

Abstract
The IL-6R/STAT3 pathway has been reported to critically contribute to the pathogenesis of multiple myeloma (MM) and to protect MM cells from apoptosis. However, recently we could demonstrate that MM cells become independent of the IL-6R/STAT3 pathway if they are cocultured with bone marrow stromal cells (BMSCs), suggesting that the BM microenvironment stimulates IL-6-independent pathways that exert a pro-survival effect. It was therfore the aim of this study to analyze the underlying mechanism of this phenomenon. Pathway analysis revealed that BMSCs stimulate STAT3 via the IL-6R, and MAPK in parts via IL-6R-independent mechanisms. Abolition of MEK1, 2 activity with PD98059, or of ERK1,2 through siRNA constructs, was insufficient to induce apoptosis. However, the combined disruption of the IL-6R/STAT3 and MEK1,2/ERK1,2 pathways led to strong induction of apoptosis even in the presence of BMSCs. Thus, disruption of the MEK/ERK pathway restores IL-6/STAT3 dependence of MM cells in the presence of BMSCs indicating that BMSC-mediated induction of the MEK/MAPK pathway is the mechanism by which BMSCs render MM cells IL-6/STAT3 idependent. Consequently, in the presence of cells from the BM microenvironment the combined targeting of different (and independently activated) pathways is required to efficiently induce apoptosis of MM cells. This effect was observed with MM cell lines and with primary MM cells and might have direct implications for the development of future therapeutic strategies for MM.
Prognostic Significance of BCL-2 Expression in Patients with Multiple Myeloma.
Authors: Hashmi Mehmood , Kojouri Kiarash , Yang Jian , Patel Sunil , Khan Mohammad , Sylvester-Kohrt Nancy , Kamble Rammurti , Selby George B. , Ozer Howard , Kharfan-Dabaja Mohamed A. 
Published: April 2020

Abstract
Background: multiple myeloma (MM) remains an incurable disease that accounts for approximately 10% of all hematologic cancers. Patients (pts) with MM treated with conventional chemotherapy have a median overall survival of 36–42 months. Various adverse prognostic factors have been described in MM, including elevated B2 microglobulin, hypoalbuminemia, cytogenetic abnormalities of chromosome 13 (13q14 del, monosomy 13), and elevated CRP among others. BCL-2 mediates anti-apoptotic pathways and its expression has been implicated with lack of response to certain chemotherapeutic agents and unfavorable prognosis in different human malignancies.
Objective: to evaluate the prognostic significance of BCL-2 expression in patients with MM.
Methods: adult pts who were diagnosed with MM at our institution since 1/1/90 were retrospectively analyzed. Expression of BCL-2 was determined by immunohistochemistry (IHC) staining of deparafinized bone marrow specimens. Median survival was calculated for BCL-2 (+) and BCL-2 (−) groups. Survival curves were estimated according to the Kaplan-Meier method, and were compared with the use of the log-rank test. Median survivals were also compared by using Wilcoxon-Mann-Whitney test. Two-tailed alpha level of 0.05 was considered statistically significant. SAS® software (Cary, NC) was used for statistical analysis.
Results: sixty-six patients were initially identified. Forty-four pts were excluded because bone marrow samples were not available for IHC staining. Twenty-two pts (11 males, 11 females), with a median age at the time of diagnosis of 64 years (range: 46 to 83 years) were included for this analysis. Fourteen pts (64%) had positive BCL-2 bone marrow specimens, including 4 mild, 3 moderate, and 7 strong staining, while for 8 pts (36%), bone marrow specimens did not stain for BCL-2. Twelve pts (86%) from BCL-2 (+) group and 4 pts (50%) from BCL-2 (−) group died during the period of follow-up; 8 pts are still alive at the time of this analysis, 23 to 71 months (median: 39 months) after diagnosis. The median survival among BCL-2 (+) pts was 35 months (95% CI: 24 – 69 months) and among BCL-2 (−) pts was 47 months (95% CI: 13 months - N/A). Overall survival did not differ between the two groups (p-values for log-rank and Wilcoxon-Mann-Whitney tests, 0.8256 and 0.7072, respectively).
Conclusion: although our study did not show correlation between BCL-2 expression and overall survival, median survival among BCL-2 (−) pts with MM was 34% longer than BCL-2 (+) pts. The authors recognize the limitations associated with this analysis including the retrospective nature of the study, small sample size, and exclusion of a large number of patients. This study highlights the need for prospective evaluation of the prognostic significance of BCL-2 expression among pts with MM.
Characteristics of Immunoglobulin Deficiency in Patients with Untreated Multiple Myeloma Stage II and III.
Authors: Hamel Markus , Poenisch Wolfram , Hasenclever Dirk , Gelbrich Goetz , Loeffler Markus , Ali Haifa K. Al , Lehmann Stefanie , Niederwieser Dietger , Merkle Karlheinz 
Published: April 2020

