BioMed Research International: Hematology The latest articles from Hindawi © 2017 , Hindawi Limited . All rights reserved. Epigenetic Guardian: A Review of the DNA Methyltransferase DNMT3A in Acute Myeloid Leukaemia and Clonal Haematopoiesis Tue, 14 Feb 2017 00:00:00 +0000 Acute myeloid leukaemia (AML) is a haematological malignancy characterized by clonal stem cell proliferation and aberrant block in differentiation. Dysfunction of epigenetic modifiers contributes significantly to the pathogenesis of AML. One frequently mutated gene involved in epigenetic modification is DNMT3A (DNA methyltransferase-3-alpha), a DNA methyltransferase that alters gene expression by de novo methylation of cytosine bases at CpG dinucleotides. Approximately 22% of AML and 36% of cytogenetically normal AML cases carry DNMT3A mutations and around 60% of these mutations affect the R882 codon. These mutations have been associated with poor prognosis and adverse survival outcomes for AML patients. Advances in whole-exome sequencing techniques have recently identified a large number of DNMT3A mutations present in clonal cells in normal elderly individuals with no features of haematological malignancy. Categorically distinct from other preleukaemic conditions, this disorder has been termed clonal haematopoiesis of indeterminate potential (CHIP). Further insight into the mutational landscape of CHIP may illustrate the consequence of particular mutations found in DNMT3A and identify specific “founder” mutations responsible for clonal expansion that may contribute to leukaemogenesis. This review will focus on current research and understanding of DNMT3A mutations in both AML and CHIP. Sabah F. Chaudry and Timothy J. T. Chevassut Copyright © 2017 Sabah F. Chaudry and Timothy J. T. Chevassut. All rights reserved. Existence of HbF Enhancer Haplotypes at HBS1L-MYB Intergenic Region in Transfusion-Dependent Saudi β-Thalassemia Patients Thu, 09 Feb 2017 00:00:00 +0000 Background and Objectives. β-Thalassemia and sickle cell disease are genetic disorders characterized by reduced and abnormal β-globin chain production, respectively. The elevation of fetal hemoglobin (HbF) can ameliorate the severity of these disorders. In sickle cell disease patients, the HbF level elevation is associated with three quantitative trait loci (QTLs), BCL11A, HBG2 promoter, and HBS1L-MYB intergenic region. This study elucidates the existence of the variants in these three QTLs to determine their association with HbF levels of transfusion-dependent Saudi β-thalassemia patients. Materials and Methods. A total of 174 transfusion-dependent β-thalassemia patients and 164 healthy controls from Eastern Province of Saudi Arabia were genotyped for fourteen single nucleotide polymorphisms (SNPs) from the three QTL regions using TaqMan assay on real-time PCR. Results. Genotype analysis revealed that six alleles of HBS1L-MYB QTL (rs9376090C , rs9399137C , rs4895441G , rs9389269C , rs9402686A , and rs9494142C ) were predominantly associated with β-thalassemia. In addition, haplotype analysis revealed that haplotypes of HBS1L-MYB (GCCGCAC ) and HBG2 (GTT ) were also predominantly associated with β-thalassemia. Furthermore, the HBS1L-MYB region also exhibited association with the high HbF cohort. Conclusion. The stimulation of HbF gene expression may provide alternative therapies for the amelioration of the disease severity of β-thalassemia. Cyril Cyrus, Chittibabu Vatte, J. Francis Borgio, Abdullah Al-Rubaish, Shahanas Chathoth, Zaki A. Nasserullah, Sana Al Jarrash, Ahmed Sulaiman, Hatem Qutub, Hassan Alsaleem, Alhusain J. Alzahrani, Martin H. Steinberg, and Amein K. Al Ali Copyright © 2017 Cyril Cyrus et al. All rights reserved. Elevated Plasma Neutrophil Gelatinase-Associated Lipocalin Level as a Risk Factor for Anemia in Patients with Systemic Inflammation Thu, 29 Dec 2016 13:08:24 +0000 Studies on neutrophil gelatinase-associated lipocalin (NGAL) as an iron-regulatory protein are limited. This study investigated the relationships between plasma NGAL levels and indices of anemia in 187 patients with systemic inflammation. Plasma NGAL levels were significantly higher in patients with anemia versus in patients without anemia (185 ng/mL versus 98 ng/mL; P < 0.001). Serum iron levels were lower in patients with NGAL > 156 ng/mL than in those with NGAL ≤ 156 ng/mL (27.4 ± 25.3 µg/dL versus 58.1 ± 43.5 µg/dL; P < 0.001). In a receiver operating characteristic curve, the diagnostic ability of NGAL to identify anemia was superior to that of high-sensitivity C-reactive protein [0.712 (95% CI, 0.618–0.787) versus 0.649 (95% CI, 0.573–0.744); P < 0.01]. In a multivariate logistic regression analysis, the elevated NGAL level was significantly associated with the presence of anemia after adjusting for potential confounders [odds ratio, 1.30 (95% CI, 1.07–2.58); P = 0.010]. In conclusion, enhanced NGAL production may contribute to the development of anemia in patients with systemic inflammation. Jong Weon Choi, Tatsuyoshi Fujii, and Noriyoshi Fujii Copyright © 2016 Jong Weon Choi et al. All rights reserved. Clinical and Laboratory Features of Acute Porphyria: A Study of 36 Subjects in a Chinese Tertiary Referral Center Tue, 29 Nov 2016 13:23:55 +0000 Porphyria is a group of eight metabolic disorders characterized by defects in heme biosynthesis. The presentation of porphyria is highly variable, and the symptoms are nonspecific, which accounts in part for delays in establishing a diagnosis. In this study, we report the characteristics of 36 Chinese acute porphyria patients. Most of them were female (33/36), and the median age was 25.3 years (range 18–45 years). The most frequent presenting symptom was abdominal pain (32/36). Hyponatremia was the most common electrolyte abnormality (29/36), and the serum sodium concentration was significantly negatively correlated with convulsion (). Genetic testing provided a precise diagnosis of the patients. Genetic analysis of the porphobilinogen deaminase (PBGD) gene was performed for 10 subjects. Of them, 9 were found to harbor a mutation in the PBGD gene, proving a diagnosis of acute intermittent porphyria, and, in 1 case, a novel Cys209Term mutation was found. Jing Yang, Qianlong Chen, Hang Yang, Baolai Hua, Tienan Zhu, Yongqiang Zhao, Huadong Zhu, Xuezhong Yu, Li Zhang, and Zhou Zhou Copyright © 2016 Jing Yang et al. All rights reserved. Synergistic Cytotoxic Effect of L-Asparaginase Combined with Decitabine as a Demethylating Agent in Pediatric T-ALL, with Specific Epigenetic Signature Sun, 27 Nov 2016 13:37:53 +0000 T-Acute Lymphoblastic Leukemia (T-ALL) remains a subgroup of pediatric ALL, with a lower response to standard chemotherapy. Some recent studies established the fundamental role of epigenetic aberrations such as DNA hypermethylation, to influence patients’ outcome and response to chemotherapy. Moreover, L-asparaginase is an important chemotherapeutic agent for treatment of ALL and resistance to this drug has been linked to ASNS expression, which can be silenced through methylation. Therefore, we tested whether the sensitivity of T-ALL cell lines towards L-asparaginase is correlated to the epigenetic status of ASNS gene and whether the sensitivity can be modified by concurrent demethylating treatment. Hence we treated different T-ALL cell lines with L-asparaginase and correlated different responses to the treatment with ASNS expression. Then we demonstrated that the ASNS expression was dependent on the methylation status of the promoter. Finally we showed that, despite the demethylating effect on the ASNS gene expression, the combined treatment with the demethylating agent Decitabine could synergistically improve the L-asparaginase sensitivity in those T-ALL cell lines characterized by hypermethylation of the ASNS gene. In conclusion, this preclinical study identified an unexpected synergistic activity of L-asparaginase and Decitabine in the subgroup of T-ALL with low ASNS expression due to hypermethylation of the ASNS promoter, while it did not restore sensitivity in the resistant cell lines characterized by higher ASNS expression. Salvatore Serravalle, Salvatore N. Bertuccio, Annalisa Astolfi, Fraia Melchionda, and Andrea Pession Copyright © 2016 Salvatore Serravalle et al. All rights reserved. Assessment of the Number and Phenotype of Macrophages in the Human BMB Samples of CML Thu, 24 Nov 2016 13:04:07 +0000 Macrophages have emerged as a key player in tumor biology. However, their number and phenotype in human bone marrow of biopsy (BMB) samples of chronic myeloid leukemia (CML) and their association with disease progression from an initial chronic phase (CP) to accelerated phase (AP) to advanced blast phase (BP) are still unclear. BMB samples from 127 CML patients and 30 patients with iron-deficiency anemia (IDA) as control group were analyzed by immunohistochemistry. The expression levels of CD68, CD163, and CD206 in BMB samples of CML patients were significantly higher than those in the patients of control group (), and we observed that their positive expression was gradually elevated during the transformation of CML-CP to AP to BP (). However, the expressions of CD68, CD163, and CD206 in released group were downregulated and contrasted to these in control group; there exists statistical significance (). The percentage ratio of CD163 and CD206 to CD68 was pronounced to be increasing from CML-CP to AP to BP (). Hence, the higher proportion of CD68+, CD163+ and CD206+ macrophages in BMB samples can be considered a key factor for disease progression of CML patients. Targeting macrophages, especially the M2 phenotype may help in designing therapeutic strategies for CML. Jian-Xin Song, Zi-Jin Dian, Yan Wen, Fen Mei, Rui-Wei Li, and Ya-Lian Sa Copyright © 2016 Jian-Xin Song et al. All rights reserved. Identification of ITGA2B and ITGB3 Single-Nucleotide Polymorphisms and Their Influences on the Platelet Function Mon, 14 Nov 2016 08:41:35 +0000 The aim of the study was to investigate ITGA2B and ITGB3 genetic polymorphisms and to evaluate the variability in the platelet function in healthy Chinese subjects. The genetic sequence of the entire coding region of the ITGA2B and ITGB3 genes was investigated. Adenosine diphosphate-induced platelet aggregation, glycoprotein IIb/IIIa content, bleeding time, and coagulation indexes were detected. Thirteen variants in the ITGA2B locus and 29 variants in the ITGB3 locus were identified in the Chinese population. The rs1009312 and rs2015049 were associated with the mean platelet volume. The rs70940817 was significantly correlated with the prothrombin time. The rs70940817 and rs112188890 were related with the activated partial thromboplastin time, and ITGB3 rs4642 was correlated with the thrombin time and fibrinogen. The minor alleles of rs56197296 and rs5919 were associated with decreased ADP-induced platelet aggregation, and rs55827077 was related with decreased GPIIb/IIIa per platelet. The rs1009312, rs2015049, rs3760364, rs567581451, rs7208170, and rs117052258 were related with bleeding time. Further studies are needed to explore the clinical importance of ITGA2B and ITGB3 SNPs in the platelet function. Qian Xiang, Shun-Dong Ji, Zhuo Zhang, Xia Zhao, and Yi-Min Cui Copyright © 2016 Qian Xiang et al. All rights reserved. Role of Membrane Lipids in the Regulation of Erythrocytic Oxygen-Transport Function in Cardiovascular Diseases Sun, 30 Oct 2016 13:43:41 +0000 The composition and condition of membrane lipids, the morphology of erythrocytes, and hemoglobin distribution were explored with the help of laser interference microscopy (LIM) and Raman spectroscopy. It is shown that patients with cardiovascular diseases (CVD) have significant changes in the composition of their phospholipids and the fatty acids of membrane lipids. Furthermore, the microviscosity of the membranes and morphology of the erythrocytes are altered causing disordered oxygen transport by hemoglobin. Basic therapy carried out with the use of antiaggregants, statins, antianginals, beta-blockers, and calcium antagonists does not help to recover the morphofunctional properties of erythrocytes. Based on the results the authors assume that, for the relief of the ischemic crisis and further therapeutic treatment, it is necessary to include, in addition to cardiovascular disease medicines, medication that increases the ability of erythrocytes’ hemoglobin to transport oxygen to the tissues. We assume that the use of LIM and Raman spectroscopy is advisable for early diagnosis of changes in the structure and functional state of erythrocytes when cardiovascular diseases develop. Victor V. Revin, Natalia V. Gromova, Elvira S. Revina, Maria I. Martynova, Angelina I. Seikina, Nadezhda V. Revina, Oksana G. Imarova, Ilia N. Solomadin, Alexander Yu. Tychkov, and Nikolai Zhelev Copyright © 2016 Victor V. Revin et al. All rights reserved. Conditioning with Fludarabine-Busulfan versus Busulfan-Cyclophosphamide Is Associated with Lower aGVHD and Higher Survival but More Extensive and Long Standing Bone Marrow Damage Mon, 24 Oct 2016 07:30:47 +0000 Acute graft-versus-host disease (aGVHD) is a major complication of allogeneic hematopoietic stem cell transplantation (allo-HSCT) and a major cause of nonrelapse mortality after allo-HSCT. A conditioning regimen plays a pivotal role in the development of aGVHD. To provide a platform for studying aGVHD and evaluating the impact of different conditioning regimens, we established a murine aGVHD model that simulates the clinical situation and can be conditioned with Busulfan-Cyclophosphamide (Bu-Cy) and Fludarabine-Busulfan (Flu-Bu). In our study, BALB/c mice were conditioned with Bu-Cy or Flu-Bu and transplanted with bone marrow cells and splenocytes from either allogeneic (C57BL/6) or syngeneic (BALB/c) donors. The allogeneic recipients conditioned with Bu-Cy had shorter survivals (), more severe clinical manifestations, and higher hepatic and intestinal pathology scores, associated with increased INF-γ expression and diminished IL-4 expression in serum, compared to allogeneic recipients conditioned with Flu-Bu. Moreover, higher donor-derived T-cell infiltration and severely impaired B-cell development were seen in the bone marrow of mice, exhibiting aGVHD and conditioned with Flu-Bu. Our study showed that the conditioning regimen with Bu-Cy resulted in more severe aGVHD while the Flu-Bu regimen was associated with more extensive and long standing bone marrow damage. Xin He, YongBin Ye, XiaoJun Xu, Jing Wang, YuXian Huang, GuangYang Weng, MingWan Zhang, and KunYuan Guo Copyright © 2016 Xin He et al. All rights reserved. Wnt-β-Catenin Signaling Promotes the Maturation of Mast Cells Sun, 23 Oct 2016 13:53:48 +0000 Mast cells play an important role in the pathogenesis of allergic diseases. Immature mast cells migrate into peripheral tissues from the bone marrow and undergo complete maturation. Interestingly, mast cells have characteristics similar to hematopoietic stem cells (HSCs), such as self-renewal and c-kit expression. In HSCs, Wnt signaling is involved in their maintenance and differentiation. On the other hand, the relation between Wnt signaling and mast cell differentiation is poorly understood. To study whether Wnt signals play a role in the maturation of mast cells, we studied the effect of Wnt proteins on mast cell maturation of bone marrow-derived mast cells (BMMCs). The expression levels of CD81 protein and histidine decarboxylase mRNA and activity of mast cell-specific protease were all elevated in BMMCs treated with Wnt5a. In addition, Wnt5a induced the expression of Axin2 and TCF mRNA in BMMCs. These results showed that Wnt5a could promote the maturation of mast cells via the canonical Wnt signaling pathway and provide important insights into the molecular mechanisms underlying the differentiation of mast cells. Tomoko Yamaguchi, Misae Nishijima, Katsuhisa Tashiro, and Kenji Kawabata Copyright © 2016 Tomoko Yamaguchi et al. All rights reserved. Monitoring Minimal Residual Disease in the Myeloproliferative Neoplasms: Current Applications and Emerging Approaches Thu, 20 Oct 2016 16:08:04 +0000 The presence of acquired mutations within the JAK2, CALR, and MPL genes in the majority of patients with myeloproliferative neoplasms (MPN) affords the opportunity to utilise these mutations as markers of minimal residual disease (MRD). Reduction of the mutated allele burden has been reported in response to a number of therapeutic modalities including interferon, JAK inhibitors, and allogeneic stem cell transplantation; novel therapies in development will also require assessment of efficacy. Real-time quantitative PCR has been widely adopted for recurrent point mutations with assays demonstrating the specificity, sensitivity, and reproducibility required for clinical utility. More recently, approaches such as digital PCR have demonstrated comparable, if not improved, assay characteristics and are likely to play an increasing role in MRD monitoring. While next-generation sequencing is increasingly valuable as a tool for diagnosis of MPN, its role in the assessment of MRD requires further evaluation. Karl Haslam and Stephen E. Langabeer Copyright © 2016 Karl Haslam and Stephen E. Langabeer. All rights reserved. Clinical Application of Revised Laboratory Classification Criteria for Antiphospholipid Antibody Syndrome: Is the Follow-Up Interval of 12 Weeks Instead of 6 Weeks Significantly Useful? Wed, 17 Aug 2016 16:54:31 +0000 Background. According to revised classification criteria of true antiphospholipid antibody syndrome, at least one of three antiphospholipid antibodies should be present on two or more occasions at least 12 weeks apart. However, it can be inconvenient to perform follow-up tests with interval of 12 weeks. We investigated clinical application of follow-up tests with interval of 12 weeks. Method. Totals of 67, 199, and 332 patients tested positive initially for the lupus anticoagulants confirm, the anti- glycoprotein-I antibody, and the anti-cardiolipin antibody test, respectively, from Jan 2007 to Jul 2009. We investigated clinical symptoms of patients, follow-up interval, and results of each test. Results. Among patients with initial test positive, 1.5%–8.5% were subjected to follow-up tests at interval of more than 12 weeks. Among 25 patients with negative conversion in tests, patients with interval of more than 12 weeks showed clinical symptom positivity of 33.3%, which was higher than that of 12.5% with 6–12 weeks. Among 34 patients with persistent test positive, clinical symptoms positivity trended to be more evident in patients at interval of 6–12 weeks (47.4% versus 26.7%, ) than more than 12 weeks. Conclusion. Less than 10% of patients with initial test positive had follow-up tests at interval of more than 12 weeks and the patients with persistent test positive at interval of more than 12 weeks showed trends toward having lower clinical symptoms than 6–12 weeks. More research is needed focused on the evidence that follow-up test at interval of more than 12 weeks should be performed instead of 6 weeks. Sang Hyuk Park, Seongsoo Jang, Chan-Jeoung Park, and Hyun-Sook Chi Copyright © 2016 Sang Hyuk Park et al. All rights reserved. Comparison of Three Chemotherapy Regimens in Elderly Patients with Diffuse Large B Cell Lymphoma: Experience at a Single National Reference Center in Mexico Sun, 10 Jul 2016 08:59:22 +0000 Background. Although chemotherapy added to rituximab is a standard of care for diffuse large B cell lymphoma (DLBCL), treatment of patients ≥65 years of age remains controversial due to comorbidities. Methods. This is a retrospective, comparative, nonrandomized study of patients ≥65 years of age, who were diagnosed with DLBCL but not previously treated. Demographic characteristics and comorbidities were analyzed. Three rituximab-containing treatment regimens (standard RCHOP, anthracycline dose-reduced RChOP, and RCOP) were compared. Descriptive analyses were conducted. Survival was calculated with the Kaplan-Meier method, and differences were compared with the log-rank test. Results. In total, 141 patients with a median age of 73.9 years were studied. The three treatment groups had comparable demographic characteristics. The overall response was 77%, 72.5%, and 59% in groups treated with RCHOP, RChOP, and RCOP, respectively. After multivariate analysis, the factors influencing the overall survival were the presence of B symptoms, poor performance status (ECOG ≥ 3), and febrile neutropenia. Factors influencing disease-free survival were febrile neutropenia, high-intermediate and high-risk IPI scores, and treatment without anthracycline. Conclusion. A higher ORR (overall response rate) was achieved with standard RCHOP, which influenced DFS and OS, although it was not statistically significant compared with the other groups. Interventional phase 3 trials testing new molecules in patients aged 70 to 80 years and older are required to improve the prognosis within this growing population. Diana Nolasco-Medina, Nancy Reynoso-Noveron, Alejandro Mohar-Betancourt, Alejandro Aviles-Salas, Osvaldo García-Perez, and Myrna Candelaria Copyright © 2016 Diana Nolasco-Medina et al. All rights reserved. Dermoscopic Follow-Up of the Skin towards Acute Graft-versus-Host-Disease in Patients after Allogeneic Hematopoietic Stem Cell Transplantation Thu, 30 Jun 2016 12:04:34 +0000 Background. Acute graft-versus-host disease (aGVHD) involving skin is one of the most frequent complications of allogeneic hematopoietic stem cell transplantation (alloHSCT), usually diagnosed based on clinical manifestations. So far, skin biopsy with histopathological evaluation is the only method to confirm the diagnosis. Objective. In this prospective study we monitored alloHSCT recipients by dermoscopy in order to assess its utility as an alternative noninvasive tool to early diagnose acute GVHD. Methods. Thirteen consecutive patients who received alloHSCT were examined clinically and dermoscopically towards aGVHD [days 28 (±7), 56 (±7), and 100 (±7)], as well as in each patient who developed cutaneous aGVHD diagnosed according to clinical criteria (Glucksberg scale). Results. Six patients (46%) developed symptoms of cutaneous acute GVHD (grade 1, ; grade 2, ). Dermoscopic evaluation revealed pinkish or reddish background and well-visible, multiple thin telangiectasias. Conclusion. To our knowledge, this is the first report on the use of dermoscopy to evaluate skin involvement in the course of acute GVHD suggesting its role as a diagnostic tool in follow-up of GVHD, which can be also used before clinical symptoms occur. Grazyna Kaminska-Winciorek, Tomasz Czerw, Tomasz Kruzel, and Sebastian Giebel Copyright © 2016 Grazyna Kaminska-Winciorek et al. All rights reserved. Platelet Activation: The Mechanisms and Potential Biomarkers Wed, 15 Jun 2016 09:51:33 +0000 Beyond hemostasis and thrombosis, an increasing number of studies indicate that platelets play an integral role in intercellular communication, mediating inflammatory and immunomodulatory activities. Our knowledge about how platelets modulate inflammatory and immunity has greatly improved in recent years. In this review, we discuss recent advances in the pathways of platelet activation and potential application of platelet activation biomarkers to diagnosis and prediction of disease states. Seong-Hoon Yun, Eun-Hye Sim, Ri-Young Goh, Joo-In Park, and Jin-Yeong Han Copyright © 2016 Seong-Hoon Yun et al. All rights reserved. Health-Related Quality of Life in Children and Adolescents with Hereditary Bleeding Disorders and in Children and Adolescents with Stroke: Cross-Sectional Comparison to Siblings and Peers Sun, 15 May 2016 13:55:33 +0000 Objectives. To investigate self-reported health-related quality of life (HrQoL) in children and adolescents with chronic medical conditions compared with siblings/peers. Methods. Group 1 (6 treatment centers) consisted of 74 children/adolescents aged 8–16 years with hereditary bleeding disorders (HBD), 12 siblings, and 34 peers. Group 2 (one treatment center) consisted of 70 children/adolescents with stroke/transient ischemic attack, 14 siblings, and 72 peers. HrQoL was assessed with the “revised KINDer Lebensqualitätsfragebogen” (KINDL-R) questionnaire. Multivariate analyses within groups were done by one-way ANOVA and post hoc pairwise single comparisons by Student’s -tests. Adjusted pairwise comparisons were done by hierarchical linear regressions with individuals nested within treatment centers (group 1) and by linear regressions (group 2), respectively. Results. No differences were found in multivariate analyses of self-reported HrQoL in group 1, while in group 2 differences occurred in overall wellbeing and all subdimensions. These differences were due to differences between patients and peers. After adjusting for age, gender, number of siblings, and treatment center these differences persisted regarding self-worth () and friend-related wellbeing (). Conclusions. In children with HBD, HrQoL was comparable to siblings and peers. In children with stroke/TIA HrQoL was comparable to siblings while peers, independently of relevant confounder, showed better self-worth and friend-related wellbeing. Bruno Neuner, Sylvia von Mackensen, Susanne Holzhauer, Stephanie Funk, Robert Klamroth, Karin Kurnik, Anne Krümpel, Susan Halimeh, Sarah Reinke, Michael Frühwald, and Ulrike Nowak-Göttl Copyright © 2016 Bruno Neuner et al. All rights reserved. Antipsychotic Drugs Inhibit Platelet Aggregation via P2Y1 and P2Y12 Receptors Wed, 16 Mar 2016 12:18:46 +0000 Antipsychotic drugs (APDs) used to treat clinical psychotic syndromes cause a variety of blood dyscrasias. APDs suppress the aggregation of platelets; however, the underlying mechanism remains unknown. We first analyzed platelet aggregation and clot formation in platelets treated with APDs, risperidone, clozapine, or haloperidol, using an aggregometer and rotational thromboelastometry (ROTEM). Our data indicated that platelet aggregation was inhibited, that clot formation time was increased, and that clot firmness was decreased in platelets pretreated with APDs. We also examined the role two major adenosine diphosphate (ADP) receptors, P2Y1 and P2Y12, play in ADP-mediated platelet activation and APD-mediated suppression of platelet aggregation. Our results show that P2Y1 receptor stimulation with ADP-induced calcium influx was inhibited by APDs in human and rats’ platelets, as assessed by in vitro or ex vivo approach, respectively. In contrast, APDs, risperidone and clozapine, alleviated P2Y12-mediated cAMP suppression, and the release of thromboxane A2 and arachidonic acid by activated platelets decreased after APD treatment in human and rats’ platelets. Our data demonstrate that each APD tested significantly suppressed platelet aggregation via different mechanisms. Chang-Chieh Wu, Fu-Ming Tsai, Mao-Liang Chen, Semon Wu, Ming-Cheng Lee, Tzung-Chieh Tsai, Lu-Kai Wang, and Chun-Hua Wang Copyright © 2016 Chang-Chieh Wu et al. All rights reserved. Thrombin Maybe Plays an Important Role in MK Differentiation into Platelets Tue, 15 Mar 2016 13:33:24 +0000 Objectives. After development and differentiation, megakaryocytes (MKs) can produce platelets. As is well known, thrombopoietin (TPO) can induce MKs to differentiate. The effect of thrombin on MKs differentiation is not clear. In this study, we used a human megakaryoblastic leukemia cell line (Meg-01) to assess the effect of thrombin on MKs differentiation. Methods. In order to interrogate the role of thrombin in Meg-01 cells differentiation, the changes of morphology, cellular function, and expression of diverse factors were analyzed. Results. The results show that thrombin suppresses Meg-01 cells proliferation and induces apoptosis and cell cycle arrest. Thrombin upregulates the expression of CD41b, which is one of the most important MK markers. Globin transcription factor 1 (GATA-1), an important transcriptional regulator, controls MK development and maturation. The expression of GATA-1 is also upregulated by thrombin in Meg-01 cells. The expression of B-cell lymphoma 2 (Bcl-2), an apoptosis-inhibitory protein, is downregulated by thrombin. Phosphorylated protein kinase B (p-AKT) and phosphorylated extracellular signal-regulated kinase (p-ERK) were upregulated by thrombin in Meg-01 cells. All the results are consistent with Meg-01 cells treated with TPO. Discussion and Conclusion. In conclusion, all these data indicate that thrombin maybe plays an important role in MK differentiation into platelets. However, whether the platelet-like particles are certainly platelets remains unknown. Xiao-Lei Yang, Meng-Kai Ge, De-Kui Mao, Ying-Tao Lv, Shu-Yan Sun, and Ai-Ping Yu Copyright © 2016 Xiao-Lei Yang et al. All rights reserved. Biological Insights into Myeloma and Other B Cell Malignancies Tue, 16 Feb 2016 13:35:32 +0000 Mariateresa Fulciniti, Nicola Amodio, Michele Cea, Patricia Maiso, and Abdel Kareem Azab Copyright © 2016 Mariateresa Fulciniti et al. All rights reserved. E2A Antagonizes PU.1 Activity through Inhibition of DNA Binding Thu, 28 Jan 2016 15:46:35 +0000 Antagonistic interactions between