bluuebunny/crossref_metadata_2025 huggingface dataset:
Improving Big Data Projects Requirement Understandability by Using Big Data Use Case Diagram
Authors: Sathis Kumar B. , Sountharrajan S.
Published: October 2019

In today’s Business world, big data projects play an important role in increasing the profit of the business. Success of the big data project lies on the complete understanding of the customer requirement. A big data project not only gives the solution for day to day functionality
of the system but also gives a solution to improve the business. Hence understanding the requirement is very important. Most of the big data projects are based on the object oriented approach. Most of the software industry relies on UML use case model to specify, analyze and design the big
data project. The difference between other project and big data project is data characteristics. The three V characters, volume, velocity and variety play major role in the design decision of big data project. The existing UML use case diagram does not have a facility to represent the big
data V characteristics. In this paper we are proposing big data use case diagram, suitable for specifying big data characteristics. The research result shows that big data use case diagram improves the understandability for the requirements of the big data project.
Structural biology of the TGFβ family
Authors: Goebel Erich J , Hart Kaitlin N , McCoy Jason C , Thompson Thomas B
Published: October 2019

The transforming growth factor beta (TGFβ) signaling pathway orchestrates a wide breadth of biological processes, ranging from bone development to reproduction. Given this, there has been a surge of interest from the drug development industry to modulate the pathway – at several points. This review discusses and provides additional context for several layers of the TGFβ signaling pathway from a structural biology viewpoint. The combination of structural techniques coupled with biophysical studies has provided a foundational knowledge of the molecular mechanisms governing this high impact, ubiquitous pathway, underlying many of the current therapeutic pursuits. This work seeks to consolidate TGFβ-related structural knowledge and educate other researchers of the apparent gaps that still prove elusive. We aim to highlight the importance of these structures and provide the contextual information to understand the contribution to the field, with the hope of advancing the discussion and exploration of the TGFβ signaling pathway.

Impact statement

The transforming growth factor beta (TGFβ) signaling pathway is a multifacetted and highly regulated pathway, forming the underpinnings of a large range of biological processes. Here, we review and consolidate the key steps in TGFβ signaling using literature rooted in structural and biophysical techniques, with a focus on molecular mechanisms and gaps in knowledge. From extracellular regulation to ligand–receptor interactions and intracellular activation cascades, we hope to provide an introductory base for understanding the TGFβ pathway as a whole.
Ham-Wasserman Lecture
Authors: Wang Zhen-yi
Published: October 2019

Conventional treatment of acute leukemia involves the use of cytotoxic agents (chemotherapy), but other strategies have been explored. All-trans retinoic acid (ATRA) and arsenic have clearly been effective in the treatment of acute promyelocytic leukemia (APL), which creates the possibility that other types of acute leukemia can be conquered by selectively inducing differentiation and/or apoptosis. A great number of investigations have been performed to elucidate the mechanisms and search for effective agents in the treatment of other types of acute leukemia by these new strategies. Progress at the molecular level has been achieved in explaining the mechanisms of action of ATRA and arsenic compounds, and several new agents have emerged, although their clinical effectiveness remains to be confirmed. Mechanism-/gene-based targeted therapy and a combination of different strategies will improve the treatment of acute leukemia.
Comprehensive Management of the CLL Patient: A Holistic Approach
Authors: Shanafelt Tait D. , Kay Neil E.
Published: October 2019

The current management of B-chronic lymphocytic leukemia (CLL) is no longer straightforward for the practicing hematologist. Rapid advances in diagnostic precision, methods of predicting prognosis, understanding of natural history of CLL, recognition of clinical complications, clarification of the quality of life (QOL) issues facing the CLL patient, and the exciting array of novel treatment approaches have made the care of the CLL patient more demanding. This review is focused on summarizing these advances in order to provide a framework for integrating this knowledge into routine hematologic practice.
Monitoring after successful therapy for chronic myeloid leukemia
Authors: Branford Susan
Published: October 2019

Monitoring response to therapy for patients with chronic myeloid leukemia using an effective strategy is fundamental for achieving optimal patient outcomes. It will allow the initiation of timely therapeutic intervention for patients with a suboptimal response or kinase inhibitor therapy failure. Evidence is mounting that reaching molecular targets early in therapy is as important as the initial hematologic and cytogenetic response for the identification of patients who may have a poorer outcome. When the molecular target of a major molecular response is achieved at 18 months, patients reach a safe haven where loss of response is rare. However, this benefit is dependent on continuous drug adherence in most patients. As some patients reach their second decade of successful imatinib therapy, how long will frequent response monitoring be necessary? Assuming that very late relapse will be extremely rare for responding patients remaining on kinase inhibitor therapy, there are reasons for maintaining a regular molecular monitoring frequency, including monitoring adherence assessment and confirming sustained undetectable BCR-ABL1 for those considering a discontinuation trial and for late molecular recurrence in patients who maintain response after treatment discontinuation.
Point/counterpoint: early-stage Hodgkin lymphoma and the role of radiation therapy
Authors: Meyer Ralph M. , Hoppe Richard T.
Published: October 2019