Abstract
Introduction: It is well known that multiple myeloma (MM) is often associated with immunoglobulin (Ig) deficiency and increased risk of infection. We evaluated the Ig levels in 150 patients with MM stage II and III as part of a phase III randomised study comparing the effectiveness of melphalane and prednisolone versus bendamustine and prednisolone.
Patients and methods: 150 patients at a median age of 68 years, with untreated MM stage II, n = 19, and stage III, n = 131, were evaluated. MM type was IgG, IgA, light chain, and biclonal in 106 (70,7 %), 32 (21,3 %), 11 (7,5 %), 1 (0,7 %) patients respectively. Serum Ig levels were measured in all patients prior to chemotherapy. Normal values for IgG were 7,02 – 14,6 g/l, for IgA 0,24 – 3,81 g/l, and for IgM 0,54 – 2,27 g/l. Only Ig classes other than the myeloma protein class were evaluated.
Results: Patients with IgG MM had a median serum IgA of 0,4 g/l (range 0 – 7,9), and a median IgM level of 0,2 g/l (0 – 5,6). In 106 patients with IgG MM, IgA serum levels were below, within normal, and above normal values in 36 (34 %), 69 (65,1 %), and 1 (0,9 %) patients, respectively. On the other hand, IgM levels in these patients were below, within normal, and above normal values in 94 (88,7 %), 9 (8,5 %), and 3 (2,8 %) patients, respectively. Median IgG for IgA MM patients was 3,7 g/l (range 0,1 – 11,8), and median IgM in these 32 patients was 0,2 g/l (range 0 – 1,3). IgG levels were below and within normal in 28 (87,5 %), 4 (12,5 %) patients, respectively. Similarly, IgM level were below, and within normal range in 30 (93,8 %), and 2 (6,3 %) patients with IgA MM. Lastly, median IgG in patients with light chain myeloma was 6,3 g/l (range 2,1 – 9,7), median IgA, and median IgM were 0,4 g/l (range 0,1 – 1,7), and 0,3 g/l (range 0,1 – 0,8), respectively. IgG, IgA, and IgM were below normal values in 8 (72,7 %), 3 (27,3 %), and 9 (81,8 %), respectively.
Conclusion: Most patients with untreated MM have markedly reduced serum Ig levels. The correlation of reduced Ig level to the incidence of infections is being evaluated. The effect of chemotherapy on Ig levels is currently being assessed in our phase III study.
Recombinant Human Erythropoietin (rHuEpo) as an Anti-Myeloma Immunomodulatory Agent.
Authors: Prutci-Sagiv Sari , Katz Odelia , Yeyni Lidor , Skaat Alon , Barzilay Eran , Neumann Drorit , Mittelman Moshe 
Published: April 2020