transcription factors contribute to cell fate decisions made by multipotent hematopoietic progenitor cells. Concentration of the transcription factor PU.1 affects myeloid/lymphoid development with high levels of PU.1 directing myeloid cell fate acquisition at the expense of B cell differentiation. High levels of PU.1 may be required for myelopoiesis in order to overcome inhibition of its activity by transcription factors that promote B cell development. The B cell transcription factors, E2A and EBF, are necessary for commitment of multipotential progenitors and lymphoid primed multipotential progenitors to lymphocytes. In this report we hypothesized that factors required for early B cell commitment would bind to PU.1 and antagonize its ability to induce myeloid differentiation. We investigated whether E2A and/or EBF associate with PU.1. We observed that the E2A component, E47, but not EBF, directly binds to PU.1. Additionally E47 represses PU.1-dependent transactivation of the MCSFR promoter through antagonizing PU.1’s ability to bind to DNA. Exogenous E47 expression in hematopoietic cells inhibits myeloid differentiation. Our data suggest that E2A antagonism of PU.1 activity contributes to its ability to commit multipotential hematopoietic progenitors to the lymphoid lineages. Jason H. Rogers, Kristin S. Owens, Jeffrey Kurkewich, Nathan Klopfenstein, Sangeeta R. Iyer, M. Celeste Simon, and Richard Dahl Copyright © 2016 Jason H. Rogers et al. All rights reserved. Plasma Levels of Monocyte Chemotactic Protein-1 Are Associated with Clinical Features and Angiogenesis in Patients with Multiple Myeloma Wed, 27 Jan 2016 10:02:45 +0000 The aim of this pilot study was to determine the plasma levels of monocyte chemotactic protein-1 (MCP-1) and possible associations with angiogenesis and the main clinical features of untreated patients with multiple myeloma (MM). ELISA was used to determine plasma MCP-1 levels in 45 newly diagnosed MM patients and 24 healthy controls. The blood vessels were highlighted by immunohistochemical staining, and computer-assisted image analysis was used for more objective and accurate determination of two parameters of angiogenesis: microvessel density (MVD) and total vascular area (TVA). The plasma levels of MCP-1 were compared to these parameters and the presence of anemia, renal dysfunction, and bone lesions. A significant positive correlation was found between plasma MCP-1 concentrations and TVA (). The MCP-1 levels were significantly higher in MM patients with evident bone lesions (), renal dysfunction (), or anemia (). Therefore, our preliminary results found a positive association between plasma MCP-1 levels, angiogenesis (expressed as TVA), and clinical features in patients with MM. However, additional prospective studies with a respectable number of patients should be performed to authenticate these results and establish MCP-1 as a possible target of active treatment. Toni Valković, Emina Babarović, Ksenija Lučin, Sanja Štifter, Merica Aralica, Irena Seili-Bekafigo, Antica Duletić-Načinović, and Nives Jonjić Copyright © 2016 Toni Valković et al. All rights reserved. Qualitative/Chemical Analyses of Ankaferd Hemostat and Its Antioxidant Content in Synthetic Gastric Fluids Tue, 26 Jan 2016 12:40:22 +0000 Introduction. Ankaferd hemostat (ABS) is the first topical haemostatic agent involving the red blood cell-fibrinogen interactions. The antihemorrhagic efficacy of ABS has been tested in controlled clinical trials. The drug induces the formation of an encapsulated complex protein web with vital erythroid aggregation. The aim of this study is to detect the essential toxicity profile and the antioxidant molecules inside ABS. Methods. The pesticides were analyzed by GC-MS and LC-MS. The determination by ICP-MS after pressure digestion was performed for the heavy metals. HPLC was used for the detection of mycotoxins. Dioxin Response Chemically Activated Luciferase Gene Expression method was used for the dioxin evaluation. TOF-MS and spectra data were evaluated to detect the antioxidants and other molecules. Results. TOF-MS spectra revealed the presence of several antioxidant molecules (including tocotrienols, vitamin E, tryptophan, estriol, galangin, apigenin, oenin, 3,4-divanillyltetrahydrofuran, TBHQ, thymol, BHA, BHT, lycopene, glycyrrhetinic acid, and tomatine), which may have clinical implications in the pharmacobiological actions of ABS. Conclusion. The safety of ABS regarding the presence of heavy metals, pesticides, mycotoxins, GMO and dioxins, and PCBs was demonstrated. Thus the present toxicological results indicated the safety of ABS. The antioxidant content of ABS should be investigated in future studies. Ahmet Koluman, Nejat Akar, Umit Y. Malkan, and Ibrahim C. Haznedaroglu Copyright © 2016 Ahmet Koluman et al. All rights reserved. MicroRNAs: Novel Crossroads between Myeloma Cells and the Bone Marrow Microenvironment Mon, 04 Jan 2016 12:43:52 +0000 Multiple myeloma (MM) is a hematologic malignancy of differentiated plasma cells that accumulate in the bone marrow, where a complex microenvironment made by different cell types supports proliferation, survival, and drug resistance of tumor cells. MicroRNAs (miRNAs) are short non-coding RNAs that regulate gene expression at posttranscriptional level. Emerging evidence indicates that miRNAs are aberrantly expressed or functionally deregulated in MM cells as the result of multiple genetic or epigenetic mechanisms and that also the tumor microenvironment regulates MM cell functions by miRNAs. Consistently, modulation of miRNA levels in MM cells has been demonstrated to impair their functional interaction with the bone marrow microenvironment and to produce significant antitumor activity even able to overcome the protective bone marrow milieu. This review will describe the most recent findings on miRNA function in the context of MM bone marrow microenvironment, focusing on the therapeutic potential of miRNA-based approaches. Lavinia Raimondi, Angela De Luca, Eugenio Morelli, Gianluca Giavaresi, Pierosandro Tagliaferri, Pierfrancesco Tassone, and Nicola Amodio Copyright © 2016 Lavinia Raimondi et al. All rights reserved. ZNF423 and ZNF521: EBF1 Antagonists of Potential Relevance in B-Lymphoid Malignancies Wed, 16 Dec 2015 12:56:53 +0000 The development of the B-lymphoid cell lineage is tightly controlled by the concerted action of a network of transcriptional and epigenetic regulators. EBF1, a central component of this network, is essential for B-lymphoid specification and commitment as well as for the maintenance of the B-cell identity. Genetic alterations causing loss of function of these B-lymphopoiesis regulators have been implicated in the pathogenesis of B-lymphoid malignancies, with particular regard to B-cell acute lymphoblastic leukaemias (B-ALLs), where their presence is frequently detected. The activity of the B-cell regulatory network may also be disrupted by the aberrant expression of inhibitory molecules. In particular, two multi-zinc finger transcription cofactors named ZNF423 and ZNF521 have been characterised as potent inhibitors of EBF1 and are emerging as potentially relevant contributors to the development of B-cell leukaemias. Here we will briefly review the current knowledge of these factors and discuss the importance of their functional cross talk with EBF1 in the development of B-cell malignancies. Maria Mesuraca, Emanuela Chiarella, Stefania Scicchitano, Bruna Codispoti, Marco Giordano, Giovanna Nappo, Heather M. Bond, and Giovanni Morrone Copyright © 2015 Maria Mesuraca et al. All rights reserved. Deep Response in Multiple Myeloma: A Critical Review Thu, 10 Dec 2015 12:24:50 +0000 Novel and more effective treatment strategies against multiple myeloma (MM) have significantly prolonged patients’ survival and raised interest in the depth of response and its association with clinical outcome. Minimal residual disease (MRD) has emerged as one of the most relevant prognostic factors in MM and should be included in a new definition of complete response (CR). Although further standardization is still required, MRD monitoring should be applied in prospective clinical trials as a sensitive tool to compare and evaluate the efficacy of different treatment strategies, particularly in the consolidation and maintenance settings, and implement individualized therapy-monitoring approaches. Here, we review current definition of deep response in MM, advantages and limitations of current MRD assessment assays, clinical evidences for MRD monitoring as a prognostic tool for therapeutic decisions in MM, and challenges to develop uniform criteria for MRD monitoring. Mariateresa Fulciniti, Nikhil C. Munshi, and Joaquin Martinez-Lopez Copyright © 2015 Mariateresa Fulciniti et al. All rights reserved. Mechanisms of Drug Resistance in Relapse and Refractory Multiple Myeloma Mon, 16 Nov 2015 13:59:31 +0000 Multiple myeloma (MM) is a hematological malignancy that remains incurable because most patients eventually relapse or become refractory to current treatments. Although the treatments have improved, the major problem in MM is resistance to therapy. Clonal evolution of MM cells and bone marrow microenvironment changes contribute to drug resistance. Some mechanisms affect both MM cells and microenvironment, including the up- and downregulation of microRNAs and programmed death factor 1 (PD-1)/PD-L1 interaction. Here, we review the pathogenesis of MM cells and bone marrow microenvironment and highlight possible drug resistance mechanisms. We also review a potential molecular targeting treatment and immunotherapy for patients with refractory or relapse MM. Wen-Chi Yang and Sheng-Fung Lin Copyright © 2015 Wen-Chi Yang and Sheng-Fung Lin. All rights reserved. The Applicability of the International Staging System in Chinese Patients with Multiple Myeloma Receiving Bortezomib or Thalidomide-Based Regimens as Induction Therapy: A Multicenter Analysis Sun, 08 Nov 2015 07:52:00 +0000 The International Staging System (ISS) is the most important prognostic system for multiple myeloma (MM). It was identified in the era of conventional agents. The outcome of MM has significantly changed by novel agents. Thus the applicability of ISS system in the era of novel agents in Chinese patients needs to be demonstrated. We retrospectively analyzed the clinical outcomes and prognostic significance of ISS system in 1016 patients with newly diagnosed multiple myeloma in Chinese patients between 2008 and 2012, who received bortezomib- or thalidomide-based regimens as first-line therapy. The median overall survival (OS) of patients for ISS stages I/II/III was not reached/55.4 months/41.7 months (), and the median progression-free survival (PFS) was 30/29.5/25 months (), respectively. Statistically significant difference in survival was confirmed among three ISS stages in thalidomide-based group, but not between ISS stages I and II in bortezomib-based group. These findings suggest that ISS system can predict the survival in the era of novel agents in Chinese MM patients, and bortezomib may have the potential to partially overcome adverse effect of risk factors on survival, especially in higher stage of ISS system. Jing Lu, Jin Lu, Aijun Liu, Weijun Fu, Juan Du, Xiaojun Huang, Wenming Chen, and Jian Hou Copyright © 2015 Jing Lu et al. All rights reserved. Study of the Structure, Oxygen-Transporting Functions, and Ionic Composition of Erythrocytes at Vascular Diseases Tue, 27 Oct 2015 12:42:50 +0000 The present paper explores the role of erythrocytes in the pathogenesis of vascular diseases. The state of erythrocytes, their ionic composition and structure, and properties of erythrocytes hemoglobin were studied by using laser interference microscopy, Raman scattering spectroscopy, and capillary electrophoresis. In patients suffering from vascular disorders we identified statistically significant changes in the shape of erythrocytes, their ionic composition, and redistribution of hemoglobin throughout cells. Viktor V. Revin, Natalia V. Gromova, Elvira S. Revina, Natalya A. Mel’nikova, Larisa A. Balykova, Ilia N. Solomadin, Alexander Yu. Tychkov, Nadezhda V. Revina, Oksana Yu. Gromova, Irina V. Anashkina, and Viktor A. Yakushkin Copyright © 2015 Viktor V. Revin et al. All rights reserved. Mechanism of Action of Bortezomib and the New Proteasome Inhibitors on Myeloma Cells and the Bone Microenvironment: Impact on Myeloma-Induced Alterations of Bone Remodeling Thu, 22 Oct 2015 13:37:03 +0000 Multiple myeloma (MM) is characterized by a high capacity to induce alterations in the bone remodeling process. The increase in osteoclastogenesis and the suppression of osteoblast formation are both involved in the pathophysiology of the bone lesions in MM. The proteasome inhibitor (PI) bortezomib is the first drug designed and approved for the treatment of MM patients by targeting the proteasome. However, recently novel PIs have been developed to overcome bortezomib resistance. Interestingly, several preclinical data indicate that the proteasome complex is involved in both osteoclast and osteoblast formation. It is also evident that bortezomib either inhibits osteoclast differentiation induced by the receptor activator of nuclear factor kappa B (NF-κB) ligand (RANKL) or stimulates the osteoblast differentiation. Similarly, the new PIs including carfilzomib and ixazomib can inhibit bone resorption and stimulate the osteoblast differentiation. In a clinical setting, PIs restore the abnormal bone remodeling by normalizing the levels of bone turnover markers. In addition, a bone anabolic effect was described in responding MM patients treated with PIs, as demonstrated by the increase in the osteoblast number. This review summarizes the preclinical and clinical evidence on the effects of bortezomib and other new PIs on myeloma bone disease. Fabrizio Accardi, Denise Toscani, Marina Bolzoni, Benedetta Dalla Palma, Franco Aversa, and Nicola Giuliani Copyright © 2015 Fabrizio Accardi et al. All rights reserved. Mechanisms and Clinical Applications of Genome Instability in Multiple Myeloma Thu, 22 Oct 2015 13:07:55 +0000 Ongoing genomic instability represents a hallmark of multiple myeloma (MM) cells, which manifests largely as whole chromosome- or translocation-based aneuploidy. Importantly, although it supports tumorigenesis, progression and, response to treatment in MM patients, it remains one of the least understood components of malignant transformation in terms of molecular basis. Therefore these aspects make the comprehension of genomic instability a pioneering strategy for novel therapeutic and clinical speculations to use in the management of MM patients. Here we will review mechanisms mediating genomic instability in MM cells with an emphasis placed on pathogenic mutations affecting DNA recombination, replication and repair, telomere function and mitotic regulation of spindle attachment, centrosome function, and chromosomal segregation. We will discuss the mechanisms by which genetic aberrations give rise to multiple pathogenic events required for myelomagenesis and conclude with a discussion of the clinical applications of these findings in MM patients. Antonia Cagnetta, Davide Lovera, Raffaella Grasso, Nicoletta Colombo, Letizia Canepa, Filippo Ballerini, Marino Calvio, Maurizio Miglino, Marco Gobbi, Roberto Lemoli, and Michele Cea Copyright © 2015 Antonia Cagnetta et al. All rights reserved. Phylogeny and Ontogeny of Erythropoiesis Wed, 21 Oct 2015 13:21:44 +0000 Wataru Nunomura, Aurora M. Cianciarullo, Takashi Kato, Ritsuko Shimizu, and Malgorzata Witeska Copyright © 2015 Wataru Nunomura et al. All rights reserved. Biology of Heme in Mammalian Erythroid Cells and Related Disorders Sun, 18 Oct 2015 13:31:08 +0000 Heme is a prosthetic group comprising ferrous iron (Fe2+) and protoporphyrin IX and is an essential cofactor in various biological processes such as oxygen transport (hemoglobin) and storage (myoglobin) and electron transfer (respiratory cytochromes) in addition to its role as a structural component of hemoproteins. Heme biosynthesis is induced during erythroid differentiation and is coordinated with the expression of genes involved in globin formation and iron acquisition/transport. However, erythroid and nonerythroid cells exhibit distinct differences in the heme biosynthetic pathway regulation. Defects of heme biosynthesis in developing erythroblasts can have profound medical implications, as represented by sideroblastic anemia. This review will focus on the biology of heme in mammalian erythroid cells, including the heme biosynthetic pathway as well as the regulatory role of heme and human disorders that arise from defective heme synthesis. Tohru Fujiwara and Hideo Harigae Copyright © 2015 Tohru Fujiwara and Hideo Harigae. All rights reserved. Phylogenetic and Ontogenetic View of Erythroblastic Islands Sun, 18 Oct 2015 12:18:24 +0000 Erythroblastic islands are a hallmark of mammalian erythropoiesis consisting of a central macrophage surrounded by and interacting closely with the maturing erythroblasts. The macrophages are thought to serve many functions such as supporting erythroblast proliferation, supplying iron for hemoglobin, promoting enucleation, and clearing the nuclear debris; moreover, inhibition of erythroblastic island formation is often detrimental to erythropoiesis. There is still much not understood about the role that macrophages and microenvironment play in erythropoiesis and insights may be gleaned from a comparative analysis with erythropoietic niches in nonmammalian vertebrates which, unlike mammals, have erythrocytes that retain their nucleus. The phylogenetic development of erythroblastic islands in mammals in which the erythrocytes are anucleate underlines the importance of the macrophage in erythroblast enucleation. Katie M. Giger and Theodosia A. Kalfa Copyright © 2015 Katie M. Giger and Theodosia A. Kalfa. All rights reserved. Analysis of Hereditary Elliptocytosis with Decreased Binding of Eosin-5-maleimide to Red Blood Cells Sun, 18 Oct 2015 12:02:34 +0000 Flow cytometric test for analyzing the eosin-5-maleimide (EMA) binding to red blood cells has been believed to be a specific method for diagnosing hereditary spherocytosis (HS). However, it has been reported that diseases other than HS, such as hereditary pyropoikilocytosis (HPP) and Southeast Asian ovalocytosis (SAO), which are forms in the category of hereditary elliptocytosis (HE), show decreased EMA binding to red blood cells. We analyzed EMA binding to red blood cells in 101 healthy control subjects and 42 HS patients and obtained a mean channel fluorescence (MCF) cut-off value of 36.4 (sensitivity 0.97, specificity 0.95). Using this method, we also analyzed 12 HE patients. Among them, four HE patients showed the MCF at or below the cut-off value. It indicates that some HE patients have decreased EMA binding to red blood cells. Two of these four HE patients were classified as common HE, and two were spherocytic HE with reduced spectrin. This study demonstrates that, in addition to patients with HPP or SAO, some HE patients have decreased EMA binding to red blood cells. Shin-ichiro Suemori, Hideho Wada, Hidekazu Nakanishi, Takayuki Tsujioka, Takashi Sugihara, and Kaoru Tohyama Copyright © 2015 Shin-ichiro Suemori et al. All rights reserved. Physicochemical Aspects of the Plasmodium chabaudi-Infected Erythrocyte Sun, 18 Oct 2015 11:37:42 +0000 Membrane electrochemical potential is a feature of the molecular profile of the cell membrane and the two-dimensional arrangement of its charge-bearing molecules. Plasmodium species, the causative agents of malaria, are intracellular parasites that remodel host erythrocytes by expressing their own proteins on erythrocyte membranes. Although various aspects of the modifications made to the host erythrocyte membrane have been extensively studied in some human Plasmodium species (such as Plasmodium falciparum), details of the structural and molecular biological modifications made to host erythrocytes by nonhuman Plasmodium parasites have not been studied. We employed zeta potential analysis of erythrocytes parasitized by P. chabaudi, a nonhuman Plasmodium parasite. From these measurements, we found that the surface potential shift was more negative for P. chabaudi-infected erythrocytes than for P. falciparum-infected erythrocytes. However, electron microscopic analysis of the surface of P. chabaudi-infected erythrocytes did not reveal any modifications as compared with nonparasitized erythrocytes. These results suggest that differences in the membrane modifications found herein represent unique attributes related to the pathogenesis profiles of the two different malaria parasite species in different host animals and that these features have been acquired through parasite adaptations acquired over long evolutionary time periods. Eri H. Hayakawa, Seiki Kobayashi, and Hiroyuki Matsuoka Copyright © 2015 Eri H. Hayakawa et al. All rights reserved. Diverse of Erythropoiesis Responding to Hypoxia and Low Environmental Temperature in Vertebrates Sun, 18 Oct 2015 11:37:16 +0000 Erythrocytes are responsible for transporting oxygen to tissue and are essential for the survival of almost all vertebrate animals. Circulating erythrocyte counts are tightly regulated and respond to erythrocyte mass and oxygen tension. Since the discovery of erythropoietin, the erythropoietic responses to environment and tissue oxygen tension have been investigated in mice and human. Moreover, it has recently become increasingly clear that various environmental stresses could induce the erythropoiesis via various modulating systems, while all vertebrates live in various environments and habitually adapt to environmental stress. Therefore, it is considered that investigations of erythropoiesis in vertebrates provide a lead to the various erythropoietic responses to environmental stress. This paper comparatively introduces the present understanding of erythropoiesis in vertebrates. Indeed, there is a wide range of variations in vertebrates’ erythropoiesis. This paper also focused on erythropoietic responses to environmental stress, hypoxia, and lowered temperature in vertebrates. Shun Maekawa and Takashi Kato Copyright © 2015 Shun Maekawa and Takashi Kato. All rights reserved. Origins of the Vertebrate Erythro/Megakaryocytic System Sun, 18 Oct 2015 09:42:58 +0000 Vertebrate erythrocytes and thrombocytes arise from the common bipotent thrombocytic-erythroid progenitors (TEPs). Even though nonmammalian erythrocytes and thrombocytes are phenotypically very similar to each other, mammalian species have developed some key evolutionary improvements in the process of erythroid and thrombocytic differentiation, such as erythroid enucleation, megakaryocyte endoreduplication, and platelet formation. This brings up a few questions that we try to address in this review. Specifically, we describe the ontology of erythro-thrombopoiesis during adult hematopoiesis with focus on the phylogenetic origin of mammalian erythrocytes and thrombocytes (also termed platelets). Although the evolutionary relationship between mammalian and nonmammalian erythroid cells is clear, the appearance of mammalian megakaryocytes is less so. Here, we discuss recent data indicating that nonmammalian thrombocytes and megakaryocytes are homologs. Finally, we hypothesize that erythroid and thrombocytic differentiation evolved from a single ancestral lineage, which would explain the striking similarities between these cells. Ondrej Svoboda and Petr Bartunek Copyright © 2015 Ondrej Svoboda and Petr Bartunek. All rights reserved. Early Fluid Resuscitation and High Volume Hemofiltration Decrease Septic Shock Progression in Swine Mon, 12 Oct 2015 08:02:05 +0000 This study aimed to assess the effects of early fluid resuscitation (EFR) combined with high volume hemofiltration (HVHF) on the cardiopulmonary function and removal of inflammatory mediators in a septic shock swine model. Eighteen swine were randomized into three groups: control (extracorporeal circulating blood only), continuous renal replacement therapy (CRRT) (; ultrafiltration volume = 25 mL/Kg/h), and HVHF (; ultrafiltration volume = 85 mL/Kg/h). The septic shock model was established by intravenous infusion of lipopolysaccharides (50 µg/kg/h). Hemodynamic parameters (arterial pressure, heart rate, cardiac output, stroke volume variability, left ventricular contractility, systemic vascular resistance, and central venous pressure), vasoactive drug parameters (dose and time of norepinephrine and hourly fluid intake), pulmonary function (partial oxygen pressure and vascular permeability), and cytokines (interleukin-6 and interleukin-10) were observed. Treatment resulted in significant changes at 4–6 h. HVHF was beneficial, as shown by the dose of vasoactive drugs, fluid intake volume, left ventricular contractility index, and partial oxygen pressure. Both CRRT and HVHF groups showed improved removal of inflammatory mediators compared with controls. In conclusion, EFR combined with HVHF improved septic shock in this swine model. The combination decreased shock progression, reduced the need for vasoactive drugs, and alleviated the damage to cardiopulmonary functions. Ping Zhao, Ruiqiang Zheng, Lu Xue, Min Zhang, and Xiaoyan Wu Copyright © 2015 Ping Zhao et al. All rights reserved. Inhibition of P-Selectin and PSGL-1 Using Humanized Monoclonal Antibodies Increases the Sensitivity of Multiple Myeloma Cells to Bortezomib Sun, 11 Oct 2015 09:17:57 +0000 Multiple myeloma (MM) is a plasma cell malignancy localized in the bone marrow. Despite the introduction of novel therapies majority of MM patients relapse. We have previously shown that inhibition of P-selectin and P-selectin glycoprotein ligand-1 (PSGL-1) play a key role in proliferation of MM and using small-molecule inhibitors of P-selectin/PSGL-1 sensitized MM cells to therapy. However, these small-molecule inhibitors had low specificity to P-selectin and showed poor pharmacokinetics. Therefore, we tested blocking of P-selectin and PSGL-1 using functional monoclonal antibodies in order to sensitize MM cells to therapy. We have demonstrated that inhibiting the interaction between MM cells and endothelial and stromal cells decreased proliferation in MM cells and in parallel induced loose-adhesion to the primary tumor site to facilitate egress. At the same time, blocking this interaction in vivo led to MM cells retention in the circulation and delayed homing to the bone marrow, thus exposing MM cells to bortezomib which contributed to reduced tumor growth and better mice survival. This study provides a better understanding of the biology of P-selectin and PSGL-1 and their roles in dissemination and resensitization of MM to treatment. Barbara Muz, Feda Azab, Pilar de la Puente, Scott Rollins, Richard Alvarez, Ziad Kawar, and Abdel Kareem Azab Copyright © 2015 Barbara Muz et al. All rights reserved. Targeted Therapy in Hematological Malignancies: From Basic Research to Clinical Practice Tue, 08 Sep 2015 06:26:12 +0000 Haiqing Ma, Saradhi Mallampati, Gang An, and Jin Wang Copyright © 2015 Haiqing Ma et al. All rights reserved. MicroRNA181a Is Overexpressed in T-Cell Leukemia/Lymphoma and Related to Chemoresistance Mon, 07 Sep 2015 07:41:03 +0000 MicroRNAs (miRs) play an important role in tumorogenesis and chemoresistance in lymphoid malignancies. Comparing with reactive hyperplasia, miR181a was overexpressed in 130 patients with T-cell leukemia/lymphoma, including acute T-cell lymphoblastic leukemia (), T-cell lymphoblastic lymphoma (), peripheral T-cell lymphoma, not otherwise specified (), anaplastic large cell lymphoma (), and angioimmunoblastic T-cell lymphoma (). Irrespective to histological subtypes, miR181a overexpression was associated with increased AKT phosphorylation. In vitro, ectopic expression of miR181a in HEK-293T cells significantly enhanced cell proliferation, activated AKT, and conferred cell resistance to doxorubicin. Meanwhile, miR181a expression was upregulated in Jurkat cells, along with AKT activation, during exposure to chemotherapeutic agents regularly applied to T-cell leukemia/lymphoma treatment, such as doxorubicin, cyclophosphamide, cytarabine, and cisplatin. Isogenic doxorubicin-resistant Jurkat and H9 cells were subsequently developed, which also presented with miR181a overexpression and cross-resistance to cyclophosphamide and cisplatin. Meanwhile, specific inhibition of miR181a enhanced Jurkat and H9 cell sensitivity to chemotherapeutic agents, further indicating that miR181a was involved in acquired chemoresistance. Collectively, miR181a functioned as a biomarker of T-cell leukemia/lymphoma through modulation of AKT pathway. Related to tumor cell chemoresistance, miR181a could be a potential therapeutic target in treating T-cell malignancies. Zi-Xun Yan, Zhong Zheng, Wen Xue, Ming-Zhe Zhao, Xiao-Chun Fei, Li-Li Wu, Li-Min Huang, Christophe Leboeuf, Anne Janin, Li Wang, and Wei-Li Zhao Copyright © 2015 Zi-Xun Yan et al. All rights reserved. Clinical Safety and Immunogenicity of Tumor-Targeted, Plant-Made Id-KLH Conjugate Vaccines for Follicular Lymphoma Sun, 06 Sep 2015 13:54:08 +0000 We report the first evaluation of plant-made conjugate vaccines for targeted treatment of B-cell follicular lymphoma (FL) in a Phase I safety and immunogenicity clinical study. Each recombinant personalized immunogen consisted of a tumor-derived, plant-produced idiotypic antibody (Ab) hybrid comprising the hypervariable regions of the tumor-associated light and heavy Ab chains, genetically grafted onto a common human IgG1 scaffold. Each immunogen was produced in Nicotiana benthamiana plants using twin magnICON vectors expressing the light and heavy chains of the idiotypic Ab. Each purified Ab was chemically linked to the carrier protein keyhole limpet hemocyanin (KLH) to form a conjugate vaccine. The vaccines were administered to FL patients over a series of 6 subcutaneous injections in conjunction with the adjuvant Leukine (GM-CSF). The 27 patients enrolled in the study had previously received non-anti-CD20 cytoreductive therapy followed by 4 months of immune recovery prior to first vaccination. Of 11 patients who became evaluable at study conclusion, 82% (9/11) displayed a vaccine-induced, idiotype-specific cellular and/or humoral immune response. No patients showed serious adverse events (SAE) related to vaccination. The fully scalable plant-based manufacturing process yields safe and immunogenic personalized FL vaccines that can be produced within weeks of obtaining patient biopsies. Daniel Tusé, Nora Ku, Maurizio Bendandi, Carlos Becerra, Robert Collins Jr., Nyla Langford, Susana Inogés Sancho, Ascensión López-Díaz de Cerio, Fernando Pastor, Romy Kandzia, Frank Thieme, Franziska Jarczowski, Dieter Krause, Julian K.