The results of recent clinical trials for the management of limited-stage Hodgkin lymphoma have led to considerable debate, especially regarding the role of radiation therapy. This review highlights those recent trials and provides perspectives regarding their interpretation from a radiation oncologist and a hematologist. The trial protocol is available at http://www.nejm.org/doi/suppl/10.1056/NEJMoa1111961/suppl_file/nejmoa1111961_protocol.pdf.
Immunoconjugates and new molecular targets in hairy cell leukemia
Authors: Kreitman Robert J.
Published: October 2019

Hairy cell leukemia (HCL) is a B-cell malignancy that in its classic form is exquisitely sensitive to single-agent purine analog therapy, but that is associated in many patients with late relapse and eventual purine analog resistance. Minimal residual disease, which is present in most patients achieving complete remission with purine analogs, retains Ags that are ideal for targeted therapy. Rituximab, which targets CD20, is active as a single agent, particularly if combined with purine analogs. Recombinant immunotoxins targeting either CD25 or CD22 and containing truncated Pseudomonas exotoxin have achieved major responses in relapsed/refractory HCL. Moxetumomab pasudotox in phase 1 testing achieved responses in 86% of such patients (complete in 46%) without dose limiting toxicity and often without MRD. Soluble CD22 has been used for improved detection and monitoring of HCL, particularly the poor-prognosis variant that lacks CD25. Ig rearrangements unique for each HCL patient have been cloned, sequenced, and followed by real-time quantitative PCR using sequence-specific reagents. Analysis of these rearrangements has identified an unmutated IGVH4-34–expressing poor-prognosis variant with immunophenotypic characteristics of either classic or variant HCL. The BRAF V600E mutation, reported in 50% of melanomas, is present in > 85% of HCL cases that are both classic and express rearrangements other than IGVH4-34, making HCL a potential target for specific inhibitors of BRAF V600E. Additional targets are being defined in both classic and variant HCL, which should improve both detection and therapy.
THE VALUE OF PENICILLIN IN THE TREATMENT OF AGRANULOCYTOSIS CAUSED BY THIOURACIL
Authors: TYSON MARY CATHERINE , VOGEL PETER , ROSENTHAL NATHAN
Published: October 2019

Thiouracil has been found to be an effective drug in the treatment of hyperthyroidism. Agranulocytosis following its use occurred in nine cases, four of which terminated fatally. In five others a complete and rapid recovery took place following penicillin therapy. The latter drug is believed to be ideal for all cases of agranulocytosis, and especially those in which chemotherapy has been used and may have been responsible for the condition. Thus far we have not seen any report of any untoward effect on the hemopoietic system from the use of penicillin.

The use of antibacterial agents for the treatment of agranulocytosis was suggested by Dameshek and Wolfson21 in 1942. It was believed by these authors that patients with agranulocytosis died not of the leukopenia per se but of the sepsis which developed secondarily to the lack of granulocytes. Two very severe cases of aminopyrine agranulocytosis treated with sulfathiazole made complete recoveries. For the treatment of sulfonamide agranulocytosis, it was suggested that a preparation differing from that which had already been used be given. With the discovery of penicillin, and its complete lack of possible deleterious effect on the bone marrow, its use was suggested by Dameshek17 (1944). A report on the beneficial effects of this medication in a case of sulfonamide agranulocytosis was later reported by Dameshek and Knowlton18 and similar cases by Sprague and Ferguson19 and by Meredith and Fink.20

Since sulfonamides may cause further toxic effect on the bone marrow, we feel that their use should be avoided in the treatment of agranulocytosis, especially where a history of previous use is obtained. We do not agree with others21, 22 who continue the use of sulfonamides in the treatment of leukopenia or agranulocytosis where these very drugs may have been responsible for the condition. It would seem better judgment to use penicillin, which by combating the bacterial invasion of the body and the consequent toxemia enables the patient to survive until the bone marrow cells regenerate.
ANTI-ANEMIC PROPERTIES OF THYMINE
Authors: SPIES TOM D. , FROMMEYER WALTER B. , VILTER CARL F. , ENGLISH ANN
Published: October 2019

Three patients with Addisonian pernicious anemia were selected, hospitalized and given a diet devoid of meat and meat products. After baseline hematological studies were made and checked repeatedly, daily large doses of thymine were given orally.

The clinical and hematological improvement in these three patients was in every way similar to that which follows the administration of folic acid to patients with pernicious anemia in relapse. The exact mode of action of thymine is obscure, but it is possible that folic acid may act as an enzyme or co-enzyme in the synthesis of thymine or a thymine-like compound. Such synthesis may take place in the gastro-intestinal tract. The present findings indicate that thymine has anti-anemic properties and a profound effect on the general metabolism of patients with Addisonian pernicious anemia in relapse.
Histochemical and Quantitative Studies of the In Vitro Dehydrogenase Activity of Spleen in Hypersplenism
Authors: BLACK MAURICE M. , PRESTON JOSEPH A. , SPEER FRANCIS D. , BRENOWITZ PHYLLIS
Published: October 2019

Tetrazolium chloride was used to visualize the in vitro dehydrogenase activity of splenic tissues from cases with and without hypersplenism. In addition, quantitative measurements of such activity were made in the presence and absence of fluoride, malonate and azide. The data indicate that spleen slices from cases of hypersplenism differed from the controls and were characterized by (a) increased endogenous dehydrogenase activity of the lymphocytes, particularly around the follicles, (b) a decreased azide stimulation in those cases having an initial thrombocytopenia which responded to splenectomy, and (c) a decreased fluoride stimulation in those cases having an initial leukopenia which responded to splenectomy.