Abstract
Recombinant human erythropoietin (rHuEpo), the main growth regulator of red blood cells, has been used in clinical practice for the treatment of cancer-related anemia. Recent studies suggest that rHuEpo has additional biological effects; including our finding that rHuEpo significantly prolonged the expected survival of six end-stage multiple myeloma (MM) patients whose expected survival was &lt;6 months (Mittelman et al., 2004, EJH 72, 155–65). The course of disease in these six MM patients led us to hypothesize that Epo might have an anti-neoplastic or anti-myeloma effect. This latter observation was further supported by studies, demonstrating a T cell mediated anti-myeloma effect of rHuEpo in mice models (Mittelman et al., 2001, PNAS 98, 5181–6). The mechanisms underlying the effects of Epo on the immune system are not fully clear. Our data show the effects of rHuEpo on the T-cell as well as the B-cell lineage. We found that the ratio of CD4+ to CD8+ T cells, which is decreased in MM patients, increases to the normal range in MM patients who are receiving rHuEpo treatment. rHuEpo-treated MM patients show a significant improvement in the level of mitogen-mediated activation of mononuclear cells, as measured by the early activator marker CD69. Furthermore, mononuclear proliferation in rHuEpo-treated MM patients was increased compared with MM patients who were not on rHuEpo treatment. Elucidating the anti-neoplastic effects of Epo treatment and obtaining a better insight into the immunomodulatory features of these effects will open new avenues in the treatment of hematological malignancies such as MM and related disorders. Moreover, the better understanding of the immunomodulatory properties of Epo may also be applied in other diseases manifesting immune abnormalities and immunocompromised subjects.
Effects of A771726, the Active Metabolite of Leflunomide,on Multiple Myeloma Cells Proliferation in Vitro.
Authors: Yang Shaosong , Venkataraman Girish , Krett Nancy L. , Rosen Steve T. 
Published: April 2020

Abstract
Multiple Myeloma (MM) is a clonal B-cell malignancy characterized by the accumulation of terminally differentiated plasma cell in the bone morrow, which accounts for 10% of all hematological cancers. Despite major advance of last decade, MM remains an incurable disease in need of new therapeutic approaches. Leflunomide was originally developed as an immunosuppressive drug capable of alleviating the severity of autoimmune disease and preventing allograft and xenograft rejection and it has been FDA approved to use in the treatment of rheumatoid arthritis (RA) since 1998. It rapidly converted in gastrointestinal tract and plasma to open ring metabolite A771726. In current study, we examine the effects of A771726 to the growth of MM cell lines in vivo and show that A771726 inhibits the proliferation of four different well-characterized MM cell lines (MM.1, U266, RPMI8226 and MDR10V) in a dose- and time- dependent manner, regardless their sensitivity to the conventional chemotherapies. Cell cycle analyses further demonstrate that G1-S phase transition is blocked by A771726 in those cells. Moreover, we find that, at higher concentration, A771726 is able to induce apoptosis in at least two of the cell lines tested as measured by an increase of the cells with subG1 DNA content, an increasing in caspase activity and cleavage of caspase substrate. Previous studies suggested that A771726 exerts its anti-proliferate function mostly by inhibiting de novo pyrimidine synthesis in lymphocyte and supplementation of the cultures with exogenous uridine could prevent A771726-induced cell cycle block by restoring the intracellular UTP and CTP lever. However, we find that additions of exogenous uridine can prevent neither A771726-induced cell cycle block nor apoptosis in the MM cell lines tested, suggesting a novel mechanism. It shall be noted that pharmacologically, the highest concentration of A771726 (200uM) used in our study is readily achievable in the plasma through current drug regiment of Leflunomide in RA patients. Since its approval, Leflunomide has been proved to be an effective antirheumatic drug with mild side effects. Given the results from current study, we are tempting to suggest that Leflunomide might also provide a safe alternative in the treatment of MM. The vast knowledge gained from clinical use of Leflunomide in last six year will greatly help us greatly in testing this hypothesis in a clinical setting in a near future.