-C. Ma, Shan Pandya, Victor Klimyuk, Yuri Gleba, and John E. Butler-Ransohoff Copyright © 2015 Daniel Tusé et al. All rights reserved. Targeted Therapies in Adult B-Cell Malignancies Sun, 06 Sep 2015 13:49:54 +0000 B-lymphocytes are programmed for the production of immunoglobulin (Ig) after antigen presentation, in the context of T-lymphocyte control within lymphoid organs. During this differentiation/activation process, B-lymphocytes exhibit different restricted or common surface markers, activation of cellular pathways that regulate cell cycle, metabolism, proteasome activity, and protein synthesis. All molecules involved in these different cellular mechanisms are potent therapeutic targets. Nowadays, due to the progress of the biology, more and more targeted drugs are identified, a situation that is correlated with an extended field of the targeted therapy. The full knowledge of the cellular machinery and cell-cell communication allows making the best choice to treat patients, in the context of personalized medicine. Also, focus should not be restricted to the immediate effects observed as clinical endpoints, that is, response rate, survival markers with conventional statistical methods, but it should consider the prediction of different clinical consequences due to other collateral drug targets, based on new methodologies. This means that new reflection and new bioclinical follow-up have to be monitored, particularly with the new drugs used with success in B-cell malignancies. This review discussed the principal aspects of such evident bioclinical progress. Jean-François Rossi Copyright © 2015 Jean-François Rossi. All rights reserved. Subcutaneous Administration of Bortezomib in Combination with Thalidomide and Dexamethasone for Treatment of Newly Diagnosed Multiple Myeloma Patients Sun, 06 Sep 2015 13:35:51 +0000 Objective. To investigate the efficacy and safety of the treatment of the newly diagnosed multiple myeloma (MM) patients with the therapy of subcutaneous (subQ) administration of bortezomib and dexamethasone plus thalidomide (VTD) regimen. Methods. A total of 60 newly diagnosed MM patients were analyzed. 30 patients received improved VTD regimen (improved VTD group) with the subQ injection of bortezomib and the other 30 patients received conventional VTD regimen (VTD group).The efficacy and safety of two groups were analyzed retrospectively. Results. The overall remission (OR) after eight cycles of treatment was 73.3% in the VTD group and 76.7% in the improved VTD group (). No significant differences in time to 1-year estimate of overall survival (72% versus 75%, ) and progression-free survival (median 22 months versus 25 months; ) between two groups. The main toxicities related to therapy were leukopenia, neutropenia, thrombocytopenia, asthenia, fatigue, and renal and urinary disorders. Grade 3 and higher adverse events were significantly less common in the improved VTD group (50%) than VTD group (80%, ). Conclusions. The improved VTD regimen by changing bortezomib from intravenous administration to subcutaneous injection has noninferior efficacy to standard VTD regimen, with an improved safety profile and reduced adverse events. Shenghao Wu, Cuiping Zheng, Songyan Chen, Xiaoping Cai, Yuejian Shi, Bijing Lin, and Yuemiao Chen Copyright © 2015 Shenghao Wu et al. All rights reserved. Rituximab as Single Agent in Primary MALT Lymphoma of the Ocular Adnexa Sun, 06 Sep 2015 13:33:36 +0000 Ocular Adnexal Lymphomas are the first cause of primary ocular malignancies, and among them the most common are MALT Ocular Adnexal Lymphomas. Recently systemic immunotherapy with anti-CD20 monoclonal antibody has been investigated as first-line treatment; however, the optimal management for MALT Ocular Adnexal Lymphomas is still unknown. The present study evaluated retrospectively the outcome of seven consecutive patients with primary MALT Ocular Adnexal Lymphomas, of whom six were treated with single agent Rituximab. All patients received 6 cycles of Rituximab 375 mg/mq every 3 weeks intravenously. The overall response rate was 100%; four patients (67%) achieved a Complete Remission, and two (33%) achieved a partial response. In four patients an additional Rituximab maintenance every 2-3 months was given for two years. After a median follow-up of 29 months (range 8–34), no recurrences were observed, without of therapy- or disease-related severe adverse events. None of the patients needed additional radiotherapy or other treatments. Rituximab as a single agent is highly effective and tolerable in first-line treatment of primary MALT Ocular adnexal Lymphomas. Furthermore, durable responses are achievable with the same-agent maintenance. Rituximab can be considered the agent of choice in the management of an indolent disease in whom the “quality of life” matter is of primary importance. Ombretta Annibali, Francesca Chiodi, Chiara Sarlo, Magdalena Cortes, Francesco M. Quaranta-Leoni, Carlo Quattrocchi, Antonella Bianchi, Stefano Bonini, and Giuseppe Avvisati Copyright © 2015 Ombretta Annibali et al. All rights reserved. Relationship of Baseline Hemoglobin Level with Serum Ferritin, Postphlebotomy Hemoglobin Changes, and Phlebotomy Requirements among HFE C282Y Homozygotes Wed, 26 Aug 2015 09:46:16 +0000 Objectives. We aimed to examine whether baseline hemoglobin levels in C282Y-homozygous patients are related to the degree of serum ferritin (SF) elevation and whether patients with different baseline hemoglobin have different phlebotomy requirements. Methods. A total of 196 patients (124 males and 72 females) who had undergone therapeutic phlebotomy and had SF and both pre- and posttreatment hemoglobin values were included in the study. Results. Bivariate correlation analysis suggested that baseline SF explains approximately 6 to 7% of the variation in baseline hemoglobin. The results also showed that males who had higher (≥150 g/L) baseline hemoglobin levels had a significantly greater reduction in their posttreatment hemoglobin despite requiring fewer phlebotomies to achieve iron depletion than those who had lower (<150 g/L) baseline hemoglobin, regardless of whether baseline SF was below or above 1000 µg/L. There were no significant differences between hemoglobin subgroups regarding baseline and treatment characteristics, except for transferrin saturation between male subgroups with SF above 1000 µg/L. Similar differences were observed when females with higher (≥138 g/L) baseline hemoglobin were compared with those with lower (<138 g/L) baseline hemoglobin. Conclusion. Dividing C282Y-homozygous patients into just two subgroups according to the degree of baseline SF elevation may obscure important subgroup variations. Seyed Ali Mousavi, Faiza Mahmood, Astrid Aandahl, Teresa Risopatron Knutsen, and Abid Hussain Llohn Copyright © 2015 Seyed Ali Mousavi et al. All rights reserved. Target-Specific Oral Anticoagulants—New Approaches in the Field of Oral Anticoagulation Wed, 20 May 2015 09:03:09 +0000 Helen Mani, Jonathan Douxfils, and Jack Ansell Copyright © 2015 Helen Mani et al. All rights reserved. Non-VKA Oral Anticoagulants: Accurate Measurement of Plasma Drug Concentrations Tue, 19 May 2015 09:37:00 +0000 Non-VKA oral anticoagulants (NOACs) have now widely reached the lucrative market of anticoagulation. While the marketing authorization holders claimed that no routine monitoring is required and that these compounds can be given at fixed doses, several evidences arisen from the literature tend to demonstrate the opposite. New data suggests that an assessment of the response at the individual level could improve the benefit-risk ratio of at least dabigatran. Information regarding the association of rivaroxaban and apixaban exposure and the bleeding risk is available in the drug approval package on the FDA website. These reviews suggest that accumulation of these compounds increases the risk of experiencing a bleeding complication. Therefore, in certain patient populations such as patients with acute or chronic renal impairment or with multiple drug interactions, measurement of drug exposure may be useful to ensure an optimal treatment response. More specific circumstances such as patients experiencing a haemorrhagic or thromboembolic event during the treatment duration, patients who require urgent surgery or an invasive procedure, or patient with a suspected overdose could benefit from such a measurement. This paper aims at providing guidance on how to best estimate the intensity of anticoagulation using laboratory assays in daily practice. Jonathan Douxfils, Helen Mani, Valentine Minet, Bérangère Devalet, Bernard Chatelain, Jean-Michel Dogné, and François Mullier Copyright © 2015 Jonathan Douxfils et al. All rights reserved. Non-Transfusion-Dependent Thalassemia: A Complex Mix of Genetic Entities Yet to Be Fully Discovered Mon, 06 Apr 2015 08:33:41 +0000 Paolo Ricchi, Aldo Filosa, Aurelio Maggio, and Suthat Fucharoen Copyright © 2015 Paolo Ricchi et al. All rights reserved. Polysulfated Trehalose as a Novel Anticoagulant Agent with Dual Mode of Action Tue, 17 Mar 2015 07:51:23 +0000 Physiological hemostatic balance is a coordinated outcome of counteracting coagulation and fibrinolytic systems. An imbalance of procoagulant and anticoagulant factors may result in life threatening thromboembolism. Presently, anticoagulant administration is the first line of therapy for the treatment of these conditions and several anticoagulants have been approved, including various forms of heparin. However, the polyanionic nature and multispecificity of heparin pose several complications. Generally, the polysulfated compounds with antithrombotic potential are thought to have feasible synthetic procedures with much less bleeding, thus having favourable safety profiles. Here we report the synthesis of a novel compound, trehalose octasulfate and the assessment of its anticoagulation potential. Molecular docking of trehalose and trehalose octasulfate with antithrombin showed a specificity switch in binding affinity on sulfation, where trehalose octasulfate interacts with critical residues of AT that are either directly involved in heparin binding or in the conformational rearrangement of AT on heparin binding. An in vitro analysis of trehalose octasulfate demonstrated prolonged clotting time. Lead compound when intravenously injected in occlusion induced thrombotic rats showed remarkable reduction in the size and weight of the clot at a low dose. Delay in coagulation time was observed by analysing blood plasma isolated from rats preinjected with trehalose octasulfate. A decrease in Adenosine 5′-Diphosphate (ADP) induced platelet aggregation indicated a probable dual anticoagulant and antiplatelet mechanism of action. To summarize, this study presents trehalose octasulfate as a novel, effective, dual acting antithrombotic agent. Qudsia Rashid, Mohammad Abid, Neha Gupta, Tarun Tyagi, Mohammad Z. Ashraf, and Mohamad Aman Jairajpuri Copyright © 2015 Qudsia Rashid et al. All rights reserved. Cost-Effectiveness of Anticoagulation in Patients with Nonvalvular Atrial Fibrillation with Edoxaban Compared to Warfarin in Germany Tue, 17 Mar 2015 06:51:38 +0000 We compared the cost-utility analysis for edoxaban at both doses with that of dabigatran at both doses, rivaroxaban, and apixaban (non vitamin K antagonist oral anticoagulants, NOAC) in a German population. Data of clinical outcome events were taken from edoxaban’s ENGAGE-AF, dabigatran’s RE-LY, rivaroxaban’s ROCKET, and apixaban’s ARISTOTLE trials. The base-case analyses of a 65-year-old person with a CHADS2 score >1 gained 0.17 and 0.21 quality-adjusted life years over warfarin for 30 mg od and 60 mg od edoxaban, respectively. The incremental cost-effectiveness ratio was 50.000 and 68.000 euro per quality-adjusted life years for the higher and lower dose of edoxaban (Monte Carlo simulation). These findings were also similar to those for apixaban and more cost-effective than the other NOAC regimens. The current market costs for direct oral anticoagulants are high in relation to the quality of life gained from a German public health care insurance perspective. The willingness-to-pay threshold was lowest for 60 mg edoxaban compared to all direct oral anticoagulants and for 30 mg edoxaban compared to dabigatran and rivaroxaban. Martin Krejczy, Job Harenberg, Martin Wehling, Konrad Obermann, and Gregory Y. H. Lip Copyright © 2015 Martin Krejczy et al. All rights reserved. Comment on “Clinical Comparisons of Two Free Light Chain Assays to Immunofixation Electrophoresis for Detecting Monoclonal Gammopathy” Thu, 12 Mar 2015 14:12:16 +0000 Antony R. Parker, Oscar Berlanga, and Stephen Harding Copyright © 2015 Antony R. Parker et al. All rights reserved. Recombinant Human Thrombopoietin Treatment Promotes Hematopoiesis Recovery in Patients with Severe Aplastic Anemia Receiving Immunosuppressive Therapy Wed, 11 Mar 2015 13:40:00 +0000 Objective. To assess the effectiveness of recombinant human thrombopoietin (rhTPO) in severe aplastic anemia (SAA) patients receiving immunosuppressive therapy (IST). Methods. Eighty-eight SAA patients receiving IST from January 2007 to December 2012 were included in this retrospective analysis. Of these, 40 subjects received rhTPO treatment (15000 U, subcutaneously, three times a week). rhTPO treatment was discontinued when the platelet count returned to normal range. Hematologic response, bone marrow megakaryocyte recovery, and time to transfusion independence were compared. Results. Hematologic response was achieved in 42.5%, 62.5%, and 67.5% of patients receiving rhTPO and 22.9%, 41.6%, and 47.9% of patients not receiving rhTPO at 3, 6, and 9 months after treatment, respectively (P = 0.0665, P = 0.0579, and P = 0.0847, resp.). Subjects receiving rhTPO presented an elevated number of megakaryocytes at 3, 6, and 9 months when compared with those without treatment (P = 0.025, P = 0.021, and P = 0.011, resp.). The time to platelet and red blood cell transfusion independence was shorter in patients who received rhTPO than in those without rhTPO treatment. Overall survival rate presented no differences between the two groups. Conclusion. rhTPO could improve hematologic response and promote bone marrow recovery in SAA patients receiving IST. Huaquan Wang, Qi’e Dong, Rong Fu, Wen Qu, Erbao Ruan, Guojin Wang, Hong Liu, Yuhong Wu, Jia Song, Limin Xing, Jing Guan, Lijuan Li, and Zonghong Shao Copyright © 2015 Huaquan Wang et al. All rights reserved. Endocrine and Bone Complications in β-Thalassemia Intermedia: Current Understanding and Treatment Thu, 05 Mar 2015 13:54:14 +0000 Thalassemia intermedia (TI), also known as nontransfusion dependent thalassemia (NTDT), is a type of thalassemia where affected patients do not require lifelong regular transfusions for survival but may require occasional or even frequent transfusions in certain clinical settings and for defined periods of time. NTDT encompasses three distinct clinical forms: β-thalassemia intermedia (β-TI), Hb E/β-thalassemia, and α-thalassemia intermedia (Hb H disease). Over the past decade, our understanding of the molecular features, pathophysiology, and complications of NTDT particularly β-TI has increased tremendously but data on optimal treatment of disease and its various complications are still lacking. In this paper, we shall review a group of commonly encountered complications in β-TI, mainly endocrine and bone complications. Adlette Inati, MohammadHassan A. Noureldine, Anthony Mansour, and Hussein A. Abbas Copyright © 2015 Adlette Inati et al. All rights reserved. Incidence of Alpha-Globin Gene Defect in the Lebanese Population: A Pilot Study Thu, 05 Mar 2015 12:00:22 +0000 Background. It is well established that the Mediterranean and Arab populations are at high risk for thalassemias in general and for alpha-thalassemia in particular. Yet, reports on alpha-thalassemia in Lebanon are still lacking. In this study, we aim at assessing the incidence of alpha-thalassemia in the Lebanese population. Methods. 230 newborns’ dried blood cards remaining from routine neonatal screening at the American University of Beirut Medical Center were collected for DNA extraction. Samples were screened for the 21 most common α-globin deletions and point mutations reported worldwide, through multiplex Polymerase Chain Reaction (PCR) and Reverse-Hybridization technique. Results. Upon analyses, the carrier rate of α-thalassemia was found to be 8%. Two mutations detected the −α3,7 single gene deletion found in 75% of cases and the nongene deletion α2 IVS1 [−5nt] in the remaining samples. Conclusion. This study is the first dedicated to investigate α-thalassemia trait incidence in Lebanon. Data obtained demonstrates a high carrier rate in a relatively, highly consanguineous population; it also highlighted the presence of two common mutations. These results may be of an important impact on premarital and newborn screening policies in our country. Chantal Farra, Rose Daher, Rebecca Badra, Rym el Rafei, Rachelle Bejjany, Lama Charafeddine, and Khalid Yunis Copyright © 2015 Chantal Farra et al. All rights reserved. Triggers, Inhibitors, Mechanisms, and Significance of Eryptosis: The Suicidal Erythrocyte Death Wed, 04 Mar 2015 11:26:34 +0000 Suicidal erythrocyte death or eryptosis is characterized by erythrocyte shrinkage, cell membrane blebbing, and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Triggers of eryptosis include Ca2+ entry, ceramide formation, stimulation of caspases, calpain activation, energy depletion, oxidative stress, and dysregulation of several kinases. Eryptosis is triggered by a wide variety of xenobiotics. It is inhibited by several xenobiotics and endogenous molecules including NO and erythropoietin. The susceptibility of erythrocytes to eryptosis increases with erythrocyte age. Phosphatidylserine exposing erythrocytes adhere to the vascular wall by binding to endothelial CXC-Motiv-Chemokin-16/Scavenger-receptor for phosphatidylserine and oxidized low density lipoprotein (CXCL16). Phosphatidylserine exposing erythrocytes are further engulfed by phagocytosing cells and are thus rapidly cleared from circulating blood. Eryptosis eliminates infected or defective erythrocytes thus counteracting parasitemia in malaria and preventing detrimental hemolysis of defective cells. Excessive eryptosis, however, may lead to anemia and may interfere with microcirculation. Enhanced eryptosis contributes to the pathophysiology of several clinical disorders including metabolic syndrome and diabetes, malignancy, cardiac and renal insufficiency, hemolytic uremic syndrome, sepsis, mycoplasma infection, malaria, iron deficiency, sickle cell anemia, thalassemia, glucose 6-phosphate dehydrogenase deficiency, and Wilson’s disease. Facilitating or inhibiting eryptosis may be a therapeutic option in those disorders. Elisabeth Lang and Florian Lang Copyright © 2015 Elisabeth Lang and Florian Lang. All rights reserved. The Roles of SNF2/SWI2 Nucleosome Remodeling Enzymes in Blood Cell Differentiation and Leukemia Wed, 18 Feb 2015 06:30:49 +0000 Here, we review the role of sucrose nonfermenting (SNF2) family enzymes in blood cell development. The SNF2 family comprises helicase-like ATPases, originally discovered in yeast, that can remodel chromatin by changing chromatin structure and composition. The human genome encodes 30 different SNF2 enzymes. SNF2 family enzymes are often part of multisubunit chromatin remodeling complexes (CRCs), which consist of noncatalytic/auxiliary subunit along with the ATPase subunit. However, blood cells express a limited set of SNF2 ATPases that are necessary to maintain the pool of hematopoietic stem cells (HSCs) and drive normal blood cell development and differentiation. The composition of CRCs can be altered by the association of specific auxiliary subunits. Several auxiliary CRC subunits have specific functions in hematopoiesis. Aberrant expressions of SNF2 ATPases and/or auxiliary CRC subunit(s) are often observed in hematological malignancies. Using large-scale data from the International Cancer Genome Consortium (ICGC) we observed frequent mutations in genes encoding SNF2 helicase-like enzymes and auxiliary CRC subunits in leukemia. Hence, orderly function of SNF2 family enzymes is crucial for the execution of normal blood cell developmental program, and defects in chromatin remodeling caused by mutations or aberrant expression of these proteins may contribute to leukemogenesis. Punit Prasad, Andreas Lennartsson, and Karl Ekwall Copyright © 2015 Punit Prasad et al. All rights reserved. Crosstalk between Red Blood Cells and the Immune System and Its Impact on Atherosclerosis Wed, 04 Feb 2015 06:35:04 +0000 Atherosclerosis is a chronic multifactorial disease of the arterial wall characterized by inflammation, oxidative stress, and immune system activation. Evidence exists on a pathogenic role of oxidized red blood cells (RBCs) accumulated in the lesion after intraplaque hemorrhage. This review reports current knowledge on the impact of oxidative stress in RBC modifications with the surface appearance of senescent signals characterized by reduced expression of CD47 and glycophorin A and higher externalization of phosphatidylserine. The review summarizes findings indicating that oxidized, senescent, or stored RBCs, due to surface antigen modification and release of prooxidant and proinflammatory molecules, exert an impaired modulatory activity on innate and adaptive immune cells and how this activity contributes to atherosclerotic disease. In particular RBCs from patients with atherosclerosis, unlike those from healthy subjects, fail to control lipopolysaccharide-induced DC maturation and T lymphocyte apoptosis. Stored RBCs, accompanied by shedding of extracellular vesicles, stimulate peripheral blood mononuclear cells to release proinflammatory cytokines, augment mitogen-driven T cell proliferation, and polarize macrophages toward the proinflammatory M1 activation pathway. Collectively, literature data suggest that the crosstalk between RBCs with immune cells represents a novel mechanism by which oxidative stress can contribute to atherosclerotic disease progression and may be exploited for therapeutic interventions. Brigitta Buttari, Elisabetta Profumo, and Rachele Riganò Copyright © 2015 Brigitta Buttari et al. All rights reserved. Mechanisms Linking Red Blood Cell Disorders and Cardiovascular Diseases Sun, 01 Feb 2015 13:48:18 +0000 The present paper aims to review the main pathophysiological links between red blood cell disorders and cardiovascular diseases, provides a brief description of the latest studies in this area, and considers implications for clinical practice and therapy. Anemia is associated with a special risk in proatherosclerotic conditions and heart disease and became a new therapeutic target. Guidelines must be updated for the management of patients with red blood cell disorders and cardiovascular diseases, and targets for hemoglobin level should be established. Risk scores in several cardiovascular diseases should include red blood cell count and RDW. Complete blood count and hemorheological parameters represent useful, inexpensive, widely available tools for the management and prognosis of patients with coronary heart disease, heart failure, hypertension, arrhythmias, and stroke. Hypoxia and iron accumulation cause the most important cardiovascular effects of sickle cell disease and thalassemia. Patients with congenital chronic hemolytic anemia undergoing splenectomy should be monitored, considering thromboembolic and cardiovascular risk. Ioana Mozos Copyright © 2015 Ioana Mozos. All rights reserved. Red Blood Cell Destruction in Autoimmune Hemolytic Anemia: Role of Complement and Potential New Targets for Therapy Thu, 29 Jan 2015 09:49:00 +0000 Autoimmune hemolytic anemia (AIHA) is a collective term for several diseases characterized by autoantibody-initiated destruction of red blood cells (RBCs). Exact subclassification is essential. We provide a review of the respective types of AIHA with emphasis on mechanisms of RBC destruction, focusing in particular on complement involvement. Complement activation plays a definitive but limited role in warm-antibody AIHA (w-AIHA), whereas primary cold agglutinin disease (CAD), secondary cold agglutinin syndrome (CAS), and paroxysmal cold hemoglobinuria (PCH) are entirely complement-dependent disorders. The details of complement involvement differ among these subtypes. The theoretical background for therapeutic complement inhibition in selected patients is very strong in CAD, CAS, and PCH but more limited in w-AIHA. The optimal target complement component for inhibition is assumed to be important and highly dependent on the type of AIHA. Complement modulation is currently not an evidence-based therapy modality in any AIHA, but a number of experimental and preclinical studies are in progress and a few clinical observations have been reported. Clinical studies of new complement inhibitors are probably not far ahead. Sigbjørn Berentsen and Tatjana Sundic Copyright © 2015 Sigbjørn Berentsen and Tatjana Sundic. All rights reserved. New Insights into Malignant B-Cell Disorders Wed, 28 Jan 2015 13:56:56 +0000 Marie-Christine Kyrtsonis, Kazuyuki Shimizu, Panayiotis Panayiotidis, and Gerassimos A. Pangalis Copyright © 2015 Marie-Christine Kyrtsonis et al. All rights reserved. Targeting of Prosurvival Pathways as Therapeutic Approaches against Primary Effusion Lymphomas: Past, Present, and Future Wed, 28 Jan 2015 06:58:06 +0000 Constitutively activated prosurvival pathways render cancer cells addicted to their effects. Consequently they turn out to be the Achilles’ heels whose inhibition can be exploited in anticancer therapy. Primary effusion lymphomas (PELs) are very aggressive non-Hodgkin’s B cell lymphomas, whose pathogenesis is strictly linked to Kaposi’s sarcoma herpesvirus (KSHV) infection. Here we summarized previous studies from our and other laboratories exploring the cytotoxic effect of drugs inhibiting the main prosurvival pathways activated in PEL cells. Moreover, the immunogenicity of cell death, in terms of dendritic cell (DC) activation and their potential side effect on DCs, is discussed. Marisa Granato, Roberta Santarelli, Roberta Gonnella, Antonella Farina, Pankaj Trivedi, Alberto Faggioni, and Mara Cirone Copyright © 2015 Marisa Granato et al. All rights reserved. Iron Dextran Increases Hepatic Oxidative Stress and Alters Expression of Genes Related to Lipid Metabolism Contributing to Hyperlipidaemia in Murine Model Sun, 18 Jan 2015 13:27:55 +0000 The objective of this study was to investigate the effects of iron dextran on lipid metabolism and to determine the involvement of oxidative stress. Fischer rats were divided into two groups: the standard group (S), which was fed the AIN-93M diet, and the standard plus iron group (SI), which was fed the same diet but also received iron dextran injections. Serum cholesterol and triacylglycerol levels were higher in the SI group than in the S group. Iron dextran was associated with decreased mRNA levels of pparα, and its downstream gene cpt1a, which is involved in lipid oxidation. Iron dextran also increased mRNA levels of apoB-100, MTP, and L-FABP indicating alterations in lipid secretion. Carbonyl protein and TBARS were consistently higher in the liver of the iron-treated rats. Moreover, a significant positive correlation was found between oxidative stress products, lfabp