Routine histological studies confirmed other reports that a constant structural feature of the spleen in hypersplenism is a perifollicular lymphocytic rimming.
Myeloblastic Leukemia Preceded by Prolonged Hematologic Disorder
Authors: WILLIAMS MARJORIE J.
Published: October 2019

A case with a prolonged period of blood disorder preceding the terminal development of myeloblastic leukemia is reported in some detail. The preleukemic period was characterized by anemia, neutropenia and thrombocytopenia. It extended over a period of almost nine years but was punctuated by a period of remission lasting four and one half years. In the initial phase, the marrow was hypoplastic but during the later stages, it was hyperplastic.

Brief reference is made to somewhat similar cases that have been recorded and to the ideas expressed by the authors of these reports.

It is suggested that further investigation is needed to determine whether the initial blood disease occurring in such cases is irrevocably linked to the subsequent leukemia.
Constitutional Anemia (Fanconi’s Syndrome) and Leukemia in Two Brothers
Authors: COWDELL R. H. , PHIZACKERLEY P. J. R. , PYKE D. A.
Published: October 2019

Two brothers physically very alike became ill in their third decade, one with severe bone marrow hypoplasia and the other with acute leukemia. Both died and the autopsy findings are described.

The two shared certain congenital defects, notably short stature with small heads, cutaneous pigmentation, pituitary and genital hypoplasia, and deformity of the thumbs. The brother with hypoplastic anemia also had aortic hypoplasia and his left kidney was absent.

No previous report has been discovered of the association of Fanconi’s syndrome (of bone marrow hypoplasia with congenital defects) and leukemia with similar congenital defects in brothers. In two reports of leukemia affecting cousins of patients with Fanconi's syndrome, the leukemic cousins were without congenital defects.
An Evaluation of Paraffin Sections of Aspirated Bone Marrow in Malignant Lymphomas
Authors: PETTET JACQUELINE D. , PEASE GERTRUDE L. , COOPER TALBERT
Published: October 2019

Examination of serial paraffin sections of bone-marrow aspirations has increased the known incidence of marrow involvement in malignant lymphomas. The results of this study can be seen in the table. In the 72 cases reviewed here, lesions compatible with malignant lymphoma were seen in paraffin sections in 22 cases. In 12 cases this was the only material available for diagnosis at the time. In an additional seven cases paraffin sections were considered suggestive of malignant lymphoma and in five of these a diagnosis could be made on the basis of examination of both smear and paraffin section. This procedure is particularly helpful in Hodgkin’s disease and reticulum cell sarcoma, where marrow smears are only rarely diagnostic.

In general, the patients in whom lesions were found in the bone marrow tended to have more advanced disease than those in whom lesions were not found, and abnormalities in the peripheral blood were more common. This was especially true of patients with Hodgkin’s disease, less so in those with reticulum cell sarcoma. Patients with lymphosarcoma, lymphocytic type, did not show this difference, either in the extent of the disease or in findings on examination of peripheral blood.

Diagnosis was most difficult in patients who had lymphocytic lymphosarcoma, since it was occasionally difficult to distinguish between the benign aggregations of lymphocytes which are not infrequently found in a large number of unrelated conditions and those aggregations which represented malignancy. Differentiation was made on the basis of number of aggregations, degree of circumscription, presence or absence of reaction centers and the cytologic appearance of the cells. Examination of the smears of the marrow is most helpful in these cases.

Seven patients in this series (9.7 per cent) had granulomatous lesions in the marrow. In five of these the final diagnosis was Hodgkin’s disease, in one reticulum cell sarcoma and in one follicular lymphoma. Although these lesions were not specific, the incidence is high enough to make the finding of a granuloma of some significance in any patient suspected of having malignant lymphoma.

The importance of thorough examination of any material obtained in an apparent dry tap and the value of repeated marrow-examinations is emphasized by three cases in this series.

Since in many cases diagnostic lesions are found in only one portion of the material, examination of only one section of the paraffin sections of aspirated bone marrow is not sufficient for proper evaluation. This fact is emphasized by the greater incidence of positive findings in this report than in that of Cooper and Watkins.

Studies of bone marrow should be particularly helpful in instances in which the diagnosis of malignant lymphoma is suggested by clinical features but cannot be proved by biopsy of peripheral nodes or other readily accessible tissue.
Immunophenotypic Diagnostic Score for MDS Taking into Account Various Cell Populations.
Authors: Millaret Arnaud , Galmarini Carlos , Ritouet Danielle Treille , Gerland Luc M. , Dumontet Charles M.
Published: October 2019

Immunophenotyping has proven to be an interesting approach in patients with myelodysplastic syndromes (MDS)(Maynadie et al., Blood. 2002 Oct 1;100(7):2349–56). Most studies in these diseases have focused on the analysis of blast cells. We chose to determine the potential interest of markers expressed on other bone marrow compartments, namely granulocytes, monocytes and the CD34 compartment.