expression, and iron stores. In addition, a negative correlation was found between pparα expression, TBARS, carbonyl protein, and iron stores. In conclusion, our results suggest that the increase observed in the transport of lipids in the bloodstream and the decreased fatty acid oxidation in rats, which was promoted by iron dextran, might be attributed to increased oxidative stress. Maísa Silva, Joyce Ferreira da Costa Guerra, Ana Flávia Santos Sampaio, Wanderson Geraldo de Lima, Marcelo Eustáquio Silva, and Maria Lucia Pedrosa Copyright © 2015 Maísa Silva et al. All rights reserved. Immunomodulatory Effects of the Agaricus blazei Murrill-Based Mushroom Extract AndoSan in Patients with Multiple Myeloma Undergoing High Dose Chemotherapy and Autologous Stem Cell Transplantation: A Randomized, Double Blinded Clinical Study Sun, 18 Jan 2015 08:38:19 +0000 Forty patients with multiple myeloma scheduled to undergo high dose chemotherapy with autologous stem cell support were randomized in a double blinded fashion to receive adjuvant treatment with the mushroom extract AndoSan, containing 82% of Agaricus blazei Murrill (19 patients) or placebo (21 patients). Intake of the study product started on the day of stem cell mobilizing chemotherapy and continued until the end of aplasia after high dose chemotherapy, a period of about seven weeks. Thirty-three patients were evaluable for all study endpoints, while all 40 included patients were evaluable for survival endpoints. In the leukapheresis product harvested after stem cell mobilisation, increased percentages of Treg cells and plasmacytoid dendritic cells were found in patients receiving AndoSan. Also, in this group, a significant increase of serum levels of IL-1ra, IL-5, and IL-7 at the end of treatment was found. Whole genome microarray showed increased expression of immunoglobulin genes, Killer Immunoglobulin Receptor (KIR) genes, and HLA genes in the Agaricus group. Furthermore, AndoSan displayed a concentration dependent antiproliferative effect on mouse myeloma cells in vitro. There were no statistically significant differences in treatment response, overall survival, and time to new treatment. The study was registered with NCT00970021. Jon-Magnus Tangen, Anne Tierens, Jo Caers, Marilene Binsfeld, Ole Kristoffer Olstad, Anne-Marie Siebke Trøseid, Junbai Wang, Geir Erland Tjønnfjord, and Geir Hetland Copyright © 2015 Jon-Magnus Tangen et al. All rights reserved. Fibrinolytic Activity and Dose-Dependent Effect of Incubating Human Blood Clots in Caffeic Acid Phenethyl Ester: In Vitro Assays Thu, 15 Jan 2015 11:04:02 +0000 Background. Caffeic acid phenethyl ester (CAPE) has been reported to possess time-dependent fibrinolytic activity by in vitro assay. This study is aimed at investigating fibrinolytic dose-dependent activity of CAPE using in vitro assays. Methods. Standardized human whole blood (WB) clots were incubated in either blank controls or different concentrations of CAPE (3.75, 7.50, 15.00, 22.50, and 30.00 mM). After 3 hours, D-dimer (DD) levels and WB clot weights were measured for each concentration. Thromboelastography (TEG) parameters were recorded following CAPE incubation, and fibrin morphology was examined under a confocal microscope. Results. Overall, mean DD (μg/mL) levels were significantly different across samples incubated with different CAPE concentrations, and the median pre- and postincubation WB clot weights (grams) were significantly decreased for each CAPE concentration. Fibrin removal was observed microscopically and indicated dose-dependent effects. Based on the TEG test, the Ly30 fibrinolytic parameter was significantly different between samples incubated with two different CAPE concentrations (15.0 and 22.50 mM). The 50% effective dose (ED50) of CAPE (based on DD) was 1.99 mg/mL. Conclusions. This study suggests that CAPE possesses fibrinolytic activity following in vitro incubation and that it has dose-dependent activities. Therefore, further investigation into CAPE as a potential alternative thrombolytic agent should be conducted. Abuzar Elnager, Rosline Hassan, Zamzuri Idris, Zulkifli Mustafa, Nadiah Wan-Arfah, S. A. Sulaiman, Siew Hua Gan, and Wan Zaidah Abdullah Copyright © 2015 Abuzar Elnager et al. All rights reserved. Phase II, Open Label, Randomized Comparative Trial of Ondansetron Alone versus the Combination of Ondansetron and Aprepitant for the Prevention of Nausea and Vomiting in Patients with Hematologic Malignancies Receiving Regimens Containing High-Dose Cytarabine Thu, 15 Jan 2015 06:42:34 +0000 Background. Aprepitant is a P/neurokinin-1 receptor antagonist approved for the prevention of CINV in moderate emetic risk chemotherapy. We explored its effectiveness in patients with leukemia receiving cytarabine-based chemotherapy. Methods. Patients were randomized to ondansetron (OND) 8 mg IV 30 minutes before cytarabine followed by 24 mg IV continuous infusion daily until 6–12 hours after the last dose of chemotherapy alone or with aprepitant (APREP) oral 125 mg 6–12 hrs before chemotherapy and 80 mg daily until 1 day after the last dose of chemotherapy. Results. Forty-nine patients were enrolled in each arm; 42 in OND and 41 in OND + APREP arm were evaluable for efficacy. The ORR with OND + APREP was 80% compared to 67% with OND alone (). On days 6 and 7, higher proportion of patients treated with OND + APREP were free from nausea (74%, 74% versus 68%, 67%; and 0.18, resp.). Requirement of rescue medications on days 2 and 3 was fewer in OND + APREP arm 7% and 5% compared to 21% and 16% in the OND arm, respectively ( and ). Conclusions. There was a trend for overall improvement in emesis with ondansetron plus aprepitant. The potential benefit of this approach with specific chemotherapy combinations remains to be determined. Talha Badar, Jorge Cortes, Gautam Borthakur, Susan O’Brien, William Wierda, Guillermo Garcia-Manero, Alessandra Ferrajoli, Tapan Kadia, Rebeca Poku, Hagop Kantarjian, and Gloria Mattiuzzi Copyright © 2015 Talha Badar et al. All rights reserved. HFE Genotyping in Patients with Elevated Serum Iron Indices and Liver Diseases Wed, 14 Jan 2015 14:17:05 +0000 Iron abnormalities in chronic liver disease may be the result of genetic diseases or secondary factors. The present study aimed to identify subjects with HFE-HH in order to describe the frequency of clinical manifestations, identify risk factors for iron elevation, and compare the iron profile of HFE-HH to other genotypes in liver disease patients. A total of 108 individuals with hepatic disease, transferrin saturation (TS) > 45%, and serum ferritin (SF) > 350 ng/mL were tested for HFE mutations. Two groups were characterized: C282Y/C282Y or C282Y/H63D genotypes () were the HFE hereditary hemochromatosis (HFE-HH) group; and C282Y and H63D single heterozygotes, the H63D/H63D genotype, and wild-type were considered group 2 (). Nonalcoholic liver disease, alcoholism, and chronic hepatitis C were detected more frequently in group 2, whereas arthropathy, hepatocarcinoma, diabetes, and osteoporosis rates were significantly higher in the HFE-HH group. TS > 82%, SF > 2685 ng/mL, and serum iron > 178 μg/dL were the cutoffs for diagnosis of HFE-HH in patients with liver disease. Thus, in non-Caucasian populations with chronic liver disease, HFE-HH diagnosis is more predictable in those with iron levels higher than those proposed in current guidelines for the general population. Andreia Silva Evangelista, Maria Cristina Nakhle, Thiago Ferreira de Araújo, Clarice Pires Abrantes-Lemos, Marta Mitiko Deguti, Flair José Carrilho, and Eduardo Luiz Rachid Cançado Copyright © 2015 Andreia Silva Evangelista et al. All rights reserved. Characterization of Red Blood Cells with Multiwavelength Transmission Spectroscopy Mon, 12 Jan 2015 14:02:09 +0000 Multiwavelength transmission (MWT) spectroscopy was applied to the investigation of the morphological parameters and composition of red blood cells (RBCs). The MWT spectra were quantitatively analyzed with a Mie theory based interpretation model modified to incorporate the effects of the nonsphericity and orientation of RBCs. The MWT spectra of the healthy and anemic samples were investigated for the RBC indices in open and blinded studies. When MWT performance was evaluated against a standard reference system, very good agreement between two methods, with for all indices studied, was demonstrated. The RBC morphological parameters were used to characterize three types of anemia and to draw an association between RBC morphology and anemia severity. The MWT spectra of RBCs infected with malaria parasite Plasmodium falciparum at different life cycle stages were analyzed for RBC morphological parameters. The changes in the RBC volume, surface area, aspect ratio, and hemoglobin composition were used to trace the morphological and compositional alterations in the infected RBCs occurring with parasites’ development and to provide insights into parasite-host interactions. The MWT method was shown to be reliable for determination of the RBC morphological parameters and to be valuable for identification of the RBC pathologic changes and disease states. Yulia M. Serebrennikova, Debra E. Huffman, and Luis H. Garcia-Rubio Copyright © 2015 Yulia M. Serebrennikova et al. All rights reserved. Incidence and Prognostic Impact of DNMT3A Mutations in Korean Normal Karyotype Acute Myeloid Leukemia Patients Sun, 11 Jan 2015 12:54:27 +0000 Background. DNA methyltransferase 3A (DNMT3A) mutation was recently introduced as a prognostic indicator in normal karyotype (NK) AML and we evaluated the incidence and prognostic impact of DNMT3A mutations in Korean NK AML patients. Methods. Total 67 NK AML patients diagnosed during the recent 10 years were enrolled. DNMT3A mutations were analyzed by direct sequencing and categorized into nonsynonymous variations (NSV), deleterious mutations (DM), and R882 mutation based on in silico analysis results. Clinical features and prognosis were compared with respect to DNMT3A mutation status. Results. Three novel (I158M, K219V, and E177V) and two known (R736H and R882H) NSVs were identified and the latter three were predicted as DMs. DNMT3A NSVs, DMs, and R882 mutation were identified in 14.9%–17.9%, 10.3%–10.4%, and 7.5% of patients, respectively. DNMT3A mutations were frequently detected in FLT3 ITD mutated patients (P = 0.054, 0.071, and 0.071 in NSV, DMs, and R882 mutation, resp.) but did not affect clinical features and prognosis significantly. Conclusions. Incidences of DNMT3A NSVs, DMs, and R882 mutation are 14.9%–17.9%, 10.3%–10.4%, and 7.5%, respectively, in Korean NK AML patients. DNMT3A mutations are associated with FLT3 ITD mutations but do not affect clinical outcome significantly in Korean NK AML patients. Sang Hyuk Park, Jae-Cheol Choi, Shine Young Kim, Jongyoun Yi, Seung Hwan Oh, In-Suk Kim, Hyung-Hoi Kim, Chulhun Ludgerus Chang, Eun Yup Lee, Moo-Kon Song, Ho-Jin Shin, and Joo Seop Chung Copyright © 2015 Sang Hyuk Park et al. All rights reserved. Plasma Cell Neoplasms: Genetics, Pathobiology, and New Therapeutic Strategies Sun, 28 Dec 2014 06:16:22 +0000 Dong Soon Lee, Wee-Joo Chng, and Kazuyuki Shimizu Copyright © 2014 Dong Soon Lee et al. All rights reserved. Iron as the Key Modulator of Hepcidin Expression in Erythroid Antibody-Mediated Hypoplasia Thu, 18 Dec 2014 00:11:23 +0000 Erythroid hypoplasia (EH) is a rare complication associated with recombinant human erythropoietin (rHuEPO) therapies, due to development of anti-rHuEPO antibodies; however, the underlying mechanisms remain poorly clarified. Our aim was to manage a rat model of antibody-mediated EH induced by rHuEPO and study the impact on iron metabolism and erythropoiesis. Wistar rats treated during 9 weeks with a high rHuEPO dose (200 IU) developed EH, as shown by anemia, reduced erythroblasts, reticulocytopenia, and plasmatic anti-rHuEPO antibodies. Serum iron was increased and associated with mRNA overexpression of hepatic hepcidin and other iron regulatory mediators and downregulation of matriptase-2; overexpression of divalent metal transporter 1 and ferroportin was observed in duodenum and liver. Decreased EPO expression was observed in kidney and liver, while EPO receptor was overexpressed in liver. Endogenous EPO levels were normal, suggesting that anti-rHuEPO antibodies blunted EPO function. Our results suggest that anti-rHuEPO antibodies inhibit erythropoiesis causing anemia. This leads to a serum iron increase, which seems to stimulate hepcidin expression despite no evidence of inflammation, thus suggesting iron as the key modulator of hepcidin synthesis. These findings might contribute to improving new therapeutic strategies against rHuEPO resistance and/or development of antibody-mediated EH in patients under rHuEPO therapy. J. C. Fernandes, P. Garrido, S. Ribeiro, P. Rocha-Pereira, E. Bronze-da-Rocha, L. Belo, E. Costa, F. Reis, and A. Santos-Silva Copyright © 2014 J. C. Fernandes et al. All rights reserved. Nek2 Is a Novel Regulator of B Cell Development and Immunological Response Mon, 17 Nov 2014 09:36:31 +0000 The serine/threonine kinase Nek2 is commonly found upregulated in a wide variety of neoplasms including diffuse large B cell lymphoma and multiple myeloma. High expression of Nek2 is implicated in the induction of chromosomal instability, promotion of cell proliferation, and drug resistance in tumor cells as well as a marker for poor clinical outcomes. Despite its well recorded involvement in chromosomal instability and neoplastic growth, little is known about the involvement of Nek2 in B cell development. Here we report the development of a transgenic mouse line with conditional expression of Nek2 in the B cell lineage and the effects it has on the development of B cells. Interestingly, we found that the overexpression of Nek2 does not induce spontaneous tumor formation within the transgenic mice up to 24 months after induction. Instead, overexpression of Nek2 in the B cell lineage affects the development of B cells by increasing the proportion of immature B cells in the bone marrow and decreasing B-1 B cells in peritoneal cavity. Furthermore, Nek2 transgenic mice develop spontaneous germinal centers and exhibit an enhanced T cell dependent immune response. Altogether, our data demonstrates a novel role for Nek2 in regulating B cell development and the immune response. Zhimin Gu, Wen Zhou, Junwei Huang, Ye Yang, Erik Wendlandt, Hongwei Xu, Xiao He, Guido Tricot, and Fenghuang Zhan Copyright © 2014 Zhimin Gu et al. All rights reserved. Long-Term Efficacy of Postpartum Intravenous Iron Therapy Thu, 06 Nov 2014 05:42:15 +0000 Background. The potential benefits of administering a dose of intravenous iron in patients with moderate postpartum anaemia rather than oral iron alone remains unproven. Aims. To determine whether a single injection of intravenous iron followed by a 6-week course of oral iron is as effective over 6 months in restoring normal haemoglobin levels and replenishing iron stores in women with moderate postpartum anaemia as a course of oral iron alone in women with mild postpartum anaemia. Materials and Methods. Retrospective two-arm cohort study in women with mild postpartum anaemia (haemoglobin 9.6–10.5 g/dL) prescribed iron daily for 6 weeks () and women with moderate postpartum anaemia (haemoglobin 8.5–9.5 g/dL), given a single 500 mg injection of intravenous iron followed by iron daily for 6 weeks (). Haemoglobin and ferritin were measured 6 months postpartum. Results. Haemoglobin returned to similar mean levels in both groups. Ferritin levels were statistically significantly higher in the intravenous + oral group ( μg/L versus  μg/L). Conclusions. Despite lower baseline haemoglobin, intravenous iron carboxymaltose was superior to oral iron alone in replenishing iron stores in moderate postpartum anaemia and may prove similarly beneficial in mild postpartum anaemia. Nadine Becuzzi, Roland Zimmermann, and Alexander Krafft Copyright © 2014 Nadine Becuzzi et al. All rights reserved. Clinical Outcome of Bortezomib Retreatment in Patients with Relapsed or Refractory Multiple Myeloma Thu, 30 Oct 2014 13:09:19 +0000 This retrospective study investigated the clinical efficacy and safety of bortezomib retreatment in patients with relapsed or refractory multiple myeloma (MM). A total of 30 patients who relapsed or progressed after 6 months since the last dose of their previous bortezomib therapy were included in this study. During the median 6 cycles (range: 2–12) of bortezomib retreatment, 10 (33.3%), 2 (6.7%), and 6 (20.0%) patients achieved complete response, very good partial response, and partial response, respectively. Grade 3 or 4 neutropenia (47.0%), thrombocytopenia (43.0%), anemia (10.0%), and peripheral sensory neuropathy (3.0%) were observed. The median time to progression, progression-free survival, and overall survival were 5.8 months (95% CI: 2.6–9.0), 5.5 months (95% CI: 4.2–6.8), and 13.4 months (95% CI: 6.1–20.7), respectively. Patients who received bortezomib retreatment 12 months from initial last therapy had a 1-year OS rate of 65.8% (95% CI: 43.5–88.1) while patients receiving retreatment after 6–12 months interval had a 1-year OS rate of 41.7% (95% CI: 13.9–69.5) (). In conclusion, this study demonstrates that retreatment with bortezomib is an effective strategy for patients with MM who relapsed at a long interval after initial bortezomib therapy. Jae-Sook Ahn, Sung-Hoon Jung, Seung-Shin Lee, Seo-Yeon Ahn, Deok-Hwan Yang, Yeo-Kyeoung Kim, Hyeoung-Joon Kim, and Je-Jung Lee Copyright © 2014 Jae-Sook Ahn et al. All rights reserved. Lenalidomide Induces Immunomodulation in Chronic Lymphocytic Leukemia and Enhances Antitumor Immune Responses Mediated by NK and CD4 T Cells Wed, 17 Sep 2014 11:59:19 +0000 Lenalidomide is an immunomodulatory drug with therapeutic activity in chronic lymphocytic leukemia (CLL). However, it has pleiotropic effects, and the mechanism of action responsible for its therapeutic activity has not been well defined yet. Herein, we show that lenalidomide treatment does not have an effect on the proliferation of leukemia cells, but it increases the proliferation of B cells from healthy donors. Lenalidomide did not exert a direct effect on the apoptosis of leukemia cells obtained from CLL patients, although it indirectly induced their apoptosis through the activation of nonmalignant immune cells. Thus, lenalidomide markedly increased the proliferation of NK and CD4 T cells. The effect of lenalidomide on NK cells was secondary to the induction of IL-2 production by CD4 T cells. Accordingly, depletion of T cells or blockade of IL-2 activity completely abrogated the proliferation of NK cells. Additionally, lenalidomide enhanced NK and NKT-like cell-mediated natural cytotoxicity against leukemia cells from CLL patients. Lenalidomide also upregulated CD20 expression on leukemia cells and, accordingly, it had a synergistic effect with rituximab on promoting antibody-dependent cell-mediated cytotoxicity against primary leukemia cells. Overall, these observations provide a support for combining lenalidomide with rituximab as a treatment in CLL. Andrea Acebes-Huerta, Leticia Huergo-Zapico, Ana Pilar Gonzalez-Rodriguez, Azahara Fernandez-Guizan, Angel R. Payer, Alejandro López-Soto, and Segundo Gonzalez Copyright © 2014 Andrea Acebes-Huerta et al. All rights reserved. New Insights into Monoclonal B-Cell Lymphocytosis Thu, 11 Sep 2014 11:19:38 +0000 Monoclonal B-cell lymphocytosis (MBL) is a premalignant condition characterized by the presence of less than 5000/μL circulating clonal B cells in otherwise healthy individuals. Three subcategories have been identified according to the immunophenotypic features: CLL-like, CD5(+) atypical, and CD5(−) MBL. CLL-like MBL is by far the most frequent and best studied category and further divided in low-count [LC] and high-count [HC] MBL, based on a cutoff value of 500/μL clonal B cells. LC-MBL typically remains stable and probably does not represent a truly premalignant condition, but rather an age-related immune senescence. On the other hand, HC-MBL is closely related to CLL-Rai0, bearing similar immunogenetic profile, and is associated with an annual risk of progression to CLL requiring therapy at a rate of 1.1%. Currently there are no reproducible factors for evaluating the risk of progression to CLL. CD5(−) MBL is characterized by an immunophenotype consistent with marginal zone origin and displays many similarities with marginal zone lymphomas (MZL), mainly the splenic MZL. The cutoff value of 5000/μL clonal B cells cannot probably be applied in CD5(−) MBL, requiring a new definition to describe those cases. Christina Kalpadakis, Gerassimos A. Pangalis, Sotirios Sachanas, Theodoros P. Vassilakopoulos, Stavroula Kyriakaki, Penelope Korkolopoulou, Efstathios Koulieris, Maria Moschogiannis, Xanthi Yiakoumis, Pantelis Tsirkinidis, Marie-Christine Kyrtsonis, Georgia Levidou, Helen A. Papadaki, Panayiotis Panayiotidis, and Maria K. Angelopoulou Copyright © 2014 Christina Kalpadakis et al. All rights reserved. Erratum to “Preventive Strategies against Bleeding due to Nonvitamin K Antagonist Oral Anticoagulants” Thu, 11 Sep 2014 06:37:46 +0000 Sarah Lessire, Anne-Sophie Dincq, Jonathan Douxfils, Bérangère Devalet, Jean-Baptiste Nicolas, Anne Spinewine, Anne-Sophie Larock, Jean-Michel Dogné, Maximilien Gourdin, and François Mullier Copyright © 2014 Sarah Lessire et al. All rights reserved. Alterations in Red Blood Cell Deformability during Storage: A Microfluidic Approach Wed, 10 Sep 2014 06:01:47 +0000 Red blood cells (RBCs) undergo extensive deformation when travelling through the microcapillaries. Deformability, the combined result of properties of the membrane-cytoskeleton complex, the surface area-to-volume ratio, and the hemoglobin content, is a critical determinant of capillary blood flow. During blood bank storage and in many pathophysiological conditions, RBC morphology changes, which has been suggested to be associated with decreased deformability and removal of RBC. While various techniques provide information on the rheological properties of stored RBCs, their clinical significance is controversial. We developed a microfluidic approach for evaluating RBC deformability in a physiologically meaningful and clinically significant manner. Unlike other techniques, our method enables a high-throughput determination of changes in deformation capacity to provide statistically significant data, while providing morphological information at the single-cell level. Our data show that, under conditions that closely mimic capillary dimensions and flow, the capacity to deform and the capacity to relax are not affected during storage in the blood bank. Our data also show that altered cell morphology by itself does not necessarily affect deformability. Judith C. A. Cluitmans, Venkatachalam Chokkalingam, Arno M. Janssen, Roland Brock, Wilhelm T. S. Huck, and Giel J. C. G. M. Bosman Copyright © 2014 Judith C. A. Cluitmans et al. All rights reserved. Timing of Peripheral Blood Stem Cell Yield: Comparison of Alternative Methods with the Classic Method for CD34+ Cell Determination Mon, 08 Sep 2014 10:52:26 +0000 Hematopoietic stem cells (HSCs), still represent a certain mystery in biology, have a unique property of dividing into equal cells and repopulating the hematopoietic tissue. This potential enables their use in transplantation treatments. The quality of the HSC grafts for transplantation is evaluated by flow cytometric determination of the CD34+ cells, which enables optimal timing of the first apheresis and the acquisition of maximal yield of the peripheral blood stem cells (PBSCs). To identify a more efficient method for evaluating CD34+ cells, we compared the following alternative methods with the reference method: hematopoietic progenitor cells (HPC) enumeration (using the Sysmex XE-2100 analyser), detection of CD133+ cells, and quantification of aldehyde dehydrogenase activity in the PBSCs. 266 aphereses (84 patients) were evaluated. In the preapheretic blood, the new methods produced data that were in agreement with the reference method. The ROC curves have shown that for the first-day apheresis target, the optimal predictive cut-off value was 0.032 cells/mL for the HPC method (sensitivity 73.4%, specificity 69.3%). HPC method exhibited a definite practical superiority as compared to other methods tested. HPC enumeration could serve as a supplementary method for the optimal timing of the first apheresis; it is simple, rapid, and cheap. I. Fatorova, M. Blaha, M. Lanska, D. Vokurkova, V. Rezacova, and P. Zak Copyright © 2014 I. Fatorova et al. All rights reserved. Management of Non-Vitamin K Antagonist Oral Anticoagulants in the Perioperative Setting Wed, 03 Sep 2014 06:22:07 +0000 The field of oral anticoagulation has evolved with the arrival of non-vitamin K antagonist oral anticoagulants (NOACs) including an anti-IIa agent (dabigatran etexilate) and anti-Xa agents (rivaroxaban and apixaban). The main specificities of these drugs are predictable pharmacokinetics and pharmacodynamics but special attention should be paid in the elderly, in case of renal dysfunction and in case of emergency. In addition, their perioperative management is challenging, especially with the absence of specific antidotes. Effectively, periods of interruption before surgery or invasive procedures depend on half-life and keeping a permanent balance between bleeding and thromboembolic risks. In addition, few data regarding the link between plasma concentrations and their effects are provided. Routine laboratory tests are altered by NOACs and quantitative measurements are not widely performed. This paper provides a review on the management of NOACs in the perioperative setting, including the estimation of the bleeding and thrombotic risk, the periods of interruption, the indication of heparin bridging, the usefulness of laboratory tests before surgery or invasive procedure, and the time of resuming. Most data are based on expert’s opinions. Anne-Sophie Dincq, Sarah Lessire, Jonathan Douxfils, Jean-Michel Dogné, Maximilien Gourdin, and François Mullier Copyright © 2014 Anne-Sophie Dincq et al. All rights reserved. Glycoprotein IIb/IIIa and P2Y12 Induction by Oligochitosan Accelerates Platelet Aggregation Thu, 28 Aug 2014 15:26:01 +0000 Platelet membrane receptor glycoprotein IIb/IIIa (gpiibiiia) is a receptor detected on platelets. Adenosine diphosphate (ADP) activates gpiibiiia and P2Y12, causing platelet aggregation and thrombus stabilization during blood loss. Chitosan biomaterials were found to promote surface induced hemostasis and were capable of activating blood coagulation cascades by enhancing platelet aggregation. Our current findings show that the activation of the gpiibiiia complex and the major ADP receptor P2Y12 is required for platelet aggregation to reach hemostasis following the adherence of various concentrations of chitosan biomaterials [7% N,O-carboxymethylchitosan (NO-CMC) with 0.45 mL collagen, 8% NO-CMC, oligochitosan (O-C), and oligochitosan 53 (O-C 53)]. We studied gpiibiiia and P2Y12 through flow cytometric analysis and western blotting techniques. The highest expression of gpiibiiia was observed with Lyostypt (74.3 ± 7.82%), followed by O-C (65.5 ± 7.17%). Lyostypt and O-C resulted in gpiibiiia expression increases of 29.2% and 13.9%, respectively, compared with blood alone. Western blot analysis revealed that only O-C 53 upregulated the expression of P2Y12 (1.12 ± 0.03-fold) compared with blood alone. Our findings suggest that the regulation of gpiibiiia and P2Y12 levels could be clinically useful to activate platelets to reach hemostasis. Further, we show that the novel oligochitosan is able to induce the increased expression of gpiibiiia and P2Y12, thus accelerating platelet aggregation in vitro. Mercy Halleluyah Periayah, Ahmad Sukari Halim, Nik Soriani Yaacob, Arman Zaharil Mat Saad, Abdul Rahim Hussein, Ahmad Hazri Abdul Rashid, and Zanariah Ujang Copyright © 2014 Mercy Halleluyah Periayah et al. All rights reserved. Clinical Features and Molecular Analysis of Hb H Disease in Taiwan Thu, 28 Aug 2014 08:28:53 +0000 Thalassemia is highly prevalent in Taiwan, but limited data are available about the association between genotypes and clinical manifestations in Taiwanese patients with Hb H disease. Here, we studied α-globin gene abnormalities and clinical features in Taiwanese patients with Hb H disease. Of the 90 patients, sixty-four (71.1%) were deletional and twenty-six (28.9%) were nondeletional Hb H disease. The () type of -thalassemia mutation was detected in the majority of patients (>95%). The most common genotype was (), followed by (). After further investigation of the genotype-phenotype correlation in 68 patients, we found that patients with nondeletional Hb H disease had more severe clinical features than those with deletional Hb H disease, including younger age at diagnosis, more requirement of blood transfusions, and larger proportion of patients with splenomegaly, hepatomegaly or jaundice. This is probably a consequence of the lower hemoglobin levels and the higher Hb H levels. The clinical severity was highly variable even among patients with an identical genotype, and the diversity was much more profound among patients with () genotype. Therefore, predicting the phenotype directly from the genotype in Hb H disease remains relatively difficult in Taiwan. Yu-Hua Chao, Kang-Hsi Wu, Han-Ping Wu, Su-Ching Liu, Ching-Tien Peng, and Maw-Sheng Lee Copyright © 2014 Yu-Hua Chao et al. All rights reserved. Peripheral Blood WT1 Expression Predicts Relapse in AML Patients Undergoing Allogeneic Stem Cell Transplantation Sun, 17 Aug 2014 12:10:14 +0000 To evaluate if WT1 expression may predict relapse after allo-SCT, we analyzed WT1 levels on peripheral blood (PB) and bone marrow (BM) before and after allo-SCT in 24 AML patients with WT1 overexpression at diagnosis. Five copies of WT1/ABL × 104 from PB were identified as the threshold value that correlated with relapse after allo-SCT. The same correlation was not identified when WT1 expression was assessed from bone marrow (BM). Eight out of 11 (73%) patients with a pre-allo-SCT PB-WT1 ≥ 5 and 4/13 (31%) patients with a pre-allo-SCT PB-WT1 < 5 relapsed, respectively (P = 0.04). The incidence of relapse was higher in patients with PB-WT1 ≥ 5 measured after allo-SCT, at the 3rd (56% versus 38%; P = 0.43) and at the 6th month (71% versus 20%; P = 0.03). Patients with pretransplant PB-WT1 < 5 had significantly better 2-year OS and LFS than patients with a PB-WT1 ≥ 5 (81% versus 0% and 63% versus 20%) (P = 0.02). Our data suggest the usefulness of WT1 monitoring from PB to predict the relapse in allotransplanted AML patients and to modulate the intensity of conditioning and/or the posttransplant immunosuppression in an attempt to reduce the posttransplant relapse risk. Michele Malagola, Cristina Skert, Giuseppina Ruggeri, Alessandro Turra, Rossella Ribolla, Valeria Cancelli, Federica Cattina, Elisa Alghisi, Simona Bernardi, Simone Perucca, Andrea Di Palma, Erika Borlenghi, Chiara Pagani, Giuseppe Rossi, Luigi Caimi, and Domenico Russo Copyright © 2014 Michele Malagola et al. All rights reserved. New Insights in the Mobilization of Hematopoietic Stem Cells in Lymphoma and Multiple Myeloma Patients Thu, 14 Aug 2014 12:10:51 +0000 Following chemotherapy and/or the administration of growth factors, such as granulocyte-colony stimulated factor (G-CSF), hematopoietic stem cells (HSC) mobilize from bone marrow to peripheral blood. This review aims to systematically present the structure of the HSC “niche” and elucidate the mechanisms of their mobilization. However, this field is constantly evolving and new pathways and molecules have been shown to contribute to the mobilization process. Understanding the importance and the possible primary pathophysiologic role of each pathway is rather difficult, since they share various overlapping components. The primary initiating event for the mobilization of HSC is chemotherapy-induced endogenous G-CSF production or exogenous G-CSF administration. G-CSF induces proliferation and expansion of the myelomonocytic series, which leads to proteolytic enzyme activation. These enzymes result in disruption of various receptor-ligand bonds, which leads to the disanchorage of HSC from the bone marrow stroma. In everyday clinical practice, CXC chemokine receptor-4 (CXCR4) antagonists are now being used as mobilization agents in order to improve HSC collection. Furthermore, based on the proposed mechanisms of HSC mobilization, novel mobilizing agents have been developed and are currently evaluated in preclinical and clinical studies. Maria K. Angelopoulou, Pantelis Tsirkinidis, Georgios Boutsikas, Theodoros P. Vassilakopoulos, and Panayiotis Tsirigotis Copyright © 2014 Maria K. Angelopoulou et al. All rights reserved. Clonotypic Analysis of Immunoglobulin Heavy Chain Sequences in Patients with Waldenström’s Macroglobulinemia: Correlation with MYD88 L265P Somatic Mutation Status, Clinical Features, and Outcome Thu, 14 Aug 2014 00:00:00 +0000 We performed IGH clonotypic sequence analysis in WM in order to determine whether a preferential IGH gene rearrangement was observed and to assess IGHV mutational status in blood and/or bone marrow samples from 36 WM patients. In addition we investigated the presence of MYD88 L265P somatic mutation. After IGH VDJ locus amplification, monoclonal VDJ rearranged fragments were sequenced and analyzed. MYD88 L265P mutation was detected by AS-PCR. The most frequent family usage was IGHV3 (74%); IGHV3-23 and IGHV3-74 segments were used in 26% and 17%, respectively. Somatic hypermutation was seen in 91% of cases. MYD88 L265P mutation was found in 65,5% of patients and absent in the 3 unmutated. These findings did not correlate with clinical findings and outcome. Conclusion. IGH genes’ repertoire differed in WM from those observed in other B-cell disorders with a recurrent IGHV3-23 and IGHV3-74 usage; monoclonal IGHV was mutated in most cases, and a high but not omnipresent prevalence of MYD88 L265P mutation was observed. In addition, the identification of 3 patients with unmutated IGHV gene segments, negative for the MYD88 L265P mutation, could support the hypothesis that an extra-germinal B-cell may represent the originating malignant cell in this minority of WM patients. Loizos Petrikkos, Marie-Christine Kyrtsonis, Maria Roumelioti, George Georgiou, Anna Efthymiou, Tatiana Tzenou, and Panayiotis Panayiotidis Copyright © 2014 Loizos Petrikkos et al. All rights reserved. Establishment of Cell Lines from Both Myeloma Bone Marrow and Plasmacytoma: SNU_MM1393_BM and SNU_MM1393_SC from a Single Patient Tue, 12 Aug 2014 11:49:30 +0000 Purpose. We tried to establish clinically relevant human myeloma cell lines that can contribute to the understanding of multiple myeloma (MM). Materials and Methods. Mononuclear cells obtained from MM patient’s bone marrow were injected via tail vein in an NRG/SCID mouse. Fourteen weeks after the injection, tumor developed at subcutis of the mouse. The engraftment of MM cells into mouse bone marrow (BM) was also observed. We separated and cultured cells from subcutis and BM. Results. After the separation and culture of cells from subcutis and BM, we established two cell lines originating from a single patient (SNU_MM1393_BM and SNU_MM1393_SC). Karyotype of the two newly established MM cell lines showed tetraploidy which is different from the karyotype of the patient (diploidy) indicating clonal evolution. In contrast to SNU_MM1393_BM, cell proliferation of SNU_MM1393_SC was IL-6 independent. SNU_MM1393_BM and SNU_MM1393_SC showed high degree of resistance against bortezomib compared to U266 cell line. SNU_MM1393_BM had the greater lethality compared to SNU_MM1393_SC. Conclusion. Two cell lines harboring different site tropisms established from a single patient showed differences in cytokine response and lethality. Our newly established cell lines could be used as a tool to understand the biology of multiple myeloma. Youngil Koh, Woo-June Jung, Kwang-Sung Ahn, and Sung-Soo Yoon Copyright © 2014 Youngil Koh et al. All rights reserved. Serum Soluble TACI, a BLyS Receptor, Is a Powerful Prognostic Marker of Outcome in Chronic Lymphocytic Leukemia Wed, 06 Aug 2014 00:00:00 +0000 BLyS is involved in CLL biology and its low soluble serum levels related to a shorter time to first treatment (TFT). TACI is a BLyS receptor and can be shed from cells’ surface and circulate in soluble form (sTACI). We investigated the impact of serum BLyS and sTACI levels at diagnosis in CLL patients and their relationship with disease parameters and patients’ outcome. Serum BLyS was determined in 73 patients, while sTACI in 60. Frozen sera drawn at diagnosis were tested by ELISA. sTACI concentrations correlated with BLyS (), b2-microglobulin (), anemia (), thrombocytopenia (), Binet stage (), and free light chains ratio (). Soluble BLyS levels below median and sTACI values above median were related to shorter TFT ( and 0.007). During a ten-year followup, sTACI levels, but not BLyS, correlated with survival (). In conclusion, we confirmed the prognostic significance of soluble BLyS levels with regard to TFT in CLL patients, and, more importantly, we showed for the first time that sTACI is a powerful prognostic marker, related to parameters of disease activity and staging and, more importantly, to TFT and OS. Marie-Christine Kyrtsonis, Katerina Sarris, Efstathios Koulieris, Dimitrios Maltezas, Eftychia Nikolaou, Maria K. Angelopoulou, Vassiliki Bartzis, Tatiana Tzenou, Maria Dimou, Mariana P. Siakandaris, Nora A. Viniou, Sotirios Sachanas, Christina Kalpadakis, Petros P. Sfikakis, Gerassimos A. Pangalis, and Panayiotis Panayiotidis Copyright © 2014 Marie-Christine Kyrtsonis et al. All rights reserved. PARP1-Driven Apoptosis in Chronic Lymphocytic Leukemia Sun, 03 Aug 2014 08:25:27 +0000 Chronic lymphocytic leukemia (CLL) is considered a malignancy resulting from defects in apoptosis. For this reason, targeting apoptotic pathways in CLL may be valuable for its management. Poly [ADP-ribose] polymerase 1 (PARP1) is the main member of a family of nuclear enzymes that act as DNA damage sensors. Through binding on DNA damaged structures, PARP1 recruits repair enzymes and serves as a survival factor, but if the damage is severe enough, its action may lead the cell to apoptosis through caspase activation, or necrosis. We measured the PARP1 mRNA and protein pretreatment levels in 26 patients with CLL and the corresponding posttreatment levels in 15 patients after 3 cycles of immunochemotherapy, as well as in 15 healthy blood donors. No difference was found between the pre- and posttreatment levels of PARP1, but we found a statistically significant relative increase of the 89 kDa fragment of PARP1 that is cleaved by caspases in the posttreatment samples, indicating PARP1-related apoptosis in CLL patients after treatment. Our findings constitute an important step in the field, especially in the era of PARP1 inhibitors, and may serve as a base for future clinical trials with these agents in CLL. Panagiotis T. Diamantopoulos, Maria Sofotasiou, Vasiliki Papadopoulou, Katerina Polonyfi, Theodoros Iliakis, and Nora-Athina Viniou Copyright © 2014 Panagiotis T. Diamantopoulos et al. All rights reserved. Newborn Screening for Sickle Cell Disease: Technical and Legal Aspects of a German Pilot Study with 38,220 Participants Wed, 23 Jul 2014 09:50:11 +0000 Sickle cell disease (SCD) does not occur in the indigenous German population, but with the increasing number of immigrants from countries at high risk for hemoglobinopathies, the question emerges whether or not a newborn screening program (NBS) for SCD disease should be initiated in Germany anyhow. We have recently shown that in Berlin, a city with a very large immigrant population, the incidence of SCD is considerable, but our findings are insufficient to make a decision for the country as a whole. In this paper we will show that a large body of epidemiological data can be generated in a relatively short period of time, with a very high degree of precision and at relatively little expense—a result that might motivate other working groups to start such a pilot project locally. We examined previously collected dried blood cards that were up to six months old, using high performance liquid chromatography (HPLC) as first method and capillary electrophoresis (CE) as second method. A single, part-time laboratory technician processed 38,220 samples in a period of 162 working days. The total costs per sample including all incidentals (as well as labor costs) were EUR 1.44. Claudia Frömmel, Annemarie Brose, Jeannette Klein, Oliver Blankenstein, and Stephan Lobitz Copyright © 2014 Claudia Frömmel et al. All rights reserved. Management of the Bleeding Patient Receiving New Oral Anticoagulants: A Role for Prothrombin Complex Concentrates Sun, 20 Jul 2014 11:53:29 +0000 Ease of dosing and simplicity of monitoring make new oral anticoagulants an attractive therapy in a growing range of clinical conditions. However, newer oral anticoagulants interact with the coagulation cascade in different ways than traditional warfarin therapy. Replacement of clotting factors will not reverse the effects of dabigatran, rivaroxaban, or apixaban. Currently, antidotes for these drugs are not widely available. Fortunately, withholding the anticoagulant and dialysis are freqnently effective treatments, particularly with rivaroxaban and dabigatran. Emergent bleeding, however, requires utilization of Prothrombin Complex Concentrates (PCCs). PCCs, in addition to recombinant factor VIIa, are used to activate the clotting system to reverse the effects of the new oral anticoagulants. In cases of refractory or emergent bleeding, the recommended factor concentrate in our protocols differs between the new oral anticoagulants. In patients taking dabigatran, we administer an activated PCC (aPCC) [FELBA] due to reported benefit in human in vitro studies. Based on human clinical trial evidence, the 4-factor PCC (Kcentra) is suggested for patients with refractory rivaroxaban- or apixaban-associated hemorrhage. If bleeding continues, recombinant factor VIIa may be employed. With all of these new procoagulant agents, the risk of thrombosis associated with administration of factor concentrates must be weighed against the relative risk of hemorrhage. Lisa M. Baumann Kreuziger, Joseph C. Keenan, Colleen T. Morton, and David J. Dries Copyright © 2014 Lisa M. Baumann Kreuziger et al. All rights reserved. Targeted Therapy for HM1.24 (CD317) on Multiple Myeloma Cells Thu, 17 Jul 2014 07:26:18 +0000 Multiple myeloma (MM) still remains an incurable disease, at least because of the existence of cell-adhesion mediated drug-resistant MM cells and/or continuous recruitment of presumed MM cancer stem cell-like cells (CSCs). As a new alternative treatment modality, immunological approaches using monoclonal antibodies (mAbs) and/or cytotoxic T lymphocytes (CTLs) are now attracting much attention as a novel strategy attacking MM cells. We have identified that HM1.24 [also known as bone marrow stromal cell antigen 2 (BST2) or CD317] is overexpressed on not only mature MM cells but also MM CSCs. We then have developed a humanized mAb to HM1.24 and defucosylated version of the mAb to adapt to clinical practice. Moreover, we have successfully induced HM1.24-specific CTLs against MM cells. The combination of these innovative therapeutic modalities may likely exert an anti-MM activity by evading the drug resistance mechanism and eliminating presumed CSCs in MM. Takeshi Harada and Shuji Ozaki Copyright © 2014 Takeshi Harada and Shuji Ozaki. All rights reserved. Dabigatran in Secondary Stroke Prevention: Clinical Experience with 106 Patients Tue, 15 Jul 2014 12:06:13 +0000 Introduction. Our aim was to analyze our clinical experience with dabigatran etexilate in secondary stroke prevention. Methods. We retrospectively included patients starting dabigatran etexilate for secondary stroke prevention from March 2010 to December 2012. Efficacy and safety variables were registered. Results. 106 patients were included, median follow-up of 12 months (range 1–31). Fifty-six females (52.8%), mean age 76.4 (range 50–95, SD 9.8), median CHADS2 4 (range 2–6), CHA2DS2-VASc 5 (range 2–9), and HAS-BLED 2 (range 1–5). Indication for dabigatran etexilate was ischemic stroke in 101 patients and acute cerebral hemorrhage (CH) due to warfarin in 5 (4.7%). Dabigatran etexilate 110 mg bid was prescribed in 71 cases (67%) and 150 mg bid was prescribed in the remaining. Seventeen patients (16%) suffered 20 complications during follow-up. Ischemic complications (10) were 6 transient ischemic attacks (TIA), 3 ischemic strokes, and 1 acute coronary syndrome. Hemorrhagic complications (10) were CH (1), gastrointestinal bleeding (6), mild hematuria (2), and mild metrorrhagia (1), leading to dabigatran etexilate discontinuation in 3 patients. Patients with previous CH remained uneventful. Three patients died (pneumonia, congestive heart failure, and acute cholecystitis) and 9 were lost during follow-up. Conclusions. Dabigatran etexilate was safe and effective in secondary stroke prevention in clinical practice, including a small number of patients with previous history of CH. Alicia DeFelipe-Mimbrera, Araceli Alonso Cánovas, Marta Guillán, Consuelo Matute, Susana Sainz de la Maza, Antonio Cruz, Rocío Vera, and Jaime Masjuan Copyright © 2014 Alicia DeFelipe-Mimbrera et al. All rights reserved. Comparison of the Freiburg and Charlson Comorbidity Indices in Predicting Overall Survival in Elderly Patients with Newly Diagnosed Multiple Myeloma Thu, 10 Jul 2014 08:36:36 +0000 Multiple myeloma occurs primarily in elderly patients. Considering the high prevalence of comorbidities, comorbidity is an important issue for the management of myeloma. However, the impact of comorbidity on clinical outcomes has not been fully investigated. We retrospectively analyzed patients with newly diagnosed myeloma. Comorbidities were assessed based on the Charlson comorbidity index (CCI) and the Freiburg comorbidity index (FCI). The CCI is a summary measure of 19 comorbid conditions. FCI is determined by performance status, renal impairment, and lung disease. This study included 127 patients with a median age of 71 years. Approximately half of the patients had additional disorders at the time of diagnosis, and diabetes mellitus was the most frequent diagnosis (18.9%). The most significant factors for prognosis among patient-related conditions were a history of solid cancer and performance status (ECOG ≥ 2). The FCI score was divided into 3 groups (0, 1, and 2-3), and the CCI score was divided into 2 groups (2-3 and ≥4). FCI was a strong prognostic tool for OS () and predicted clinical outcome better than CCI (). In conclusion, FCI was more useful than CCI in predicting overall survival in elderly patients with myeloma. Sung Min Kim, Moon Jin Kim, Hyun Ae Jung, Kihyun Kim, Seok Jin Kim, Jun Ho Jang, Won Seog Kim, and Chul Won Jung Copyright © 2014 Sung Min Kim et al. All rights reserved. Novel Approach to Reactive Oxygen Species in Nontransfusion-Dependent Thalassemia Wed, 09 Jul 2014 11:13:21 +0000 The term Nontransfusion dependent thalassaemia (NTDT) was suggested to describe patients who had clinical manifestations that are too severe to be termed minor yet too mild to be termed major. Those patients are not entirely dependent on transfusions for survival. If left untreated, three main factors are responsible for the clinical sequelae of NTDT: ineffective erythropoiesis, chronic hemolytic anemia, and iron overload. Reactive oxygen species (ROS) generation in NTDT patients is caused by 2 major mechanisms. The first one is chronic hypoxia resulting from chronic anemia and ineffective erythropoiesis leading to mitochondrial damage and the second is iron overload also due to chronic anemia and tissue hypoxia leading to increase intestinal iron absorption in thalassemic patients. Oxidative damage by reactive oxygen species (generated by free globin chains and labile plasma iron) is believed to be one of the main contributors to cell injury, tissue damage, and hypercoagulability in patients with thalassemia. Independently increased ROS has been linked to a myriad of pathological outcomes such as leg ulcers, decreased wound healing, pulmonary hypertension, silent brain infarcts, and increased thrombosis to count a few. Interestingly many of those complications overlap with those found in NTDT patients. Paul I. Tyan, Amr H. Radwan, Assaad Eid, Anthony G. Haddad, David Wehbe, and Ali T. Taher Copyright © 2014 Paul I. Tyan et al. All rights reserved. Beta-Thalassaemia Intermedia: Evaluation of Endocrine and Bone Complications Mon, 07 Jul 2014 11:01:38 +0000 Objective. Data about endocrine and bone disease in nontransfusion-dependent thalassaemia (NTDT) is scanty. The aim of our study was to evaluate these complications in β-TI adult patients. Methods. We studied retrospectively 70 β-TI patients with mean followup of 20 years. Data recorded included age, gender, haemoglobin and ferritin levels, biochemical and endocrine tests, liver iron concentration (LIC) from , transfusion regimen, iron chelation, hydroxyurea, splenectomy, and bone mineralization by dual X-ray absorptiometry. Results. Thirty-seven (53%) males and 33 (47%) females were studied, with mean age years, mean haemoglobin  g/dL, median ferritin 537 (range 14–4893), and mean LIC  mg Fe/g dw. Thirty-three patients (47%) had been transfused, occasionally (24/33; 73%) or regularly (9/33; 27%); 37/70 (53%) had never been transfused; 34/70 patients had been splenectomized (49%); 39 (56%) were on chelation therapy; and 11 (16%) were on hydroxyurea. Endocrinopathies were found in 15 patients (21%): 10 hypothyroidism, 3 hypogonadism, 2 impaired glucose tolerance (IGT), and one diabetes. Bone disease was observed in 53/70 (76%) patients, osteoporosis in 26/53 (49%), and osteopenia in 27/53 (51%). Discussion and Conclusions. Bone disease was found in most patients in our study, while endocrinopathies were highly uncommon, especially hypogonadism. We speculate that low iron burden may protect against endocrinopathy development. M. Baldini, A. Marcon, R. Cassin, F. M. Ulivieri, D. Spinelli, M. D. Cappellini, and G. Graziadei Copyright © 2014 M. Baldini et al. All rights reserved. Implications of Heterogeneity in Multiple Myeloma Wed, 02 Jul 2014 10:03:48 +0000 Multiple myeloma is the second most common hematologic malignancy in the world. Despite improvement in outcome, the disease is still incurable for most patients. However, not all myeloma are the same. With the same treatment, some patients can have very long survival whereas others can have very short survival. This suggests that there is underlying heterogeneity in myeloma. Studies over the years have revealed multiple layers of heterogeneity. First, clinical parameters such as age and tumor burden could significantly affect outcome. At the genetic level, there are also significant heterogeneity ranging for chromosome numbers, genetic translocations, and genetic mutations. At the clonal level, there appears to be significant clonal heterogeneity with multiple clones coexisting in the same patient. At the cell differentiation level, there appears to be a hierarchy of clonally related cells that have different clonogenic potential and sensitivity to therapies. These levels of complexities present challenges in terms of treatment and prognostication as well as monitoring of treatment. However, if we can clearly delineate and dissect this heterogeneity, we may also be presented with unique opportunities for precision and personalized treatment of myeloma. Some proof of concepts of such approaches has been demonstrated. Sanjay de Mel, Su Hong Lim, Moon Ley Tung, and Wee-Joo Chng Copyright © 2014 Sanjay de Mel et al. All rights reserved. MMSET: Role and Therapeutic Opportunities in Multiple Myeloma Tue, 01 Jul 2014 10:37:19 +0000 Recurrent chromosomal translocations are central to the pathogenesis, diagnosis, and prognosis of hematologic malignancies. The translocation t(4; 14)(p16; q32) is one of the most common translocations in multiple myeloma (MM) and is associated with very poor prognosis. The t(4; 14) translocation leads to the simultaneous overexpression of two genes, FGFR3 (fibroblast growth factor receptor 3) and MMSET (multiple myeloma SET domain), both of which have potential oncogenic activity. However, approximately 30% of t(4; 14) MM patients do not express FGFR3 and have poor prognosis irrespective of FGFR3 expression, whereas MMSET overexpression is universal in t(4; 14) cases. In this review, we provide an overview of recent findings regarding the oncogenic roles of MMSET in MM and its functions on histone methylation. We also highlight some of MMSET partners and its downstream signalling pathways and discuss the potential therapeutics targeting MMSET. Zhigang Xie and Wee Joo Chng Copyright © 2014 Zhigang Xie and Wee Joo Chng. All rights reserved. Bone Marrow-Derived Mesenchymal Cell Differentiation toward Myogenic Lineages: Facts and Perspectives Thu, 26 Jun 2014 12:25:30 +0000 Bone marrow-derived mesenchymal stem cells (BM-MSCs) are valuable platforms for new therapies based on regenerative medicine. BM-MSCs era is coming of age since the potential of these cells is increasingly demonstrated. In fact, these cells give origin to osteoblasts, chondroblasts, and adipocyte precursors in vitro, and they can also differentiate versus other mesodermal cell types like skeletal muscle precursors and cardiomyocytes. In our short review, we focus on the more recent manipulations of BM-MSCs toward skeletal and heart muscle differentiation, a growing field of obvious relevance considering the toll of muscle disease (i.e., muscular dystrophies), the heavier toll of heart disease in developed countries, and the still not completely understood mechanisms of muscle differentiation and repair. Daniela Galli, Marco Vitale, and Mauro Vaccarezza Copyright © 2014 Daniela Galli et al. All rights reserved. Aberrant Frequency of IL-10-Producing B Cells and Its Association with Treg/Th17 in Adult Primary Immune Thrombocytopenia Patients Thu, 26 Jun 2014 09:13:00 +0000 Background. Regulatory B cells (Breg) are a distinct B cell subset with immunoregulatory properties. Pivotal to Breg function is interleukin-10. This study was to investigate the role of IL-10-producing B cell (B10) and its association with Treg and Th17 subsets in immune thrombocytopenia (ITP) patients. Methods. Peripheral blood mononuclear cells from ITP patients and controls were stimulated with PMA, ionomycin, and Brefeldin A. The frequencies of CD19+IL-10+ B cells, CD3+CD4+IL-17+ Th17 cells, and CD4+CD25hiFoxp3+ Treg cells were analyzed by flow cytometry. The mRNA expression of Foxp3 and RORγt was detected by real-time quantitative PCR. Results. The number of B10 cells was elevated in ITP patients. After first-line therapies, it remained at high level in patients who achieved complete or partial response but decreased in those who acquired no response. There was a positive correlation between B10 cells and Tregs in ITP both before and after therapies. The ratio of Treg/Th17 decreased in ITP, and it strongly correlated with B10 cells. Conclusions. The frequency of B10 cells is elevated in ITP and it correlates with both the Tregs counts and the Treg/Th17 ratio. B10 cells to regulate functional T cell subsets might be impaired in patients with ITP. Fanli Hua, Lili Ji, Yanxia Zhan, Feng Li, Shanhua Zou, Lihang Chen, Song Gao, Ying Li, Hao Chen, and Yunfeng Cheng Copyright © 2014 Fanli Hua et al. All rights reserved. Increased Oxidative Damage Associated with Unfavorable Cytogenetic Subgroups in Chronic Lymphocytic Leukemia Thu, 26 Jun 2014 08:54:34 +0000 Oxidative stress contributes to genomic instability in chronic lymphocytic leukemia (CLL), but its relationship with the acquisition of specific chromosomal abnormalities is unknown. We recruited 55 untreated CLL patients and assessed 8-oxo-2′-deoxyguanosine (8-oxo-dG), glutathione, and malondialdehyde (MDA) levels, and we compared them among the cytogenetic subgroups established using fluorescence in situ hybridization (FISH). Significant increases in 8-oxo-dG and/or MDA were observed in patients with unfavorable cytogenetic aberrations (17p and 11q deletions) compared to the 13q deletion group. TP53 deletion patients exhibited a diminished DNA repair efficiency. Finally, cases with normal FISH also showed enhanced 8-oxo-dG, which could result in adverse outcomes. Rosa Collado, David Ivars, Isabel Oliver, Carmen Tormos, Mercedes Egea, Amparo Miguel, Guillermo T. Sáez, and Félix Carbonell Copyright © 2014 Rosa Collado et al. All rights reserved. Changes in Osteoblastic Activity in Patient Who Received Bortezomib as Second Line Treatment for Plasma Cell Myeloma: A Prospective Multicenter Study Mon, 23 Jun 2014 08:44:31 +0000 We conducted a prospective multicenter study identifying the role of bortezomib in patients with relapsed or refractory plasma cell myeloma (PCM) in bone resorption and formation via bone turnover markers. A total of 104 patients received at least 1 cycle of bortezomib. Most of them had advanced disease (). Among them, 75 patients completed 4 cycles of treatment. Most of the patients (81.7%) were treated in combination with steroid. After the 4th cycle treatment, 47 of 75 patients achieved CR, nCR, VGPR, and PR (64.4%), while 26 patients achieved less than PR (35.6%). The proportion of patients who achieved ≥ PR increased as patients received more treatment cycles, reaching 90% after the 8th cycle. DKK-1 levels decreased significantly posttreatment. Bone formation markers (bALP and OC) and osteoclast regulator such as sRANKL also decreased significantly. These findings were observed primarily in patients who received steroid and who had a longer disease duration. While sRANKL demonstrated significant reduction posttreatment, osteoprotegerin (OPG) level did not significantly change posttreatment, resulting in a decreased sRANKL/OPG ratio (). In conclusion, our clinical data suggest that treatment with bortezomib and steroid may