Fourteen marrow samples from patients with MDS and 10 normal bone marrow were analyzed. The median age in the MDS group and the control group was 64.4 and 59.8 years, respectively. Flow cytometry analysis of bone marrow samples was performed on a Facs Calibur, using four-color panels. The following antigens were analyzed: CD7, CD11c, CD14, CD15, CD16, CD33, CD34, CD45, CD36, CD38, CD56, CD71, CD117 and HLA-DR. The various compartments analyzed were defined using CD45/SSC gating, combined with SSC/CD34 gating for CD34+ cells. The median percentage of blasts in the MDS marrows was < 5%. Seven patients had a hemoglobin value < 100 g/dl. Five patients had a leukocyte count < 3.109/l. Six patients had a platelet count < 100.1012/L. Threshold values were determined by ROC analysis. Comparison of the expression levels of markers between MDS and normal bone marrows was performed using a Wilcoxon test. Analysis was performed on bone marrow granulocytes, monocytes and CD34 cells.

Granulocytes were found to express higher levels of CD33 in MDS than in normal marrows (p=0.007). CD34 + cells in MDS marrows expressed significantly higher levels of CD11c (p=0.007), CD117 (p=0.023) and CD33 (p=0.013) than their normal counterparts, with a trend towards lower expression of CD71 (p=0.053). Monocytes in MDS marrows expressed significantly lower levels of CD16 (p=0.006), CD36 (p=0.004) and CD117 (p=0.007) than their normal counterparts, with a trend towards lower expression of CD11c (p=0.059). We did not observe any differences in the antigen expression of MDS erythroblasts in comparison to their normal counterparts. A diagnostic score (0–7) was constructed taking into account the following seven parameters: expression of CD33 on granulocytes; CD11c, CD33 and CD117 on CD34+ cells; CD16, CD36 and CD117 on monocytes. This score distinguished all cases of MDS samples from controls. These results suggest that analysis of cell populations other than blasts could be useful in patients with MDS and that an immunophenotypic panel composed of markers commonly used in the diagnosis of acute leukemias could constitute a useful tool, in complement to morphological studies, for the diagnosis of myelodysplastic syndromes.
Management of Patients with Multiple Myeloma (MM) Failing Total Therapy 2 (TT 2) According to Thalidomide (THAL) Randomization.
Authors: Barlogie Bart , Rasmussen Erik , Tricot Guido , Crowley John
Published: October 2019

As of August 04, 104 of 668 patients enrolled in TT 2 have relapsed/progressed, 61 of whom had been randomized to no THAL and 43 to THAL. The 2 groups were comparable in 14 pre-treatment features examined including the presence of cytogenetic abnormalities (CA), LDH, B2M and albumin. At the time of treatment failure, which occurred at a similar median time from study enrollment of 23 months, higher proportions of patients on THAL had CA (63% vs 37%, p=.03) and higher levels of bone marrow plasmacytosis >33% (46% vs 12%, p=.001). Their subsequent management included THAL + DEX in 25%; Velcade + THAL in 16% and DT-PACE in 10% of those failing on the no THAL arm; compared to THAL + DEX in 12%; Velcade + THAL in 14% and DT-PACE in 21% of those relapsing on the THAL arm. Subsequent salvage response was 51% for those relapsing on no THAL and 26% of those on THAL (relapsing on THAL) (p=.028). Importantly, survival from the time of TT 2 failure was significantly better when patients had not received prior THAL than when THAL was part of the initial management (median of 29 vs 8 mos, p=.0001).

Univarately adverse features for post-failure survival included the presence of CA at the time of failure, IgA isotype, LDH at baseline prior to TT 2 and initial randomization to THAL. On multivariate analysis, baseline LDH (HR 2.6, p=.0006) and THAL randomization (HR 2.6, p=.0006) were highly associated with inferior post-TT 2 failure survival (R2 =38%). Indeed, those on the THAL arm had inferior survival regardless of CA, present either at baseline or at relapse (Figure 1). Our preliminary conclusion is that initial use of THAL for the primary management of symptomatic MM reduces the subsequent salvage potential, due to the emergence of a more aggressive disease exhibiting CA at relapse and greater tumor burden (high bone marrow plasmacytosis). Thus, randomized trials are needed to determine whether thalidomide is more appropriately used up front or for the management of relapse in order to achieve the longest overall survival.