rearrange the metabolic balance between osteoblast and osteoclast activities in PCM. Ki-Seong Eom, Seok Jin Kim, Je-Jung Lee, Cheolwon Suh, Jin Seok Kim, Sung-Soo Yoon, Byung Soo Kim, Hye Jin Kang, Young Jin Choi, Chul Soo Kim, Yang Soo Kim, Jae-Yong Kwak, Yoo Jin Kim, Young Don Joo, Yeung-Chul Mun, Deog Yeon Jo, Joon Seong Park, Chi-Young Park, Sung-Hyun Kim, and Chang-Ki Min Copyright © 2014 Ki-Seong Eom et al. All rights reserved. p53 Abnormalities and Potential Therapeutic Targeting in Multiple Myeloma Tue, 17 Jun 2014 08:26:21 +0000 p53 abnormalities are regarded as an independent prognostic marker in multiple myeloma. Patients harbouring this genetic anomaly are commonly resistant to standard therapy. Thus, various p53 reactivating agents have been developed in order to restore its tumour suppressive abilities. Small molecular compounds, especially, have gained popularity in its efficacy against myeloma cells. For instance, promising preclinical results have steered both nutlin-3 and PRIMA-1 into phase I/II clinical trials. This review summarizes different modes of p53 inactivation in myeloma and highlights the current p53-based therapies that are being utilized in the clinic. Finally, we discuss the potential and promise that the novel small molecules possess for clinical application in improving the treatment outcome of myeloma. P. J. Teoh and W. J. Chng Copyright © 2014 P. J. Teoh and W. J. Chng. All rights reserved. Preventive Strategies against Bleeding due to Nonvitamin K Antagonist Oral Anticoagulants Mon, 16 Jun 2014 09:16:33 +0000 Dabigatran etexilate (DE), rivaroxaban, and apixaban are nonvitamin K antagonist oral anticoagulants (NOACs) that have been compared in clinical trials with existing anticoagulants (warfarin and enoxaparin) in several indications for the prevention and treatment of thrombotic events. All NOACs presented bleeding events despite a careful selection and control of patients. Compared with warfarin, NOACs had a decreased risk of intracranial hemorrhage, and apixaban and DE (110 mg BID) had a decreased risk of major bleeding from any site. Rivaroxaban and DE showed an increased risk of major gastrointestinal bleeding compared with warfarin. Developing strategies to minimize the risk of bleeding is essential, as major bleedings are reported in clinical practice and specific antidotes are currently not available. In this paper, the following preventive approaches are reviewed: improvement of appropriate prescription, identification of modifiable bleeding risk factors, tailoring NOAC’s dose, dealing with a missed dose as well as adhesion to switching, bridging and anesthetic procedures. Lessire Sarah, Dincq Anne-Sophie, Douxfils Jonathan, Devalet Bérangère, Nicolas Jean-Baptiste, Spinewine Anne, Larock Anne-Sophie, Dogné Jean-Michel, Gourdin Maximilien, and Mullier François Copyright © 2014 Lessire Sarah et al. All rights reserved. Efficacy and Pharmacologic Data of Second-Generation Tyrosine Kinase Inhibitor Nilotinib in BCR-ABL-Positive Leukemia Patients with Central Nervous System Relapse after Allogeneic Stem Cell Transplantation Sun, 15 Jun 2014 10:58:24 +0000 Central nervous system (CNS) involvement is a severe complication of BCR-ABL-positive leukemia after allogenic stem cell transplantation (alloSCT) associated with fatal outcome. Although second-generation tyrosine-kinase inhibitors (TKI) such as nilotinib have shown activity in systemic BCR-ABL+ disease, little data exists on their penetration and efficacy within the CNS. Four patients (3 male, 1 female; age 15–49) with meningeal relapse after alloSCT and subsequent treatment with nilotinib were identified. A total of 17 cerebrospinal fluid (csf) and serum samples were assessed for nilotinib concentration and patient outcome was recorded. Nilotinib concentrations showed a low median csf/plasma ratio of 0.53% (range 0.23–1.5%), yet pronounced clinical efficacy was observed with long-lasting responses (>1 year) in three patients. Comparison with historical data showed a trend towards superior efficacy of nilotinib versus imatinib. Despite poor csf penetration, nilotinib showed significant clinical activity in CNS relapse of BCR-ABL+ leukemias. As nilotinib has a high protein-binding affinity, the low-protein concentration in csf could translate into a relatively higher amount of free and therefore active nilotinib in csf as compared to blood, possibly explaining the observed efficacy. Thus, treatment with a 2nd generation TKI warrants further investigation and should be considered in cases of CNS relapse of BCR-ABL-positive leukemia after alloSCT. Mark Reinwald, Eberhard Schleyer, Philipp Kiewe, Igor Wolfgang Blau, Thomas Burmeister, Stefan Pursche, Martin Neumann, Michael Notter, Eckhard Thiel, Wolf-Karsten Hofmann, Hans-Jochem Kolb, Stefan Burdach, and Hans-Ulrich Bender Copyright © 2014 Mark Reinwald et al. All rights reserved. Blood Donation, Being Asian, and a History of Iron Deficiency Are Stronger Predictors of Iron Deficiency than Dietary Patterns in Premenopausal Women Wed, 11 Jun 2014 11:50:17 +0000 This study investigated dietary patterns and nondietary determinants of suboptimal iron status (serum ferritin < 20 μg/L) in 375 premenopausal women. Using multiple logistic regression analysis, determinants were blood donation in the past year [OR: 6.00 (95% CI: 2.81, 12.82); ], being Asian [OR: 4.84 (95% CI: 2.29, 10.20); ], previous iron deficiency [OR: 2.19 (95% CI: 1.16, 4.13); ], a “milk and yoghurt” dietary pattern [one SD higher score, OR: 1.44 (95% CI: 1.08, 1.93); ], and longer duration of menstruation [days, OR: 1.38 (95% CI: 1.12, 1.68); ]. A one SD change in the factor score above the mean for a “meat and vegetable” dietary pattern reduced the odds of suboptimal iron status by 79.0% [OR: 0.21 (95% CI: 0.08, 0.50); ] in women with children. Blood donation, Asian ethnicity, and previous iron deficiency were the strongest predictors, substantially increasing the odds of suboptimal iron status. Following a “milk and yoghurt” dietary pattern and a longer duration of menstruation moderately increased the odds of suboptimal iron status, while a “meat and vegetable” dietary pattern reduced the odds of suboptimal iron status in women with children. Kathryn L. Beck, Cathryn A. Conlon, Rozanne Kruger, Anne-Louise M. Heath, Christophe Matthys, Jane Coad, Beatrix Jones, and Welma Stonehouse Copyright © 2014 Kathryn L. Beck et al. All rights reserved. Expression of Myeloid Antigen in Neoplastic Plasma Cells Is Related to Adverse Prognosis in Patients with Multiple Myeloma Wed, 04 Jun 2014 09:59:13 +0000 We evaluated the association between the expression of myeloid antigens on neoplastic plasma cells and patient prognosis. The expression status of CD13, CD19, CD20, CD33, CD38, CD56, and CD117 was analyzed on myeloma cells from 55 newly diagnosed patients, including 36 men (65%), of median age 61 years (range: 38–78). Analyzed clinical characteristics and laboratory parameters were as follows: serum β2-microglobulin, lactate dehydrogenase, calcium, albumin, hemoglobin, serum creatinine concentrations, bone marrow histology, and cytogenetic findings. CD13+ and CD33+ were detected in 53% and 18%, respectively. Serum calcium () and LDH () concentrations were significantly higher and morphologic subtype of immature or plasmablastic was more frequent in CD33+ than in CD33− patients (). CD33 and CD13 expression demonstrate a potential prognostic impact and were associated with lower overall survival (OS; and ) in Kaplan-Meier analysis. Multivariate analysis showed that CD33 was independently prognostic of shorter progression free survival (PFS; ) and OS () with correction of clinical prognostic factors. This study showed that CD13 and CD33 expression associated with poor prognosis in patients with MM implicating the need of analysis of these markers in MM diagnosis. Hyoeun Shim, Joo Hee Ha, Hyewon Lee, Ji Yeon Sohn, Hyun Ju Kim, Hyeon-Seok Eom, and Sun-Young Kong Copyright © 2014 Hyoeun Shim et al. All rights reserved. The Positive Effects of One-Hour Intravenous Administration of Bortezomib on Peripheral Neuropathy in Multiple Myeloma Patients Wed, 04 Jun 2014 09:57:53 +0000 Bortezomib-induced peripheral neuropathy (BiPN) in multiple myeloma (MM) patients is a common and serious side effect. Currently, it has been reported that subcutaneous (SC) administration of bortezomib decreases the incidence of BiPN as compared to standard intravenous (IV) bolus injection without any differences in efficacy. However, there are reports of severe injection site reaction following SC administration of bortezomib. The aim of this study was to evaluate the response rate and incidence of BiPN following one-hour IV infusion of bortezomib. The data was retrospectively collected from MM patients who had been treated with IV administration of bortezomib for one hour. Twenty-three patients were evaluated (median age 72 years, 13 males). The median number of treatment cycles was 5 (range 2–10). The cumulative bortezomib dose was 26.0 mg/m2 (14.3–66.3) and percent of actual per expected cumulative dose was 90% (50–100). The overall response (complete response plus partial response) rate was 65%. The incidence of BiPN was 57% (n = 13) and incidence of severe neuropathy was 4% (n = 1). One-hour IV infusion of bortezomib was an effective regimen for MM with reduced incidence of severe BiPN. This route of administration of bortezomib could be an alternative mode of delivery for patients with severe injection site reactions following SC administration. Joo Young Jung, Ho Young Kim, Boram Han, Dae Ro Choi, Dae Young Zang, and Hyo Jung Kim Copyright © 2014 Joo Young Jung et al. All rights reserved. Clinical Factors Associated with Response or Survival after Chemotherapy in Patients with Waldenström Macroglobulinemia in Korea Wed, 04 Jun 2014 09:55:12 +0000 Waldenström’s macroglobulinemia (WM) is a B-cell proliferative malignancy characterized by immunoglobulin M monoclonal gammopathy and bone marrow infiltration by lymphoplasmacytic cells. Clinical features and cytogenetics of WM in Asia including Republic of Korea remain unclear. Moreover, no study has reported treatment outcomes in patients with WM treated with novel agent combined with conventional chemotherapy. This study investigated clinical features and assessed treatment outcomes with novel agent and conventional chemotherapy in Republic of Korea. Data from all () patients with newly diagnosed WM at 17 hospitals who received chemotherapy between January 2005 and December 2012 were collected retrospectively. The median age of patients was 66 years (range: 37–92 years) and male to female ratio was 5 : 1. Patients treated with novel agent combined chemotherapy displayed higher overall response rate (ORR) compared to conventional chemotherapy alone (92.9% versus 52.6%, ). The 5-year overall survival rate was 62.6% (95% confidence interval: 34.73–111.07). Use of novel agents produced higher ORR but survival benefit was not apparent due to the small number of patients and short follow-up duration. Further studies are needed to confirm the efficacy of novel agents in patients with WM. Ho Sup Lee, Kihyun Kim, Dok Hyun Yoon, Jin Seok Kim, Soo-Mee Bang, Jeong-Ok Lee, Hyeon Seok Eom, Hyewon Lee, Inho Kim, Won Sik Lee, Sung Hwa Bae, Se Hyung Kim, Mark Hong Lee, Young Rok Do, Jae Hoon Lee, Junshik Hong, Ho-Jin Shin, Ji Hyun Lee, Yeung-Chul Mun, and Chang-Ki Min Copyright © 2014 Ho Sup Lee et al. All rights reserved. Plasma Levels of Osteopontin and Vascular Endothelial Growth Factor in Association with Clinical Features and Parameters of Tumor Burden in Patients with Multiple Myeloma Wed, 04 Jun 2014 07:39:50 +0000 The aim of this pilot study was to determine the plasma levels of osteopontin (OPN) and vascular endothelial growth factor (VEGF) and find possible association between them and main clinical features and parameters of tumor burden in patient with multiple myeloma (MM). Plasma levels of OPN and VEGF were determined in 44 newly diagnosed MM patients and 24 healthy persons by ELISA method. These values were compared with the presence of anemia, renal dysfunction, and bone lesions as myeloma related clinical manifestations and with serum beta-2 microglobulin and Durie-Salmon clinical stage as prognosticators related to tumor mass. The value of OPN was significantly higher in MM patients with evident bone lesions () and there was also a positive correlation with serum beta-2 microglobulin (; ). Furthermore, patients with lower Durie-Salmon stage had significantly lower OPN and VEGF levels (; , resp.). Our preliminary results found positive association between plasma level of OPN, tumor burden, and bone destruction. Further analysis should provide information about the possible use of OPN as useful clinical biomarker for monitoring bone disease and tumor mass, as well as a prognostic factor, or a possible target for pharmacological intervention. Toni Valković, Emina Babarović, Ksenija Lučin, Sanja Štifter, Merica Aralica, Sanja Pećanić, Irena Seili-Bekafigo, Antica Duletić-Načinović, Damir Nemet, and Nives Jonjić Copyright © 2014 Toni Valković et al. All rights reserved. MicroRNA: Important Player in the Pathobiology of Multiple Myeloma Tue, 03 Jun 2014 11:30:01 +0000 Recent studies have revealed a pivotal role played by a class of small, noncoding RNAs, microRNA (miRNA), in multiple myeloma (MM), a plasma cell (PC) malignancy causing significant morbidity and mortality. Deregulated miRNA expression in patient’s PCs and plasma has been associated with tumor progression, molecular subtypes, clinical staging, prognosis, and drug response in MM. A number of important oncogenic and tumor suppressor miRNAs have been discovered to regulate important genes and pathways such as p53 and IL6-JAK-STAT signaling. miRNAs may also form complex regulatory circuitry with genetic and epigenetic machineries, the deregulation of which could lead to malignant transformation and progression. The translational potential of miRNAs in the clinic is being increasingly recognized that they could represent novel biomarkers and therapeutic targets. This review comprehensively summarizes current progress in delineating the roles of miRNAs in MM pathobiology and management. Chonglei Bi and Wee Joo Chng Copyright © 2014 Chonglei Bi and Wee Joo Chng. All rights reserved. Genome-Wide Screening of Cytogenetic Abnormalities in Multiple Myeloma Patients Using Array-CGH Technique: A Czech Multicenter Experience Mon, 02 Jun 2014 09:29:57 +0000 Characteristic recurrent copy number aberrations (CNAs) play a key role in multiple myeloma (MM) pathogenesis and have important prognostic significance for MM patients. Array-based comparative genomic hybridization (aCGH) provides a powerful tool for genome-wide classification of CNAs and thus should be implemented into MM routine diagnostics. We demonstrate the possibility of effective utilization of oligonucleotide-based aCGH in 91 MM patients. Chromosomal aberrations associated with effect on the prognosis of MM were initially evaluated by I-FISH and were found in 93.4% (85/91). Incidence of hyperdiploidy was 49.5% (45/91); del(13)(q14) was detected in 57.1% (52/91); gain(1)(q21) occurred in 58.2% (53/91); del(17)(p13) was observed in 15.4% (14/91); and t(4;14)(p16;q32) was found in 18.6% (16/86). Genome-wide screening using Agilent 44K aCGH microarrays revealed copy number alterations in 100% (91/91). Most common deletions were found at 13q (58.9%), 1p (39.6%), and 8p (31.1%), whereas gain of whole 1q was the most often duplicated region (50.6%). Furthermore, frequent homozygous deletions of genes playing important role in myeloma biology such as TRAF3, BIRC1/BIRC2, RB1, or CDKN2C were observed. Taken together, we demonstrated the utilization of aCGH technique in clinical diagnostics as powerful tool for identification of unbalanced genomic abnormalities with prognostic significance for MM patients. Jan Smetana, Jan Frohlich, Romana Zaoralova, Vladimira Vallova, Henrieta Greslikova, Renata Kupska, Pavel Nemec, Aneta Mikulasova, Martina Almasi, Ludek Pour, Zdenek Adam, Viera Sandecka, Lenka Zahradová, Roman Hajek, and Petr Kuglik Copyright © 2014 Jan Smetana et al. All rights reserved. Clinical Comparisons of Two Free Light Chain Assays to Immunofixation Electrophoresis for Detecting Monoclonal Gammopathy Tue, 27 May 2014 05:49:28 +0000 Free light chains (FLCs) are useful biomarkers for the diagnosis and monitoring of various plasma cell dyscrasias. One hundred fifty-seven samples from 120 patients for screening or monitoring of monoclonal gammopathy (MG) were included. The new N Latex FLC assays (Siemens Healthcare Diagnostics GmbH, Germany) were compared with the Freelite FLC assays (The Binding Site Ltd., UK) and the results were analyzed with those of immunofixation electrophoresis (IFE). The Freelite FLC assay showed significantly wider assay ranges than the N Latex FLC assay. The correlation coefficients of the two FLC kappa (κ) assays, lambda (λ) assays, and the κ/λ ratio were 0.9792, 0.8264, and 0.9064, respectively. The concordance rate was 84.7% for the FLC κ assays, 79.6% for FLC λ, and 89.2% for the κ/λ ratio. The clinical sensitivity and specificity of the κ/λ ratios were 72.2% and 93.6% for the Freelite assay and 64.6% and 100% for the N Latex FLC assay. Two FLC assays showed good correlations and concordance. However, the clinical sensitivity of the κ/λ ratio was higher in the Freelite FLC assays; clinical specificity was higher in the N Latex FLC assay. Both FLC assays seem to have limited clinical utility in detecting MG in certain clinical settings. Han-Sung Kim, Hyun Soo Kim, Kyu-Sung Shin, Wonkeun Song, Hyo Jung Kim, Hyeong Su Kim, and Min-Jeong Park Copyright © 2014 Han-Sung Kim et al. All rights reserved. Genetic Abnormalities in Chronic Lymphocytic Leukemia: Where We Are and Where We Go Thu, 22 May 2014 11:52:33 +0000 Chromosomal abnormalities in chronic lymphocytic leukemia (CLL) are detected in up to 80% of patients. Among them, deletions of 11q, 13q, 17p, and trisomy 12 have a known prognostic value and play an important role in CLL pathogenesis and evolution, determining patients outcome and therapeutic strategies. Standard methods used to identify these genomic aberrations include both conventional G-banding cytogenetics (CGC) and fluorescence in situ hybridization (FISH). Although FISH analyses have been implemented as the gold standard, CGC allows the identification of chromosomal translocations and complex karyotypes, the latest associated with poor outcome. Genomic arrays have a higher resolution that allows the detection of cryptic abnormalities, although these have not been fully implemented in routine laboratories. In the last years, next generation sequencing (NGS) methods have identified a wide range of gene mutations (e.g., TP53, NOTCH1, SF3B1, and BIRC3) which have improved our knowledge about CLL development, allowing us to refine both the prognostic subgroups and better therapeutic strategies. Clonal evolution has also recently arisen as a key point in CLL, integrating cytogenetic alterations and mutations in a dynamic model that improve our understanding about its clinical course and relapse. Anna Puiggros, Gonzalo Blanco, and Blanca Espinet Copyright © 2014 Anna Puiggros et al. All rights reserved. Elevated Red Blood Cell Distribution Width as a Simple Prognostic Factor in Patients with Symptomatic Multiple Myeloma Wed, 21 May 2014 07:42:30 +0000 Red blood cell distribution width (RDW) is a parameter reported in complete blood cell count tests, and has been reported as an inflammatory biomarker. Multiple myeloma (MM) is known to be associated with inflammatory microenvironments. However, the importance of RDW has been seldom studied in MM. For this study, 146 symptomatic myeloma patients with available RDW at diagnosis were retrospectively reviewed, and their characteristics were compared between two groups, those with high (>14.5%) and normal (≤14.5%) RDW. RDW was correlated to hemoglobin, MM stage, β2-microglobulin, M-protein, bone marrow plasma cells, and cellularity (). During induction, overall response rates of the two groups were similar (); however, complete response rate was higher in the normal-RDW group than it was in the high-RDW group (). With a median follow-up of 47 months, the normal-RDW group showed better progression-free survival (PFS) (24.2 versus 17.0 months, ) compared to the high-RDW group. Overall survival was not different according to the RDW level (). In multivariate analysis, elevated RDW at diagnosis was a poor prognostic factor for PFS (HR 3.21, 95% CI 1.24–8.32) after adjustment with other myeloma-related prognostic factors. RDW would be a simple and immediately available biomarker of symptomatic MM, reflecting the systemic inflammation. Hyewon Lee, Sun-Young Kong, Ji Yeon Sohn, Hyoeun Shim, Hye Sun Youn, Sangeun Lee, Hyun Ju Kim, and Hyeon-Seok Eom Copyright © 2014 Hyewon Lee et al. All rights reserved. Serum Parathyroid Hormone Is a New Potential Risk Factor in Multiple Myeloma Mon, 19 May 2014 12:15:38 +0000 We hypothesized that serum PTH might be associated with various clinicopathological parameters in multiple myeloma (MM). So we investigated the implications of serum PTH in MM patients and the relationship with other risk factors of MM. A total of 115 patients who were newly diagnosed with MM were enrolled. Serum PTH level was 24.7 ± 34.9 (ranged 0.0–284.1) pg/mL. Serum levels of IgG, IgM, FLC-lambda, albumin, and LDH were in positive correlation with serum PTH. Compared to non-high PTH (<68.3 pg/mL) group, the hazard ratio (HR) for overall survival was higher for group with high PTH level (≥68.3 pg/mL) (HR, 1.710). Furthermore, the patient group with high PTH level showed inferior progression-free survival than non-high PTH group (). Interestingly, subgroup analysis showed that serum PTH level at diagnosis was associated with risk factors and clinical outcome in MM patients, especially in complete remission group, transplantation cases, ISS stage II cases, and cases without chromosome abnormality. In conclusion, this study showed that blood PTH level in MM at diagnosis was associated with risk factors and clinical outcome in MM patients. Min-Gu Kang, Eun-Jeong Won, Hyun-Woo Choi, Hye-Ran Kim, Hyun-Jung Choi, Hye-Ri Park, Jong-Hee Shin, Soon-Pal Suh, Dong-Wook Ryang, and Myung-Geun Shin Copyright © 2014 Min-Gu Kang et al. All rights reserved. Salvage Therapy of Multiple Myeloma: The New Generation Drugs Mon, 19 May 2014 05:25:02 +0000 During the past decade, overall results of treatment of multiple myeloma (MM) have been improved and survival curves are now significantly better with respect to those obtained with historical treatment. These improvements are linked to a deeper knowledge of the biology of disease and to the introduction in clinical practice of drugs with different mechanism of action such as proteasome inhibitors and immunomodulatory drugs (IMiDs). However, MM remains in most cases an incurable disease. For patients who relapse after treatment with novel agents, the prognosis is dismal and new drugs and therapeutic strategies are required for continued disease control. In this review, we summarize new insights in salvage therapy for relapsed/refractory MM as emerging from recent clinical trials exploring the activity of bendamustine, new generation proteasome inhibitors, novel IMiDs, monoclonal antibodies, and drugs interfering with growth pathways. Alessandra Romano, Concetta Conticello, Maide Cavalli, Calogero Vetro, Cosimo Di Raimondo, Valentina Di Martina, Elena Schinocca, Alessia La Fauci, Nunziatina Laura Parrinello, Annalisa Chiarenza, and Francesco Di Raimondo Copyright © 2014 Alessandra Romano et al. All rights reserved. Recombinant Human Factor IX Produced from Transgenic Porcine Milk Sun, 18 May 2014 10:32:02 +0000 Production of biopharmaceuticals from transgenic animal milk is a cost-effective method for highly complex proteins that cannot be efficiently produced using conventional systems such as microorganisms or animal cells. Yields of recombinant human factor IX (rhFIX) produced from transgenic porcine milk under the control of the bovine -lactalbumin promoter reached 0.25 mg/mL. The rhFIX protein was purified from transgenic porcine milk using a three-column purification scheme after a precipitation step to remove casein. The purified protein had high specific activity and a low ratio of the active form (FIXa). The purified rhFIX had 11.9 -carboxyglutamic acid (Gla) residues/mol protein, which approached full occupancy of the 12 potential sites in the Gla domain. The rhFIX was shown to have a higher isoelectric point and lower sialic acid content than plasma-derived FIX (pdFIX). The rhFIX had the same -glycosylation sites and phosphorylation sites as pdFIX, but had a higher specific activity. These results suggest that rhFIX produced from porcine milk is physiologically active and they support the use of transgenic animals as bioreactors for industrial scale production in milk. Meng-Hwan Lee, Yin-Shen Lin, Ching-Fu Tu, and Chon-Ho Yen Copyright © 2014 Meng-Hwan Lee et al. All rights reserved. Polymorphism of XRCC1, XRCC3, and XPD Genes and Risk of Chronic Myeloid Leukemia Thu, 15 May 2014 12:28:33 +0000 The genetic polymorphisms of X-ray repair cross complementing group 1 (XRCC1), X-ray repair cross complementing group 3 (XRCC3), and xeroderma pigmentosum complementation group D (XPD) repair genes may lead to genetic instability and leukemogenesis. The purpose of the study was to evaluate the association between XRCC1 Arg399Gln, Arg280His and Arg194Trp, XRCC3 Thr241Met, and XPD Lys751Gln polymorphisms and the risk of developing CML in Romanian patients. A total of 156 patients diagnosed with CML and 180 healthy controls were included in this study. We found no association between CML and XRCC1 or XRCC3 variant genotypes in any of the investigated cases. A significant difference was observed in the variant genotype frequencies of the XPD Lys751Gln polymorphism between the patients with CML and control group (for variant homozygous genotypes, ; 95% –4.67; P value = 0.016 and for combined heterozygous and variant homozygous genotypes, ; 95% –2.69; P value = 0.019). This was also observed when analyzing the variant 751Gln allele (; 95% –2.11; P value = 0.008). Our results suggest that the XPD Lys751Gln variant genotype increases the risk of CML. Claudia Bănescu, Adrian P. Trifa, Smaranda Demian, Erzsebeth Benedek Lazar, Delia Dima, Carmen Duicu, and Minodora Dobreanu Copyright © 2014 Claudia Bănescu et al. All rights reserved. MYD88 L265P Mutations Are Correlated with 6q Deletion in Korean Patients with Waldenström Macroglobulinemia Wed, 07 May 2014 12:12:19 +0000 Waldenström macroglobulinemia (WM) is a malignant lymphoplasma-proliferative disorder with IgM monoclonal gammopathy. A recent whole-genome study identified MYD88 L265P as the key mutation in WM. We investigated MYD88 mutations in conjunction with cytogenetic study in 22 consecutive Korean WM patients. Conventional G-banding and interphase fluorescence in situ hybridization (FISH) were performed at regions including 6q21 using bone marrow (BM) aspirates. Sixteen patients were subjected to Sanger sequencing-based MYD88 mutation study. Five patients (28%) showed cytogenetic aberrations in G-banding. The incidence of 6q21 deletion was 17% by conventional G-banding and 37% by FISH. Ten patients (45%) showed cytogenetic aberrations using FISH: 6q deletion in eight (37%) and IGH rearrangement in four (18%). Two patients had both the 6q deletion and IGH rearrangement, and two had only the IGH rearrangement. Eleven patients (69%) presented with the MYD88 L265P mutation. MYD88 mutations were significantly associated with the presence of 6q deletions (). Six patients with the 6q deletion for whom sequencing was possible were found to harbor MYD88 mutations. The MYD88 L265P mutation was also associated with increased lymphocyte burden in BM biopsy. This is the first report of high frequency MYD88 L265P mutations in Korean WM patients. Jung-Ah Kim, Kyongok Im, Si Nae Park, Jiseok Kwon, Qute Choi, Sang Mee Hwang, Naohiro Sekiguchi, Sung-Soo Yoon, Dong Soon Lee, and Seon Young Kim Copyright © 2014 Jung-Ah Kim et al. All rights reserved. Bone Marrow Plasma Cell Assessment before Peripheral Blood Stem Cell Mobilization in Patients with Multiple Myeloma Undergoing Autologous Stem Cell Transplantation Tue, 06 May 2014 07:29:33 +0000 The current definition of complete response (CR) in multiple myeloma (MM) includes negative serum and urine immunofixation (IFE) tests and <5% bone marrow plasma cells (BMPCs). However, many studies of the prognostic impact of pretransplant response have not included BMPCs. We evaluated the prognostic impact of BMPC assessment before peripheral blood stem cell (PBSC) mobilization on subsequent transplant outcomes. BMPCs were assessed by CD138, kappa, and lambda immunostaining in 106 patients. After a median followup of 24.5 months, patients with <5% BMPCs had a significantly better progression-free survival (PFS) compared to those with ≥5% BMPCs . Patients with <5% BMPCs + serologic CR showed superior PFS compared to those with <5% BMPCs + serologic non-CR or ≥5% BMPCs + serologic non-CR . Interestingly, the prognostic impact of BMPCs was more apparent for patients who did not achieve a serologic CR compared to those with a serologic CR . We concluded that IFE negativity and <5% BMPCs before PBSC mobilization were important factors to predict PFS in patients with MM undergoing ASCT. Particularly, a significant impact of <5% BMPCs was observed in patients who did not achieve IFE negativity. Sung-Eun Lee, Jae-Ho Yoon, Seung-Hwan Shin, Ki-Seong Eom, Yoo-Jin Kim, Hee-Je Kim, Seok Lee, Seok-Goo Cho, Jong Wook Lee, Woo-Sung Min, Chong-Won Park, Myungshin Kim, and Chang-Ki Min Copyright © 2014 Sung-Eun Lee et al. All rights reserved. Psychometric Properties of the Greek Haem-A-QoL for Measuring Quality of Life in Greek Haemophilia Patients Tue, 06 May 2014 00:00:00 +0000 Background and Objectives. Health Related Quality of Life (HRQoL) is an important health outcome measure in haemophilia. The aim of this study was to assess the psychometric properties of the Greek version of Haem-A-QoL, a disease-specific questionnaire for haemophiliacs. Methods. Haem-A-QoL and SF-36 were administered to 118 adult haemophilia patients. Hypothesized scale structure, internal consistency (Cronbach’s α), and test-retest reliability, as well as various types of construct validity were evaluated. Results. Scale structure of Haem-A-QoL was confirmed, with good item convergence (87%) and discrimination (80.6%) rates. Cronbach’s α was >0.70 for all but one dimension (dealing) and test-retest reliability was significantly high. The strength of Spearman’s correlations between Haem-A-QoL and SF-36 scales ranged from 0.25 to 0.75 (). Multiple stepwise linear regression analysis revealed that all but one Haem-A-QoL dimensions were important predictors of SF-36 scales. Known-groups comparisons yielded consistent support of the instruments’ construct validity and significant relationships were identified for age, educational level, haemophilia type, disease severity, and viral infections. Conclusion. Overall, the psychometric properties of the Greek version of Haem-A-QoL, resulting from this first time administration of the instrument to Greek adult haemophiliacs, confirmed it as a reliable and valid questionnaire for assessing haemophilia-specific HRQoL in Greece. Agoritsa Varaklioti, Nick Kontodimopoulos, Olga Katsarou, and Dimitris Niakas Copyright © 2014 Agoritsa Varaklioti et al. All rights reserved. Identification of a 20-Gene Expression-Based Risk Score as a Predictor of Clinical Outcome in Chronic Lymphocytic Leukemia Patients Mon, 05 May 2014 12:56:24 +0000 Despite the improvement in treatment options, chronic lymphocytic leukemia (CLL) remains an incurable disease and patients show a heterogeneous clinical course requiring therapy for many of them. In the current work, we have built a 20-gene expression (GE)-based risk score predictive for patients overall survival and improving risk classification using microarray gene expression data. GE-based risk score allowed identifying a high-risk group associated with a significant shorter overall survival (OS) and time to treatment (TTT) , comprising 19.6% and 13.6% of the patients in two independent cohorts. GE-based risk score, and NRIP1 and TCF7 gene expression remained independent prognostic factors using multivariate Cox analyses and combination of GE-based risk score together with NRIP1 and TCF7 gene expression enabled the identification of three clinically distinct groups of CLL patients. Therefore, this GE-based risk score represents a powerful tool for risk stratification and outcome prediction of CLL patients and could thus be used to guide clinical and therapeutic decisions prospectively. Elias Bou Samra, Bernard Klein, Thérèse Commes, and Jérôme Moreaux Copyright © 2014 Elias Bou Samra et al. All rights reserved. Immunohistochemical Analysis of IL-6, IL-8/CXCR2 Axis, -STAT-3, and SOCS-3 in Lymph Nodes from Patients with Chronic Lymphocytic Leukemia: Correlation between Microvascular Characteristics and Prognostic Significance Mon, 05 May 2014 11:06:00 +0000 A number of studies have looked into the pathophysiological role of angiogenesis in CLL, but the results have often been inconsistent. We aimed to gain direct insight into the angiogenic process in lymph nodes involved by CLL, focusing on proangiogenic cytokines and microvessel morphometry. The tissue levels of VEGF, Th-2 cytokines IL-6 and IL-8, IL-8 receptor CXCR2, and tyrosine p-STAT-3/SOCS-3 axis modulating cytokine expression were evaluated immunohistochemically in 62 CLL/SLL cases. Microvascular characteristics were evaluated by image analysis. Results were analyzed with regard to clinicopathological characteristics. Proliferation centers (PCs) were less well vascularised compared to non-PC areas. IL-8 and CXCR2 expression was distinctly uncommon as opposed to IL-6, VEGF and SOCS-3, which were detected in the vast majority of cases. The latter two molecule expressions were more pronounced in the PCs in ∼40% of the cases. p-STAT-3 immunoreactivity was recorded in 66.67% of the cases with a predilection for PCs. Microvessel morphometry was unrelated to proangiogenic cytokines, p-STAT-3, SOCS-3, or survival. Microvascular caliber and VEGF expression were higher in Binet stage A, whereasIL-6 expression was higher in stage C. VEGF and p-STAT-3 exerted a favorable effect on progression, which remained significant in multivariate analysis, thereby constituting potential outcome predictors in CLL patients. Georgia Levidou, Sotirios Sachanas, Gerassimos A. Pangalis, Christina Kalpadakis, Xanthi Yiakoumis, Maria Moschogiannis, Athanasia Sepsa, Eleftheria Lakiotaki, Vassilis Milionis, Marie-Christine Kyrtsonis, Theodoros P. Vassilakopoulos, Pantelis Tsirkinidis, Flora Kontopidou, Styliani Kokoris, Marina Siakantaris, Maria Angelopoulou, Helen Papadaki, Nikolaos Kavantzas, Panayiotis Panayiotidis, Efstratios Patsouris, and Penelope Korkolopoulou Copyright © 2014 Georgia Levidou et al. All rights reserved. Guidelines on Vaccinations in Paediatric Haematology and Oncology Patients Tue, 29 Apr 2014 07:50:08 +0000 Objective. Vaccinations are the most important tool to prevent infectious diseases. Chemotherapy-induced immune depression may impact the efficacy of vaccinations in children. Patients and Methods. A panel of experts of the supportive care working group of the Italian Association Paediatric Haematology Oncology (AIEOP) addressed this issue by guidelines on vaccinations in paediatric cancer patients. The literature published between 1980 and 2013 was reviewed. Results and Conclusion. During intensive chemotherapy, vaccination turned out to be effective for hepatitis A and B, whilst vaccinations with toxoid, protein subunits, or bacterial antigens should be postponed to the less intensive phases, to achieve an adequate immune response. Apart from varicella, the administration of live-attenuated-virus vaccines is not recommended during this phase. Family members should remain on recommended vaccination schedules, including toxoid, inactivated vaccine (also poliomyelitis), and live-attenuated vaccines (varicella, measles, mumps, and rubella). By the time of completion of chemotherapy, insufficient serum antibody levels for vaccine-preventable diseases have been reported, while immunological memory appears to be preserved. Once immunological recovery is completed, usually after 6 months, response to booster or vaccination is generally good and allows patients to be protected and also to contribute to herd immunity. Simone Cesaro, Mareva Giacchino, Francesca Fioredda, Angelica Barone, Laura Battisti, Stefania Bezzio, Stefano Frenos, Raffaella De Santis, Susanna Livadiotti, Serena Marinello, Andrea Giulio Zanazzo, and Désirée Caselli Copyright © 2014 Simone Cesaro et al. All rights reserved. The Impact of Hyperglycemia on Risk of Severe Infections during Early Period of Induction Therapy in Patients with Newly Diagnosed Multiple Myeloma Wed, 16 Apr 2014 16:46:34 +0000 The association between hyperglycemia and infections during induction chemotherapy has been reported in a number of hematologic disorders. This retrospective study evaluated the incidence of hyperglycemia during induction therapy in 155 patients with newly diagnosed multiple myeloma (MM) and its effect on serious infections during the first 60 days of induction. A total of 20 (12.9%) patients developed overt hyperglycemia (≥200 mg/dL) during induction therapy. Serious infections occurred in 28 (18.1%) of 155 patients and infection-related mortality within 2 months after treatment was 0.6% (1 patient). In a univariate analysis, overt hyperglycemia, poor performance status (≥2), International Staging System III, lymphopenia (<500/μL), and elevated serum creatinine (≥2 mg/dL) were found to be associated with serious infections. In multivariate analysis, only overt hyperglycemia (HR 7.846, 95% CI 2.512–24.503, ) and poor performance status (HR 5.801, 95% CI 1.974–17.050, ) remained significant. In conclusion, this study demonstrated an association between hyperglycemia and serious infections during induction therapy in patients with MM. Sung-Hoon Jung, Hee-Chang Jang, Seung-Shin Lee, Jae-Sook Ahn, Deok-Hwan Yang, Yeo-Kyeoung Kim, Hyeoung-Joon Kim, and Je-Jung Lee Copyright © 2014 Sung-Hoon Jung et al. All rights reserved. Nutritional Biomarkers in Children and Adolescents with Beta-Thalassemia-Major: An Egyptian Center Experience Tue, 08 Apr 2014 07:04:44 +0000 Background and Aim. Trace elements and vitamins play a vital role in human body to perform its function properly. Thalassemic patients are at risk of micronutrient deficiency. This study estimated levels of vitamins A, C, E, B12, folic acid, total homocysteine (tHcy), and methylmalonic acid (MMA) along with trace elements, zinc, copper, and selenium in Beta-thalassemia-major patients. Methods. This study included 108 patients with Beta-thalassemia-major and 60 age and sex matched healthy children. Serum levels of vitamin A, E, C, tHcy, and MMA were estimated by high pressure liquid chromatography while serum levels of folic acid and B12 were estimated by thin layer chromatography. Serum zinc, copper, and selenium were determined by atomic absorption spectrometry. Results. There was a significant decrease of vitamins A, C, E, and B12 and trace elements zinc, copper, and selenium in thalassemic patients as compared to controls. tHcy and MMA were significantly elevated in patients. No significant correlations were found between the serum levels of the studied vitamins and trace elements as regards age, frequency of transfusion, duration of transfusion, and serum ferritin. Conclusion. The level of various nutritional biomarkers (vitamins A, C, E, and B12 and trace elements zinc, copper, selenium) was reduced in chronically transfused Egyptian thalassemic patient. These patients should have periodic nutritional evaluation and supplementation. Multicenter studies are highly recommended. Laila M. Sherief, Sanaa M. Abd El-Salam, Naglaa M. Kamal, Osama El safy, Mohamed A. A. Almalky, Seham F. Azab, Hemat M. Morsy, and Amal F. Gharieb Copyright © 2014 Laila M. Sherief et al. All rights reserved. Mutation Status and Immunoglobulin Gene Rearrangements in Patients from Northwest and Central Region of Spain with Chronic Lymphocytic Leukemia Sun, 30 Mar 2014 07:21:48 +0000 The aim of this study was to investigate the frequency and mutation status of the immunoglobulin heavy variable chain (IGHV) in a cohort of 224 patients from northwest and central region of Spain diagnosed with chronic lymphocytic leukemia (CLL), and to correlate it with cytogenetic abnormalities, overall survival (OS) and time to first treatment (TTFT). 125 patients had mutated IGHV, while 99 had unmutated IGHV. The most frequently used IGHV family was IGHV3, followed by IGHV1 and IGHV4. The regions IGHV3-30, IGHV1-69, IGHV3-23, and IGHV4-34 were the most commonly used. Only 3.1% of the patients belonged to the subfamily IGHV3-21 and we failed to demonstrate a worse clinical outcome in this subgroup. The IGHV4 family appeared more frequently with mutated pattern, similar to IGHV3-23 and IGHV3-74. By contrast, IGHV1-69 was expressed at a higher frequency in unmutated CLL patients. All the cases from IGHV3-11 and almost all from IGHV5-51 subfamily belonged to the group of unmutated CLL. I. González-Gascón y Marín, J. A. Hernández, A. Martín, M. Alcoceba, M. E. Sarasquete, A. Rodríguez-Vicente, C. Heras, N. de las Heras, R. Fisac, A. García de Coca, I. de la Fuente, M. Hernández-Sánchez, I. Recio, J. Galende, G. Martín-Núñez, J. M. Alonso, J. M. Hernández-Rivas, and M. González Copyright © 2014 I. González-Gascón y Marín et al. All rights reserved. β-Thalassemia Intermedia in Northern Iraq: A Single Center Experience Thu, 27 Feb 2014 08:16:26 +0000 To investigate the molecular basis of β-thalassemia intermedia in Northern Iraq and evaluate its management practices, a total of 74 patients from 51 families were enrolled. The patients were clinically and hematologically reevaluated, and had their β-thalassemia mutations characterized, as well as the number of α-globin genes and Xmn I Gγ −158 (C>T) polymorphism studied. Out of 14 β-thalassemia mutations identified, the four most common were IVS-I-6 (T>C) [33.3%], IVS-II-I (G>A) [21.1%], codon 82/83(−G) [10.1%], and codon 8 (−AA) [8.1%]. The most common contributing factors to the less severe phenotype of thalassemia intermedia were found to be the inheritance of mild β-thalassemia alleles and the Xmn I polymorphism, while concomitant α-thalassemia had a limited role. Several complications were documented including: pulmonary hypertension in 20.4%, diabetes mellitus in 1.4%, hypothyroidism in 2.9%, and heart failure in 2.7%, while no documented cases of venous thrombosis were found. Compared to their counterparts in several Mediterranean countries, it appears that our patients were much less frequently transfused and had a lower proportion of patients who were splenectomized, on iron chelation, or hydroxycarbamide therapy. Such practices require further scrutiny to ensure that a better level of care is provided and that growth retardation, skeletal changes, and other complications are prevented or reduced. Nasir A. S. Al-Allawi, Sana D. Jalal, Ameen M. Mohammad, Sharaza Q. Omer, and Raji S. D. Markous Copyright © 2014 Nasir A. S. Al-Allawi et al. All rights reserved. Autologous Stem Cell Transplantation in Elderly Patients with Multiple Myeloma: Past, Present, and Future Thu, 20 Feb 2014 07:48:56 +0000 High-dose melphalan (200 mg/m2) as conditioning regimen followed by autologous stem cell transplantation (ASCT) rescue has been established as a standard treatment for patients with multiple myeloma (MM) younger than 65 years of age. However, the role of ASCT in elderly patients older than 65 years remains controversial in the era of novel agents such as thalidomide, bortezomib, and lenalidomide. The efficacy and feasibility of ASCT have been shown in elderly patients by reducing the dose of melphalan to 100–140 mg/m2. Although the clinical benefit of reduced-intensity ASCT in elderly patients has not been clearly established in comparison with that of novel agent-based induction therapy, recent studies have demonstrated that sequential strategies of novel agent-based induction therapy and reduced-intensity ASCT followed by consolidation/maintenance with novel agents translate into better outcome in the management of elderly patients. Thus, ASCT could also be a mainstay in the initial treatment of elderly MM patients, and its indication should be evaluated based on performance status and the presence of complications and/or comorbidities of each elderly patient with MM. Shuji Ozaki and Kazuyuki Shimizu Copyright © 2014 Shuji Ozaki and Kazuyuki Shimizu. All rights reserved. TET2 Overexpression in Chronic Lymphocytic Leukemia Is Unrelated to the Presence of TET2 Variations Tue, 18 Feb 2014 16:35:32 +0000 TET2 is involved in a variety of hematopoietic malignancies, mainly in myeloid malignancies. Most mutations of TET2 have been identified in myeloid disorders, but some have also recently been described in mature lymphoid neoplasms. In contrast to the large amount of data about mutations of TET2, some data are available for gene expression. Moreover, the role of TET2 in chronic lymphocytic leukemia (CLL) is unknown. This study analyzes both TET2 expression and mutations in 48 CLL patients. TET2 expression was analyzed by exon arrays and quantitative real-time polymerase chain reaction (qRT-PCR). Next-generation sequencing (NGS) technology was applied to investigate the presence of TET2 variations. Overexpression of TET2 was observed in B-cell lymphocytes from CLL patients compared with healthy donors (P = 0.004). In addition, in CLL patients, an overexpression of TET2 was also observed in the clonal B cells compared with the nontumoral cells (P = 0.002). However, no novel mutations were observed. Therefore, overexpression of TET2 in CLL seems to be unrelated to the presence of genomic TET2 variations. María Hernández-Sánchez, Ana Eugenia Rodríguez, Alexander Kohlmann, Rocío Benito, Juan Luis García, Alberto Risueño, Encarna Fermiñán, Javier De Las Rivas, Marcos González, and Jesús-María Hernández-Rivas Copyright © 2014 María Hernández-Sánchez et al. All rights reserved. Sphingosine-1-phosphate-Mediated Mobilization of Hematopoietic Stem/Progenitor Cells during Intravascular Hemolysis Requires Attenuation of SDF-1-CXCR4 Retention Signaling in Bone Marrow Sun, 29 Dec 2013 13:37:52 +0000 Sphingosine-1-phosphate (S1P) is a crucial chemotactic factor in peripheral blood (PB) involved in the mobilization process and egress of hematopoietic stem/progenitor cells (HSPCs) from bone marrow (BM). Since S1P is present at high levels in erythrocytes, one might assume that, by increasing the plasma S1P level, the hemolysis of red blood cells would induce mobilization of HSPCs. To test this assumption, we induced hemolysis in mice by employing phenylhydrazine (PHZ). We observed that doubling the S1P level in PB from damaged erythrocytes induced only a marginally increased level of mobilization. However, if mice were exposed to PHZ together with the CXCR4 blocking agent, AMD3100, a robust synergistic increase in the number of mobilized HSPCs occurred. We conclude that hemolysis, even if it significantly elevates the S1P level in PB, also requires attenuation of the CXCR4-SDF-1 axis-mediated retention in BM niches for HSPC mobilization to occur. Our data also further confirm that S1P is a major chemottractant present in plasma and chemoattracts HSPCs into PB under steady-state conditions. However, to egress from BM, HSPCs first have to be released from BM niches by blocking the SDF-1-CXCR4 retention signal. Kasia Mierzejewska, Yuri M. Klyachkin, Janina Ratajczak, Ahmed Abdel-Latif, Magda Kucia, and Mariusz Z. Ratajczak Copyright © 2013 Kasia Mierzejewska et al. All rights reserved. Asthma Management in Sickle Cell Disease Sun, 10 Nov 2013 10:24:43 +0000 Asthma is a common comorbid factor in sickle cell disease (SCD). However, the incidence of asthma in SCD is much higher than expected compared to rates in the general population. Whether “asthma” in SCD is purely related to genetic and environmental factors or rather is the consequence of the underlying hemolytic and inflammatory state is a topic of recent debate. Regardless of the etiology, hypoxemia induced by bronchoconstriction and inflammation associated with asthma exacerbations will contribute to a cycle of sickling and subsequent complications of SCD. Recent studies confirm that asthma predisposes to complications of SCD such as pain crises, acute chest syndrome, and stroke and is associated with increased mortality. Early recognition and aggressive standard of care management of asthma may prevent serious pulmonary complications and reduce mortality. However, data regarding the management of asthma in SCD is very limited. Clinical trials are needed to evaluate the effectiveness of current asthma therapy in patients with SCD and coincident asthma, while mechanistic studies are needed to delineate the underlying pathophysiology. Esteban Gomez and Claudia R. Morris Copyright © 2013 Esteban Gomez and Claudia R. Morris. All rights reserved. UGT1A1 Gene Mutation due to Crigler-Najjar Syndrome in Iranian Patients: Identification of a Novel Mutation Mon, 28 Oct 2013 15:11:20 +0000 Crigler-Najjar syndrome (CNS) type I and type II are inherited as autosomal recessive conditions that are caused by mutations in the UGT1A1 gene. We present the analysis of UGT1A1 gene in 12 individuals from three different families. This analysis allowed us to identify one novel mutation, which was not previously described. In this study, three families with clinically diagnosed CNS referred from Khuzestan province, southwest Iran, were screened. After signing the informed consent, peripheral blood samples from the patients and their parents were collected in EDTA-containing tube followed by DNA extraction using a routine phenol-chloroform method. All five coding exons and the flanking intronic regions of the bilirubin-UGT were amplified by polymerase chain reaction (PCR) followed by DNA sequencing by Sanger method. From the first family, a 9-month-old boy was homozygous for a deletion mutation of two adjacent nucleotides including one adenosine (A) and one glutamine (G) between nucleotides 238 and 239 in exon 1 (c.238_240 del AG). In the second family, there were two affected individuals, an 11-year-old girl and a fetus, found to be homozygous for the same mutation. The third family showed a mutation at nucleotide 479 in exon 1 (Val160Glu) that has been reported previously. Molecular analysis can significantly help confirm the diagnosis of CNS, without any need for the liver biopsy, and may help the therapeutic management by ruling out more harmful causes of hyperbilirubinemia. Javad Mohammadi Asl, Mohammad Amin Tabatabaiefar, Hamid Galehdari, Kourosh Riahi, Mohammad Hosein Masbi, Zohre Zargar Shoshtari, and Fakher Rahim Copyright © 2013 Javad Mohammadi Asl et al. All rights reserved. A Pipeline with Multiplex Reverse Transcription Polymerase Chain Reaction and Microarray for Screening of Chromosomal Translocations in Leukemia Tue, 08 Oct 2013 09:25:10 +0000 Chromosome rearrangements and fusion genes present major portion of leukemogenesis and contribute to leukemic subtypes. It is practical and helpful to detect the fusion genes in clinic diagnosis of leukemia. Present application of reverse transcription polymerase chain reaction (RT-PCR) method to detect the fusion gene transcripts is effective, but time- and labor-consuming. To set up a simple and rapid system, we established a method that combined multiplex RT-PCR and microarray. We selected 15 clinically most frequently observed chromosomal rearrangements generating more than 50 fusion gene variants. Chimeric reverse primers and chimeric PCR primers containing both gene-specific and universal sequences were applied in the procedure of multiplex RT-PCR, and then the PCR products hybridized with a designed microarray. With this approach, among 200 clinic samples, 63 samples were detected to have gene rearrangements. All the detected fusion genes positive and negative were validated with RT-PCR and Sanger sequencing. Our data suggested that the RT-PCR-microarray pipeline could screen 15 partner gene pairs simultaneously at the same accuracy of the fusion gene detection with regular RT-PCR. The pipeline showed effectiveness in multiple fusion genes screening in clinic samples. Fei-Fei Xiong, Ben-Shang Li, Chun-Xiu Zhang, Hui Xiong, Shu-Hong Shen, and Qing-Hua Zhang Copyright © 2013 Fei-Fei Xiong et al. All rights reserved. Comparison of Stress-Hemoconcentration Correction Techniques for Stress-Induced Coagulation Mon, 07 Oct 2013 10:19:03 +0000 When examining stress effects on coagulation, arithmetic correction is typically used to adjust for concomitant hemoconcentration but may be inappropriate for coagulation activity assays. We examined a new physiologically relevant method of correcting for stress-hemoconcentration. Blood was drawn from healthy men () during baseline, mental stress, and recovery, and factor VII activity (FVII:C), factor VIII activity (FVIII:C), activated partial thromboplastin time (APTT), prothrombin time (PT%), fibrinogen, D-dimer, and plasma volume were determined. Three hemoconcentration correction techniques were assessed: arithmetic correction and two reconstitution techniques using baseline plasma or physiological saline. Area-under-the-curve (AUC) was computed for each technique. For FVII:C, uncorrected AUC was significantly greater than AUC corrected arithmetically. For PT%, uncorrected AUC was significantly greater than AUC corrected with saline or arithmetically. For APTT, uncorrected AUC was significantly less than AUC corrected with saline and greater than AUC corrected arithmetically. For fibrinogen, uncorrected AUC was significantly greater than AUC corrected with saline or arithmetically. For D-dimer, uncorrected AUC was significantly greater than AUC corrected arithmetically. No differences in AUC were observed for FVIII:C. Saline reconstitution seems most appropriate when adjusting for hemoconcentration effects on clotting time and activity. Stress-hemoconcentration accounted for the majority of coagulation changes. Anthony W. Austin and Stephen M. Patterson Copyright © 2013 Anthony W. Austin and Stephen M. Patterson. All rights reserved. The Fc Receptor Polymorphisms and Expression of Neutrophil Activation Markers in Patients with Sickle Cell Disease from Western India Sun, 29 Sep 2013 14:49:26 +0000 Objective. Sickle cell disease has variable clinical manifestations. Activation of neutrophils plays an important role in the initiation and propagation of vaso occlusive crises which can be analysed by determining the expression of neutrophil antigens such as CD16, CD32, and CD62L. The common FcγR polymorphisms (FcγRIIA and FcγRIIIB) are considered to influence clinical presentation. This study focuses on distribution of FcγR polymorphisms and their association with neutrophil activity among the patients from western India. Methods. In this paper 127 sickle cell anemia patients and 58 patients with sickle-β-thalassemia (median age  years) with variable clinical phenotypes along with 175 normals were investigated. FcγRs polymorphisms were analysed by RFLP and AS-PCR. Activation of neutrophils was measured by flow cytometry. Results. The genotypic frequency of the H/R genotype of FcγRIIA and the NA1/NA1 genotype of FcγRIIIB was significantly decreased in patients compared to normals (-0.0074, -0.0471, resp.). We found a significant difference in the expression of CD32 and CD62L among the patients as against normals. A significantly higher expression of CD32 was seen in the milder patients with the H/H genotype (-0.0231), whereas the expression of CD16 was higher in severe patients with the NA2/NA2 genotype (-0.0312). Conclusion. The two FcγR polymorphisms had significant association with variable phenotypes of sickle cell disease. The expression of CD62L decreased in our patients indicating activation of neutrophils. Harshada K. Kangne, Farah F. Jijina, Yazdi M. Italia, Dipti L. Jain, Anita H. Nadkarni, Maya Gupta, Vandana Pradhan, Rati D. Mukesh, Kanjaksha K. Ghosh, and Roshan B. Colah Copyright © 2013 Harshada K. Kangne et al. All rights reserved. Lenalidomide and Chronic Lymphocytic Leukemia Thu, 19 Sep 2013 11:13:10 +0000 Lenalidomide is an oral immunomodulatory drug used in multiple myeloma and myelodysplastic syndrome and most recently it has shown to be effective in the treatment of various lymphoproliferative disorders such as chronic lymphocytic leukemia (CLL) and non-Hodgkin lymphoma. The mechanism of action of lenalidomide varies depending on the pathology, and in the case of CLL, it appears to primarily act by restoring the damaged mechanisms of tumour immunosurveillance. This review discusses the potential mechanism of action and efficacy of lenalidomide, alone or in combination, in treatment of CLL and its toxic effects such as tumor lysis syndrome (TLS) and tumor flare reaction (TFR), that make its management different from other hematologic malignancies. Ana Pilar González-Rodríguez, Angel R. Payer, Andrea Acebes-Huerta, Leticia Huergo-Zapico, Monica Villa-Alvarez, Esther Gonzalez-García, and Segundo Gonzalez Copyright © 2013 Ana Pilar González-Rodríguez et al. All rights reserved. TNFα Mediated IL-6 Secretion Is Regulated by JAK/STAT Pathway but Not by MEK Phosphorylation and AKT Phosphorylation in U266 Multiple Myeloma Cells Mon, 16 Sep 2013 15:02:01 +0000 IL-6 and TNFα were significantly increased in the bone marrow aspirate samples of patients with active multiple myeloma (MM) compared to those of normal controls. Furthermore, MM patients with advanced aggressive disease had significantly higher levels of IL-6 and TNFα than those with MM in plateau phase. TNFα increased interleukin-6 (IL-6) production from MM