Figure Figure
Imatinib Mesylate in the Treatment of Systemic Mastocytosis, a Phase I/II Trial.
Authors: Droogendijk H.J. , Kluin-Nelemans J.C. , van Daele P.L.A
Published: October 2019

Introduction: mastocytosis comprimes a group of diseases characterized by abnormal proliferation and accumulation of mast cells in one or more organs. A cutaneous and systemic form of mastocytosis is distinguished. Systemic mastocytosis defines the disease process in which mast cell proliferation exceeds the skin. The clinical manifestations of systemic mastocytosis depend on the tissues involved and the tissue response to the accumulation of mast cells. Although in general the disease progresses slowly, it may develop into a malignant disease. Currently there is no cure for systemic mastocytosis. Mast cells develop from pluripotent bone marrow progenitor cells that express CD34 antigen and are dispersed as precursors which undergo proliferation and maturation in different tissues. Normal mast cell development involves the action of stam cell growth factor and c-kit receptors, which are expressed by mast cells at their different developmental stages. Deregulation and/or abnormalities of the c-kit receptor are assumed to play a causal role in disordered mast-cell proliferation. In most patients a mutation in the gene for c-kit exists. One of the mutations is the D816V mutation.

Aim of the study:imatinib mesylate, formerly called ST1571, is a potent inhibitor of c-kit receptor tyrosine kinase activity. In this study, we evaluate whether imatinib mesylate is safe and effective in the treatment of patients with systemic mastocytosis. Primary end-points of study are reduction in urinary N-methylhistamine excretion, serum tryptase activity, skin lesions, number of mast cells in sections of bone marrow, hepato-and/or splenomegaly and symptoms.Adverse effects on therapy are also considered.

Results: up to now, 10 patients with systemic mastocytosis are treated with 400 mg of imatinib mesylate orally once daily. During the first 2 weeks of the study the patients also received 30 mg of prednisolone daily. In general imatinib mesylate is well tolerated. The first results show a 38–80% reduction in urinary N-methylhistamine excretion and 30–66% reduction in serum tryptase activity. Skin lesions diminish in two of the six patients with cutaneous mastocytosis,. Number of mast cells in sections of bone marrow are reduced in 63% (5/8) of the patients. Hepato-and/or splenomegaly is slightly decreased in two of the three patients with organomegaly. Finally 60 % of all patients experiences relief of symptoms. In eight patients the D816V mutation was found. In contrast with former studies imatinib mesylate is also effective in these patients. Further results are to be awaited.

Conclusion: imatinib mesylate is safe and seems effective in the treatment of patients with systemic mastocytosis (including patients with the D816V mutation).
Enhanced Apoptosis to Chemotherapeutic Agents Is Dependent on NFκB and Bcl2-Related Proteins but Is Independent of P53 and Bax in Burkitt’s Lymphoma Cells.
Authors: Kanda Kayoko , Wong Wendy , Boxer Linda M.
Published: October 2019

Deregulated expression of c-Myc is involved in the pathogenesis and growth of Burkitt’s lymphoma cells. Aberrant NFκB activation has been linked to several forms of cancers, including Hodgkin’s lymphoma, breast cancers, and B-cell malignancies. The cytoplasmic retention of NFκB is mediated by a family of inhibitor proteins, the IκBs. We showed that expression of a super-repressor form of IκBα, IκBα-SR, resulted in decreased proliferation and an increased G0/G1 population of Raji Burkitt’s cells. No spontaneous apoptosis was observed, but there was increased sensitivity to apoptosis following treatment with chemotherapeutic agents. Further studies were performed to determine the mechanisms involved in the induction of apoptosis. Interference with NFκB activity decreased the levels of c-Myc and Bcl-xL, but there was no change in the levels of Bax, Bak, Mcl-1, or p53. Treatment with chemotherapeutic agents alone resulted in decreased expression of Mcl-1 with no change in the other proteins measured. Decreased expression of both Bcl-xL and Mcl-1 following inhibition of NFκB activity and chemotherapy (vincristine, 4HC or doxorubicin) treatment resulted in apoptosis. In support of the important role of Bcl-xL and Mcl-1, siRNA experiments revealed that decreased expression of either one alone did not lead to apoptosis, while decreased expression of both Bcl-xL and Mcl-1 resulted in the induction of apoptosis. Many Burkitt’s lymphoma cells have mutations in p53. We found that the mutated p53 was able to bind to IκBα and Iκ Bα-SR as well as to Bcl-xL, however, no up-regulation of Bax or p53 was observed in response to a chemotherapeutic agent. This indicates that mutated p53 retains the ability to bind the other proteins but has lost transcriptional function. Introduction of high levels of wild-type p53 into Raji Burkitt’s cells was able to restore the induction of Bax and Mdm2 and induce apoptosis, whereas the mutant was not. These results suggest that sensitization of Burkitt’s cells to apoptotic death with chemotherapeutic agents requires interference with NFκB activity and changes in the expression of Bcl2-family members but is independent of p53. Inhibition of NFκB activity could be useful in combination with chemotherapy for the treatment of Burkitt’s lymphoma.
Contig Array CGH at 3p14.2 Points to the FHIT Gene as the Deleted Gene in Diffuse Large B Cell Lymphoma.
Authors: Kameoka Yoshihiro , Tagawa Hiroyuki , Tsuzuki Shinobu , Ota Akinobu , Suzuki Ritsuro , Seto Masao
Published: October 2019

Diffuse large B-cell lymphoma (DLBCL) is the most common type of non-Hodgkin’s lymphoma (NHL), accounting for 30 – 40% of adult NHL. Genomic alterations in DLBCL have been investigated by various methods and many regions of amplifications and losses have been detected. Genomic deletions of the 3p arm have been reported in many solid tumors and leukemias, but no study published to date has reported genomic deletions of this region in DLBCL. Recently we demonstrated that 3p14.2 was deleted in approximately 30% of DLBCL patients and cell lines by use of a genome-wide array-comparative genomic hybridization (array-CGH) (Tagawa et al., 2004). For a more detailed examination of the genomic losses at 3p14.2, here we made use of contig BAC array for 3p14.2, and found that 12 of 27 DLBCL samples displayed losses. All of the deleted regions were located within the Fragile Histidine Triad (FHIT) gene, and the most frequently region of loss was mapped to 0.4 Mbp of the region encompassing the introns 4 and 5 and exon 5 of the FHIT gene. Genomic deletions of the FHIT gene have been observed in most common forms of cancer and it is therefore conceivable that the FHIT gene is the target one of the genomic deletion at 3p in DLBCL. To investigate the relationship between genomic deletions and transcript alterations, we performed nested reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. Aberrant transcripts or loss of expression was detected in 33% (19 of 57) of the DLBCL samples, and the lost exons of the aberrant transcripts were correlated with genomic deletions detected by array CGH. We also investigated the CpG island methylation status of the promotor of the FHIT gene by means of methylation specific PCR (MSP). One case with genomic loss of the FHIT gene, and three cases without genomic loss showed methylated allele. These findings indicate that 1) Loss of genomic material at 3q14.2 is responsible for exon losses of the FHIT gene, and 2) Genomic loss of the FHIT gene is one of the causes of the generation of aberrant transcripts. 3) Both genomic deletions and methylations are involved in the causes of inactivation of the FHIT gene.
Direct Adoptive Transfer of Cytomegalovirus-Specific CTL from Unrelated Donors to Stem Cell Transplant Patients Following Selection by HLA-Peptide Tetramers.
Authors: Cobbold Mark , Amrolia Persis , Mahendra Prem , Steward Colin , Olavarria Eduardo , Arrazi Julie , McDonald Dorothy , Tauro Sudhir , Goldman John , Craddock Charles , Moss Paul
Published: October 2019

Cytomegalovirus (CMV) infection remains an important cause of morbidity and mortality in patients undergoing allogeneic stem cell transplantation (SCT). This is particularly true in patients transplanted using volunteer unrelated donors (VUDs) where despite improvements in antiviral therapy CMV positive serostatus remains an adverse prognostic indicator. Adoptive transfer of donor-derived CMV-specific CD8+ T cell clones reduces the rate of viral reactivation but the complexity of this approach severely limits its clinical application.

We have previously reported the purification of CMV-specific CD8+ cytotoxic T lymphocytes (CMV-CTL) from the blood of stem cell transplant donors using HLA-peptide tetramers and their subsequent transfer. Six patients undergoing stem cell transplantation from HLA identical sibling donors were studied. CMV-CTL were infused on a pre-emptive basis in patients who developed evidence of CMV viremia. All patients showed prompt and durable CMV-specific immune reconstitution demonstrating durable CMV-specific immune reconstitution. CMV-CTL constituted up to 21% of the patient lymphocyte CD8+ T cell compartment and were detectable up to 24 months in all patients. There was a reduced requirement for anti-viral agents in treated patients (33%) compared with a control group (82%) p=0.11. No secondary reactivation was seen in any patient in the treatment group. No infusion-related toxicity was seen in any treated patients. We have now extended these studies to recipients of VUD transplants in whom no CMV-CTL are detectable within the first 100 days of stem cell infusion. Tetramer-selected CMV-CTL have been adoptively transferred into six patients undergoing transplantation from an unrelated donor. CMV-CTL were given either for treatment of refractory CMV infection (n=5) or prophylactically 21 days post SCT (n=1). A cell dose of between 6x102/kg and 1.25x105/kg was administered with median purity of 95% of the infused T cell dose. No infusional toxicity or acute graft-versus-host disease was noted. CMV-CTL were not present in any patient prior to therapy. Within 7 days of adoptive transfer CMV-CTL were detectable in all patients comprising 0.6% – 20% of CD8+ T cell compartment. All patients demonstrated a reduction in the viral load and 5/6 cleared their viremia. In the patient who received prophylactic therapy durable expansion of CMV-CTL were observed up to 20% of CD8+ T cell pool. Taken together these data are consistent with the hypothesis that antigen-specific CMV specific CTL can be safely infused to recipients of VUD SCT and expand directly in vivo. Such an approach has considerable potential in the treatment or prophylaxis of CMV infection in patients undergoing VUD SCT.
G12/13 Pathways Potentiate Akt Phosphorylation in Platelets Mediated by Gi/Gz Pathways in a Src Kinase-Dependent Manner.
Authors: Kim Soochong , Jin Jianguo , Kunapuli Satya P.
Published: October 2019