cells. However, the detailed mechanisms involved in signaling pathways by which TNFα promotes IL-6 secretion from MM cells are largely unknown. In our study, we found that TNFα treatments induce MEK and AKT phosphorylation. TNFα-stimulated IL-6 production was abolished by inhibition of JAK2 and IKKβ or by small interfering RNA (siRNA) targeting TNF receptors (TNFR) but not by MEK, p38, and PI3K inhibitors. Also, TNFα increased phosphorylation of STAT3 (ser727) including c-Myc and cyclin D1. Three different types of JAK inhibitors decreased the activation of the previously mentioned pathways. In conclusion, blockage of JAK/STAT-mediated NF-κB activation was highly effective in controlling the growth of MM cells and, consequently, an inhibitor of TNFα-mediated IL-6 secretion would be a potential new therapeutic agent for patients with multiple myeloma. Chansu Lee, Jeong-In Oh, Juwon Park, Jee-Hye Choi, Eun-Kyung Bae, Hyun Jung Lee, Woo June Jung, Dong Soon Lee, Kwang-Sung Ahn, and Sung-Soo Yoon Copyright © 2013 Chansu Lee et al. All rights reserved. TLR Stimulation of Bone Marrow Lymphoid Precursors from Childhood Acute Leukemia Modifies Their Differentiation Potentials Tue, 10 Sep 2013 08:32:33 +0000 Acute leukemias are the most frequent childhood malignancies worldwide and remain a leading cause of morbidity and mortality of relapsed patients. While remarkable progress has been made in characterizing genetic aberrations that may control these hematological disorders, it has also become clear that abnormalities in the bone marrow microenvironment might hit precursor cells and contribute to disease. However, responses of leukemic precursor cells to inflammatory conditions or microbial components upon infection are yet unexplored. Our previous work and increasing evidence indicate that Toll-like receptors (TLRs) in the earliest stages of lymphoid development in mice and humans provide an important mechanism for producing cells of the innate immune system. Using highly controlled co-culture systems, we now show that lymphoid precursors from leukemic bone marrow express TLRs and respond to their ligation by changing cell differentiation patterns. While no apparent contribution of TLR signals to tumor progression was recorded for any of the investigated diseases, the replenishment of innate cells was consistently promoted upon in vitro TLR exposure, suggesting that early recognition of pathogen-associated molecules might be implicated in the regulation of hematopoietic cell fate decisions in childhood acute leukemia. Elisa Dorantes-Acosta, Eduardo Vadillo, Adriana Contreras-Quiroz, Juan Carlos Balandrán, Lourdes Arriaga-Pizano, Jessica Purizaca, Sara Huerta-Yepez, Elva Jiménez, Wendy Aguilera, Aurora Medina-Sanson, Héctor Mayani, and Rosana Pelayo Copyright © 2013 Elisa Dorantes-Acosta et al. All rights reserved. Impact of Interstitial Pneumonia on the Survival and Risk Factors Analysis of Patients with Hematological Malignancy Wed, 04 Sep 2013 11:38:24 +0000 Background. The emergence of interstitial pneumonia (IP) in patients with hematological malignancy (HM) is becoming a challenging scenario in current practice. However, detailed characterization and investigation of outcomes and risk factors on survival have not been addressed. Methods. We conducted a retrospective study of 42,584 cancer patients covering the period between 1996 and 2008 using the institutional cancer registry system. Among 816 HM patients, 61 patients with IP were recognized. The clinical features, laboratory results, and histological types were studied to determine the impact of IP on survival and identify the profile of prognostic factors. Results. HM patients with IP showed a significant worse survival than those without IP in the 5-year overall survival (). The overall survival showed no significant difference between infectious pneumonia and noninfectious interstitial pneumonia (IIP versus nIIP) (). In a multivariate Cox regression model, leukocyte and platelet count were associated with increased risk of death. Conclusions. The occurrence of IP in HM patients is associated with increased mortality. Of interest, nIIP is a prognostic indicator in patients with lymphoma but not in patients with leukemia. However, aggressive management of IP in patients with HM is strongly advised, and further prospective survey is warranted. Wei-Liang Chen, Yu-Tzu Tsao, Tsun-Hou Chang, Tsu-Yi Chao, Woei-Yau Kao, Yeu Chin Chen, and Ching-Liang Ho Copyright © 2013 Wei-Liang Chen et al. All rights reserved. Analysis of the Magnetic Field Influence on the Rheological Properties of Healthy Persons Blood Mon, 02 Sep 2013 13:18:48 +0000 The influence of magnetic field on whole blood rheological properties remains a weakly known phenomenon. An in vitro analysis of the magnetic field influence on the rheological properties of healthy persons blood is presented in this work. The study was performed on blood samples taken from 25 healthy nonsmoking persons and included comparative analysis of the results of both the standard rotary method (flow curve measurement) and the oscillatory method known also as the mechanical dynamic analysis, performed before and after exposition of blood samples to magnetic field. The principle of the oscillatory technique lies in determining the amplitude and phase of the oscillations of the studied sample subjected to action of a harmonic force of controlled amplitude and frequency. The flow curve measurement involved determining the shear rate dependence of blood viscosity. The viscoelastic properties of the blood samples were analyzed in terms of complex blood viscosity. All the measurements have been performed by means of the Contraves LS40 rheometer. The data obtained from the flow curve measurements complemented by hematocrit and plasma viscosity measurements have been analyzed using the rheological model of Quemada. No significant changes of the studied rheological parameters have been found. Anna Marcinkowska-Gapinska and Honorata Nawrocka-Bogusz Copyright © 2013 Anna Marcinkowska-Gapinska and Honorata Nawrocka-Bogusz. All rights reserved. VEGF and bFGF Gene Polymorphisms in Patients with Non-Hodgkin's Lymphoma Mon, 12 Aug 2013 09:41:26 +0000 Angiogenesis and lymphangiogenesis are important in the proliferation and survival of the malignant hematopoietic neoplasms, including non-Hodgkin’s lymphomas (NHLs). Vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) play an important role in the initiation of angiogenesis. Both VEGF and bFGF have been reported to have prognostic significance in NHL. The present study aimed to determine an association between the VEGF and bFGF gene polymorphisms and disease susceptibility and progression. VEGF (rs3025039; 936 C>T) and bFGF (rs308395, −921 G>C) variants were determined in 78 NHL patients and 122 healthy individuals by PCR-RFLP technique. The presence of the VEGF 936T allele was found to significantly associate with worse prognosis of the disease (expressed by the highest International Prognostic Index (IPI)) (0.41 versus 0.20, for IPI 4 among patients having and lacking the T allele). The VEGF 936T variant was also more frequent among patients with IPI 4 than in controls (OR = 3.37, ). The bFGF −921G variant was more frequently detected among patients with aggressive as compared to those with indolent histological subtype (0.37 versus 0.18, ) and healthy individuals (0.37 versus 0.19, OR = 2.51, ). These results imply that VEGF and bFGF gene polymorphisms have prognostic significance in patients with NHL. Tomasz Wróbel, Grzegorz Mazur, Justyna Dzietczenia, Katarzyna Gębura, Kazimierz Kuliczkowski, and Katarzyna Bogunia-Kubik Copyright © 2013 Tomasz Wróbel et al. All rights reserved. Relationship between Circulating BAFF Serum Levels with Proliferating Markers in Patients with Multiple Myeloma Mon, 15 Jul 2013 11:06:25 +0000 In multiple myeloma, there are many factors influencing the growth of the malignant clone in direct and indirect manners. BAFF is a growth factor for myeloma cells. The aim of the study was to measure its circulating levels in 54 pretreatment patients, along with serum levels of other proliferation markers, such as interleukins-6, -10, and -15, CRP, and beta-2 microglobulin, as well as bone marrow plasma cell infiltration and expression of Ki-67 PI, in various stages of the disease and after effective treatment in 28 of them. Serum levels of the previously mentioned factors were measured by ELISA, whereas bone marrow plasma cell infiltration and Ki-67 expression were estimated immunohistochemically. All measured parameters were higher in pretreated myeloma patients compared to healthy population and were also increasing with the progression of the disease. They all also decreased after effective therapy. Furthermore, all pretreatment values correlated to each other. BAFF seems to be an important growth factor for myeloma plasma cells. Measuring its serum levels, along with the previously mentioned cytokines, may provide important information regarding the degree of myeloma cells’ proliferation. Therefore, they all could be used as markers of proliferation and disease activity. Michael G. Alexandrakis, Parascevi Roussou, Constantina A. Pappa, Ippokratis Messaritakis, Athina Xekalou, Nektaria Goulidaki, Anna Boula, and George Tsirakis Copyright © 2013 Michael G. Alexandrakis et al. All rights reserved. Human Platelet Antigen Alleles in 998 Taiwanese Blood Donors Determined by Sequence-Specific Primer Polymerase Chain Reaction Thu, 20 Jun 2013 09:12:01 +0000 Polymorphism of human platelet antigens (HPAs) leads to alloimmunizations and immune-mediated platelet disorders including fetal-neonatal alloimmune thrombocytopenia (FNAIT), posttransfusion purpura (PTP), and platelet transfusion refractoriness (PTR). HPA typing and knowledge of antigen frequency in a population are important in particular for the provision of HPA-matched blood components for patients with PTR. We have performed allele genotyping for HPA-1 through -6 and -15 among 998 platelet donors from 6 blood centers in Taiwan using sequence-specific primer polymerase chain reaction. The HPA allele frequency was 99.55, and 0.45% for HPA-1a and -1b; 96.49, and 3.51% for HPA-2a and -2b; 55.81, and 44.19% for HPA-3a and -3b; 99.75, and 0.25% for HPA-4a and -4b; 98.50, and 1.50% for HPA-5a and -5b; 97.75 and 2.25% for HPA-6a and -6b; 53.71 and 46.29% for HPA-15a and -15b. HPA-15b and HPA-3a, may be considered the most important, followed by HPA-2, -6, -1, -5, and -4 systems, as a cause of FNAIT, PTP, and PTR based on allele frequency. HPA-4b and HPA-5b role cannot be excluded based on their immunogenicity. A larger-scale study will now be conducted to confirm these hypotheses and to establish an apheresis donor database for the procurement of HPA-matched apheresis platelets for patients with PTR. Shun-Chung Pai, Thierry Burnouf, Jen-Wei Chen, and Liang-In Lin Copyright © 2013 Shun-Chung Pai et al. All rights reserved. Genetic Epidemiology, Hematological and Clinical Features of Hemoglobinopathies in Iran Tue, 18 Jun 2013 14:16:28 +0000 There is large variation in the molecular genetics and clinical features of hemoglobinopathies in Iran. Studying structural variants of hemoglobin demonstrated that the β-chain variants of hemoglobin S and D-Punjab are more prevalent in the Fars (southwestern Iran) and Kermanshah (western Iran) provinces, respectively. Also, α-chain variants of Hb Q-Iran and Hb Setif are prevalent in western Iran. The molecular basis and clinical severity of thalassemias are extremely heterogenous among Iranians due to the presence of multiethnic groups in the country. β-Thalassemia is more prevalent in northern and southern Iran. Among 52 different β-thalassemia mutations that have been identified among Iranian populations, IVSII-1 G:A is the most frequent mutation in most parts of the country. The presence of IVS I-5 G:C mutation with high frequency in southeastern Iran might reflect gene flow from neighboring countries. A wide spectrum of α-thalassemia alleles has been detected among Iranians with as the most prevalent α-thalassemia mutation. The prevention program of thalassemia birth in Iran has reduced the birth rate of homozygous β-thalassemia since the implementation of the program in 1997. In this review genetic epidemiology, clinical and hematological aspects of hemoglobinopathies, and the prevention programs of β-thalassemia in Iran will be discussed. Zohreh Rahimi Copyright © 2013 Zohreh Rahimi. All rights reserved. Acute Toxicity and the Effect of Andrographolide on Porphyromonas gingivalis-Induced Hyperlipidemia in Rats Thu, 13 Jun 2013 11:23:56 +0000 The aim of the current study is to evaluate the effect of andrographolide on hyperlipidemia induced by Porphyromonas gingivalis in rats. Thirty male Sprague Dawley (SD) rats were divided into five groups as follows: group 1 (vehicle) and four experimental groups (groups 2, 3, 4, and 5) were challenged orally with P. gingivalis ATCC 33277 (0.2 mL of bacterial cells/mL in 2% carboxymethylcellulose (CMC) with phosphate-buffered saline (PBS)) five times a week for one month to induce hyperlipidemia. Then, group 3 received a standard oral treatment with simvastatin 100 mg/kg, and groups 4 and 5 received oral treatment with andrographolide 20 mg/kg and 10 mg/kg, respectively, for another month. The results showed that total cholesterol (TC), low-density lipoprotein (LDL-C), and triglycerides (TG) were reduced significantly in groups treated with andrographolide. The malondialdehyde (MDA) level was low in treated groups, while antioxidant enzymes, superoxide dismutase (SOD), and glutathione peroxidase (GPx) were significantly increased in these groups (). Liver tissues of the groups treated with andrographolide reduce the accumulation of lipid droplets in hepatic tissue cells. An acute toxicity test did not show any toxicological symptoms in rats. Rami Al Batran, Fouad Al-Bayaty, Mazen M. Jamil Al-Obaidi, and Mahmood A. Abdulla Copyright © 2013 Rami Al Batran et al. All rights reserved. TET2 Mutations in Ph-Negative Myeloproliferative Neoplasms: Identification of Three Novel Mutations and Relationship with Clinical and Laboratory Findings Sat, 25 May 2013 13:50:29 +0000 High-throughput DNA sequence analysis was used to screen for TET2 mutations in peripheral blood derived DNA from 97 patients with BCR-ABL-negative myeloproliferative neoplasms (MPNs). Overall six mutations in the coding region of the gene were identified in 7 patients with an overall mutational frequency of 7.2%. In polycythemia vera patients () 2 mutations were identified (8%), and in those with essential thrombocythemia () 2 mutations (3.6%); in those with unclassifiable MPN () 3 mutations (37.5%). No primary myelofibrosis patients () harboured TET2 mutations. Three unreported mutations were identified (p.P177fs, p.C1298del, and p.P411del), the first two in patients with unclassifiable MPN, the last in a patient with essential thrombocythemia. On multivariate analysis the diagnosis of an unclassifiable MPN was significantly related to the presence of TET2 mutations (; OR: 2.81; 95% CI 1.11–7.06). We conclude that TET2 mutations occur in both JAK2 V617F-positive and -negative MPNs and are more frequent in MPN-U patients. This could represent the biological link between the different classes of myeloid malignancies. Andrea Patriarca, Donatella Colaizzo, Gianluca Tiscia, Raffaele Spadano, Silvia Di Zacomo, Antonio Spadano, Ida Villanova, Maurizio Margaglione, Elvira Grandone, and Alfredo Dragani Copyright © 2013 Andrea Patriarca et al. All rights reserved. Soluble Flt-1 Gene Delivery in Acute Myeloid Leukemic Cells Mediating a Nonviral Gene Carrier Thu, 10 Jan 2013 16:12:23 +0000 Vascular endothelial growth factor (VEGF) is a potent angiogenic factor involved in angiogenesis-mediated progression of acute myeloid leukemia (AML). Studies have reported the role of soluble form of fms-like tyrosine kinase (sFlT-1) delivery as an antitumor agent by inhibiting VEGF. This study investigates the outcome of delivery of a VEGF165 antagonist, soluble vascular endothelial growth factor receptor, namely sFLT-1, mediating lipofectamine 2000 in acute myeloid leukemic cells. A recombinant plasmid expressing sFLT-1 was constructed and transfected into the K562 and HL60 cells using lipofectamine 2000 transfection reagent. sFLT-1 expression/secretion in pVAX-sFLT-1 transfected cells was verified by RT-PCR and western blot. MTS assay was carried out to evaluate the effect of sFLT-1 on human umbilical vein endothelial cells and K562 and HL60 cells in vitro. Treatment with pVAX-sFLT-1 showed no association between sFLT-1 and proliferation of infected K562 and HL60 cells, while it demonstrated a significant inhibitory impact on the proliferation of HUVECs. The results of the current study imply that the combination of nonviral gene carrier and sFLT-1 possesses the potential to provide efficient tool for the antiangiogenic gene therapy of AML. Razieh Amini, Farid Azizi Jalilian, Abhi Veerakumarasivam, Syahril Abdullah, Ahmed S. Abdulamir, Fatemeh Nadali, and Rozita Rosli Copyright © 2013 Razieh Amini et al. All rights reserved. Insights and Hopes in Umbilical Cord Blood Stem Cell Transplantations Wed, 31 Oct 2012 15:11:09 +0000 Over 20.000 umblical cord blood transplantations (UCBT) have been carried out around the world. Indeed, UCBT represents an attractive source of donor hematopoietic stem cells (HSCs) and, offer interesting features (e.g., lower graft-versus-host disease) compared to bone marrow transplantation (BMT). Thereby, UCBT often represents the unique curative option against several blood diseases. Recent advances in the field of UCBT, consisted to develop strategies to expand umbilical stem cells and shorter the timing of their engraftment, subsequently enhancing their availability for enhanced efficacy of transplantation into indicated patients with malignant diseases (e.g., leukemia) or non-malignant diseases (e.g., thalassemia major). Several studies showed that the expansion and homing of UCBSCs depends on specific biological factors and cell types (e.g., cytokines, neuropeptides, co-culture with stromal cells). In this review, we extensively present the advantages and disadvantages of current hematopoietic stem cell transplantations (HSCTs), compared to UBCT. We further describe the importance of cord blood content and obstetric factors on cord blood selection, and report the recent approaches that can be undertook to improve cord blood stem cell expansion as well as engraftment. Eventually, we provide two majors examples underlining the importance of UCBT as a potential cure for blood diseases. Somayeh Shahrokhi, Farid Menaa, Kamran Alimoghaddam, Colin McGuckin, and Massoumeh Ebtekar Copyright © 2012 Somayeh Shahrokhi et al. All rights reserved. Umbilical Cord-Derived Mesenchymal Stem Cells for Hematopoietic Stem Cell Transplantation Wed, 03 Oct 2012 08:53:11 +0000 Hematopoietic stem cell transplantation (HSCT) is becoming an effective therapeutic modality for a variety of diseases. Mesenchymal stem cells (MSCs) can be used to enhance hematopoietic engraftment, accelerate lymphocyte recovery, reduce the risk of graft failure, prevent and treat graft-versus-host disease, and repair tissue damage in patients receiving HSCT. Till now, most MSCs for human clinical application have been derived from bone marrow. However, acquiring bone-marrow-derived MSCs involves an invasive procedure. Umbilical cord is rich with MSCs. Compared to bone-marrow-derived MSCs, umbilical cord-derived MSCs (UCMSCs) are easier to obtain without harm to the donor and can proliferate faster. No severe adverse effects were noted in our previous clinical application of UCMSCs in HSCT. Accordingly, application of UCMSCs in humans appears to be feasible and safe. Further studies are warranted. Yu-Hua Chao, Han-Ping Wu, Chin-Kan Chan, Chris Tsai, Ching-Tien Peng, and Kang-Hsi Wu Copyright © 2012 Yu-Hua Chao et al. All rights reserved. Intrabone Transplant of Cord Blood Stem Cells Establishes a Local Engraftment Store: A Functional PET/FDG Study Tue, 02 Oct 2012 12:16:31 +0000 Background. Despite advancements in comprehension of molecular mechanisms governing bone marrow (BM) homing of hematopoietic stem cells, cord blood transplant (CBT) suffers from a slow rate of hematopoietic recovery. Intrabone (IB) injection has been proposed as a method able to improve speed of BM engraftment with respect to conventional IV protocols. However, the mechanisms underlying this benefit are largely unknown. Aim. To verify whether IB-CBT determines a local engraftment able to predict the reconstitution of recipient hematopoiesis. Design and Methods. Twenty-one patients with hematologic malignancies received IB injection into both iliac crests of 3.2±0.68 * 107/kg cord blood cells. One month following IB-CBT, PET-CT imaging was performed. Maximal standardized uptake values (SUVs) were assessed in BM of both iliac crests and in all lumbar vertebrae. Results. Maximal SUV within iliac crests was higher than in lumbar vertebrae (4.1±1.7 versus 3.2±0.7, resp., 𝑃=0.01). However, metabolic activity in these two different BM districts was significantly correlated (𝑟=0.7, 𝑃<0.001). Moreover, FDG uptake values within the injection site closely predicted platelet recovery 100 days after IB-CBT (𝑟=0.72, 𝑃<0.01). Conclusions. The metabolic activity of injected BM predicts the subsequent rate of hematopoietic recovery after IB-CBT, suggesting a pivotal role of the local engraftment in the reconstitution of recipient hematopoiesis. Cecilia Marini, Marina Podestà, Michela Massollo, Selene Capitanio, Francesco Fiz, Silvia Morbelli, Massimo Brignone, Andrea Bacigalupo, Michele Piana, Francesco Frassoni, and Gianmario Sambuceti Copyright © 2012 Cecilia Marini et al. All rights reserved. Analyzing Gene Expression Profile in K562 Cells Exposed to Sodium Valproate Using Microarray Combined with the Connectivity Map Database Mon, 04 Jun 2012 09:13:28 +0000 To explore the mechanism underlying antileukaemia effect of sodium valproate, the growth and survival of the K562 cell line were investigated. Global profiles of gene expression in K562 cells exposed to sodium valproate were assessed and validated. The differentially expressed genes identified were further used to query the connectivity map database to retrieve a ranked list of compounds that act on the same intracellular targets as sodium valproate. A significant increase in cell apoptosis and a change in gene expression profile were observed in valproate-exposed K562 cells. The significant enrichment analysis of gene ontology terms for the differentially expressed genes showed that these genes were involved in many important biological processes. Eight differentially expressed genes involved in apoptosis were verified by quantitative real-time PCR. The connectivity map analysis showed gene expression profile in K562 cells exposed to sodium valproate was most similar to that of HDACi and PI3K inhibitors, suggesting that sodium valproate might exert antileukaemic action by inhibiting HDAC as well as inhibiting PI3K pathway. In conclusion, our data might provide clues to elucidate the molecular and therapeutic potential of VPA in leukaemia treatment, and the connectivity map is a useful tool for exploring the molecular mechanism of drug action. Xiang-Zhong Zhang, Ai-Hua Yin, Dong-Jun Lin, Xiao-Yu Zhu, Qian Ding, Chun-Huai Wang, and Yun-Xian Chen Copyright © 2012 Xiang-Zhong Zhang et al. All rights reserved. Evaluation of Distinct Freezing Methods and Cryoprotectants for Human Amniotic Fluid Stem Cells Cryopreservation Mon, 14 May 2012 14:22:22 +0000 Amniotic fluid (AF) was described as a potential source of mesenchymal stem cells (MSCs) for biomedicine purposes. Therefore, evaluation of alternative cryoprotectants and freezing protocols capable to maintain the viability and stemness of these cells after cooling is still needed. AF stem cells (AFSCs) were tested for different freezing methods and cryoprotectants. Cell viability, gene expression, surface markers, and plasticity were evaluated after thawing. AFSCs expressed undifferentiated genes Oct4 and Nanog; presented typical markers (CD29, CD44, CD90, and CD105) and were able to differentiate into mesenchymal lineages. All tested cryoprotectants preserved the features of AFSCs however, variations in cell viability were observed. In this concern, dimethyl sulfoxide (Me2SO) showed the best results. The freezing protocols tested did not promote significant changes in the AFSCs viability. Time programmed and nonprogrammed freezing methods could be used for successful AFSCs cryopreservation for 6 months. Although tested cryoprotectants maintained undifferentiated gene expression, typical markers, and plasticity of AFSCs, only Me2SO and glycerol presented workable viability ratios. Felipe de Lara Janz, Adriana de Aguiar Debes, Rita de Cássia Cavaglieri, Sérgio Aloísio Duarte, Carolina Martinez Romão, Antonio Fernandes Morón, Marcelo Zugaib, and Sérgio Paulo Bydlowski Copyright © 2012 Felipe de Lara Janz et al. All rights reserved. The Role of T-Cell Leukemia Translocation-Associated Gene Protein in Human Tumorigenesis and Osteoclastogenesis Thu, 24 Nov 2011 15:49:52 +0000 Synovial tissues of patients with rheumatoid arthritis (RA) include factors regulating bone resorption, such as receptor activator NF-κB ligand (RANKL), TNF-α, IL-6, IL-17, and IFN-γ. However, in addition to these cytokines, other factors expressed in synovial tissues may play a role in regulating bone resorption. In 2009, we demonstrated that novel peptides from T-cell leukemia translocation-associated gene (TCTA) protein expressed in synovial tissues from patients with RA inhibit human osteoclastogenesis, preventing cellular fusion via the interaction between TCTA protein and a putative counterpart molecule. Only a few studies on the role of TCTA protein have been reported. Genomic Southern blots demonstrated a reduced TCTA signal in three of four small cell lung cancer cell lines, suggesting the loss of one of the two copies of the gene. In the current paper, we reviewed the roles of TCTA protein in lung cancer cell lines and human osteoclastogenesis. Shigeru Kotake, Toru Yago, Manabu Kawamoto, and Yuki Nanke Copyright © 2012 Shigeru Kotake et al. All rights reserved. Employment of Oligodeoxynucleotide plus Interleukin-2 Improves Cytogenetic Analysis in Splenic Marginal Zone Lymphoma Sat, 21 May 2011 12:34:24 +0000 To compare the efficiency of novel mitogenic agents and traditional mitosis inductors, 18 patients with splenic marginal zone lymphoma (SMZL) were studied. Three cultures using oligodeoxynucleotide (ODN) plus interleukin-2 (IL-2), or TPA, or LPS were setup in each patient. Seventeen/18 cases with had moderate/good proliferation () as compared with 10/18 cases with TPA and LPS () (); 14/18 () cases with had sufficient good quality of banding as compared with 8/18 cases () with TPA and LPS. The karyotype could be defined from -stimulated cultures in all 18 patients, 14 of whom () had a cytogenetic aberration, whereas clonal aberrations could be documented in 9 and in 3 cases by stimulation with LPS and TPA, respectively. Recurrent chromosome aberrations in our series were represented by aberrations of chromosome 14q in 5 patients, by trisomy 12 and 7q deletion in 4 cases each, and by abnormalities involving 11q and 13q in two cases each. These findings show that stimulation with offers more mitotic figures of better quality and results in an increased rate of clonal aberrations in SMZL, making this method ideal for prospective studies aiming at the definition of the prognostic impact of cytogenetic aberrations in this disorder. Antonella Bardi, Francesco Cavazzini, Gian Matteo Rigolin, Elisa Tammiso, Eleonora Volta, Elisa Pezzolo, Luca Formigaro, Olga Sofritti, Giulia Daghia, Cristina Ambrosio, Lara Rizzotto, Awad E. Abass, Fiorella D'Auria, Pellegrino Musto, and Antonio Cuneo Copyright © 2011 Antonella Bardi et al. All rights reserved. Trypsin Isoinhibitors with Antiproliferative Activity toward Leukemia Cells from Phaseolus vulgaris cv “White Cloud Bean” Mon, 14 Jun 2010 12:05:00 +0000 A purification protocol that comprised ion exchange chromatography on DEAE-cellulose, affinity chromatography on Affi-gel blue gel, ion exchange chromatography on SP-Sepharose, and gel filtration by FPLC on Superdex 75 was complied to isolate two trypsin inhibitors from Phaseolus vulgaris cv “White Cloud Bean”. Both trypsin inhibitors exhibited a molecular mass of 16 kDa and reduced the activity of trypsin with an IC50 value of about 0.6 𝜇M. Dithiothreitol attenuated the trypsin inhibitory activity, signifying that an intact disulfide bond is indispensable to the activity. [Methyl-3H] thymidine incorporation by leukemia L1210 cells was inhibited with an IC50 value of 28.8 𝜇M and 21.5 𝜇M, respectively. They were lacking in activity toward lymphoma MBL2 cells and inhibitory effect on HIV-1 reverse transcriptase and fungal growth when tested up to 100 𝜇M. Jian Sun, Hexiang Wang, and Tzi Bun Ng Copyright © 2010 Jian Sun et al. All rights reserved.