Akt is a serine/threonine kinase that is activated by various agonists including thrombin and ADP in platelets, and activation of Akt in platelets is known to require Gi signaling pathways. Even though thrombin-induced Akt phosphorylation depends on secretion/Gi pathways, thrombin caused much stronger Akt phosphorylation than ADP and epinephrine. In this study, we investigated the contribution of G12/13 pathways to Akt phosphorylation mediated by Gi or Gz pathways. We used selective agonists to activate different G protein pathways. PAR4-activating peptide (AYPGKF) and thrombin failed to induce Akt phosphorylation in Gαq-deficient platelets, but Akt phosphorylation was restored to the levels achieved by AYPGKF and thrombin in wild-type platelets by selective supplement of either Gi or Gz signaling with 2-MeSADP and epinephrine, respectively. This phosphorylation of Akt was dramatically inhibited in the presence of PP2

(4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazole[3,4-d]pyrimidine), an inhibitor of Src family tyrosine kinase, but not by PP3, an inactive structural analog. Importantly, AYPGKF and thrombin induced the activation of Src kinase in Gαq-deficient platelets, suggesting the involvement of Src-dependent pathways in the G12/13 signaling in potentiation of Akt phosphorylation. When human platelets were stimulated with low concentrations of YFLLRNP, a PAR1-specific partial agonist, to selectively activate the G12/13 signaling cascade, Akt phosphorylation did not occur. However, combined stimulation of YFLLRNP-mediated G12/13 signaling and selective activation of Gi pathways caused the Akt phosphorylation. This Akt phosphorylation was blocked by the P2Y12 receptor antagonist AR-C69931MX, or the PI 3-kinase inhibitor LY294002. Platelet aggregation induced by co-activation of both G12/13 and Gi signaling was dramatically inhibited by ML-9

(1-(5-chloronaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride), an Akt selective inhibitor, suggesting an important role of Akt in platelet aggregation stimulated by combined G12/13 and Gi pathways. These data demonstrate that G12/13 signaling alone does not cause Akt phosphorylation in platelets, but G12/13 signaling has a significant role in potentiating Akt phosphorylation mediated by selective Gi or Gz signaling in human as well as mouse platelets. Finally, we conclude that Src family kinases play an important role in this Akt phosphorylation in platelets.
Red Blood Cell S-Nitrosohemoglobin Deficiency in Pulmonary Arterial Hypertension.
Authors: Pawloski John R. , McMahon Timothy J. , Moya Martin P. , Gow Andrew J. , Huang Yuh-Chin T. , Luchsinger Benjamin J. , Krichman Abigail D. , Bashore Thomas M. , Califf Robert M. , Singel David J. , Piantadosi Claude A. , Tapson Victor F. , Stamler Jonathan S.
Published: October 2019

Recent studies have suggested that binding of oxygen to hemoglobin (Hb) facilitates reactions of nitric oxide (NO) that lead to production of S-nitrosohemoglobin (SNO-Hb), and that vasodilator S-nitrosthiol (SNO) is dispensed by red blood cells (RBCs) at low oxygen tension (pO2) to dilate blood vessels. In human lungs, NO bioactivity serves to attenuate hypoxic pulmonary vasoconstriction (HPV). We therefore considered the possibility that RBC-SNO may oppose HPV and that defective vasodilation by RBCs may contribute to the pathophysiology of pulmonary arterial hypertension (PAH). Here we report that RBCs from patients with PAH exhibit substantial depletion of SNO-Hb and consequent impairment in hypoxia-mediated vasodilation. Furthermore, levels of RBC-NO correlated inversely with pulmonary artery pressures. A SNO-Hb deficiency characteristic of PAH was reproduced in control RBCs by hypoxia: loss of SNO-Hb was accompanied by a buildup of heme-NO species that are deficient in the pO2-governed intramolecular transfer of NO to cysteine thiol, yielding RBCs deficient in NO bioactivity. SNO-deficient RBCs produced exaggerated HPV responses as compared to SNO-replete RBCs. In PAH patients, SNO-Hb repletion fully restored the hypoxic vasodilator activity of RBCs. Our results suggest that a deficiency in RBC-SNO contributes to pulmonary hypertension and hypoxemia, and that repletion of RBC-SNO represents a rational strategy for treating PAH patients.
TAL1 Regulates Expression of the Band 3 Gene in Erythroid Progenitors through Both Upstream Regulatory and Proximal Promoter Elements.
Authors: Xie Jingping , Xu Zhixiong , Brandt Stephen J.
Published: October 2019

The TAL1 (also known as SCL and TCL-5) protein is a class II basic helix-loop-helix (bHLH) transcription factor that plays an important role during embryonic and adult hematopoiesis. We previously established that the Protein 4.2 (P4.2) gene is a physiologic target of TAL1 in erythroid cells and showed that tandem E box-GATA elements in its proximal promoter mediated TAL1-directed gene activation. Through database searches, we identified two E box-GATA consensus sequences in the proximal promoter of another erythroid membrane skeleton gene, that encoding the Band 3 protein. Identical to what was observed for P4.2, overexpression of wild-type TAL1 slightly increased while enforced expression of a DNA binding-defective TAL1 mutant severely reduced endogenous Band 3 gene expression in murine erythroleukemia (MEL) cells induced to differentiate with dimethyl sulfoxide (DMSO). Overexpression of Ldb1 significantly inhibited and a LIM protein interaction-defective Ldb1 mutant virtually ablated Band 3 mRNA accumulation in DMSO-induced MEL cells. Quantitative chromatin immunoprecipitation (ChIP) analysis with Tal1 antibody confirmed that Tal1 was associated with the proximal promoter of the Band 3 gene in MEL cells, while analysis of approximately 28 kb of genomic sequence spanning the Band 3 gene, including 5 kb 5′ and 3′ of the gene, revealed another consensus E box-GATA element in intron 16; however, no significant TAL1 binding was detected in this region by ChIP analysis. To identify potential non-conventional TAL1 binding site(s), scanning ChIP analysis was applied to the entire 28 kb Band 3 genomic region, and a region ~2.5 kb upstream of the major transcriptional start site was found to bind significantly more TAL1 protein than the proximal promoter. Fine mapping of TAL1 binding to this region by ChIP analysis using more highly sheared DNA and smaller sized amplicons narrowed TAL1 binding to a region of ~100 bp, which we designated as the Band 3 upstream regulatory region (B3URE). Luciferase reporter assays in transiently transfected MEL cells with vectors containing genomic sequence revealed the presence of an orientation- and position- independent repressor in this upstream region, with a 107 bp fragment retaining nearly all the repressing activity of a larger, 700 bp fragment. In contrast, coexpression of TAL1, E47, GATA-1, LMO2, and Ldb1 in COS cells led to transactivation of a reporter gene linked to the promoter-proximal E box-GATA element. These data suggest that TAL1 acts as a repressor on the upstream B3URE in undifferentiated cells and then, as differentiation proceeds, as an activator on the E box-GATA elements in the proximal promoter.
GATA-1 Directly Induces Physical Proximity between the LCR and β Globin Promoter in Erythroid Cells.
Authors: Vakoc Christopher , Dekker Job , Blobel Gerd A.
Published: October 2019

The regulation of higher-order chromatin folding is a poorly understood aspect of nuclear biology. The beta globin locus is one of the most thoroughly studied mammalian loci with regard to its chromatin organization and sub-nuclear localization. The LCR (locus control region) is a powerful enhancer that acts at great distances to regulate the expression of all beta-like globin genes. Recent evidence suggests that the LCR is in physical proximity with the active but not inactive globin gene promoters, thereby looping out up to tens of kilobases of intervening sequence. The LCR-promoter interaction is dynamically regulated throughout development and cellular differentiation. However, the transcription factors that mediate long range “loops” are unknown. Here we examined the essential erythroid transcription factor GATA-1 for its role in LCR-promoter communication using the 3C (Chromosome Conformation Capture) technique. For our studies we used a genetic complementation assay in which erythroid maturation and globin gene expression depend on the activity of a conditional, drug-activated form of GATA-1. The results suggest that GATA-1 induces proximity of the LCR with the active beta-globin promoter. Kinetic studies revealed that the LCR-promoter interaction occurs rapidly and coincides with or even precedes the onset of globin gene transcription. Moreover, “loop” formation correlates with GATA-1 occupancy as determined by ChIP assay, and occurs in the presence of cyclohexamide, indicating that the effects of GATA-1 are direct. Together, these data suggest GATA-1 is a required component for the formation of a tissue-specific chromatin loop. Additional experiments with mutant versions of GATA-1 that address which known GATA-1 cofactors are required for this novel function of GATA-1 will be discussed.
Combined Immuno-Chemotherapy (R-CHOP) Results in Significantly Superior Response Rates and Time to Treatment Failure in First Line Treatment of Patients with Lymphomplasmocytoid/ic Immunocytoma (LP-IC) - Results of a Prospective Randomized Trial of the German Low Grade Lymphoma Study Group (GLSG).
Authors: Buske Christian , Dreyling Martin H. , Eimermacher Hartmut , Boeck Hans-Peter , Pfreundschuh Michael , Metzner Bernd , Fuchs Roland , Woermann Bernhard , Truemper Lorenz H. , Hess Georg , Wandt Hannes , Ludwig Wolf-Dieter , Kreuser Ernst-Dietrich , Schimke Joerg , Weh H.-J. , Schmitz Stephan , Schmiegel W. , Unterhalt Michael , Hiddemann Wolfgang
Published: October 2019

Lymphomplasmocytoid/ic immunocytoma (LP-IC), including Waldenstrom’s macro-globulinemia and lymphoplasmacytoid lymphoma according to the Kiel classification (the latter one subsummed as a variant of B-CLL in the WHO classification), is an indolent lymphoma, which is incurable by conventional chemotherapy in the advanced stage of disease. The anti-CD20 antibody Rituximab has shown remarkable activity in indolent lymphomas, in particular when combined with chemotherapy. Based on these results the GLSG investigated the efficacy of a combined immuno-chemotherapy (R-CHOP: Rituximab 375 mg/m2 d0-1; cyclophosphamide 750 mg/m2 d1; doxorubicine 50 mg/m2 d1; vincristine 1.4 mg/m2 d1; prednisone 100 mg/m2 d1-5) versus CHOP alone as first line treatment of LP-IC in a multicenter prospective randomized phase III trial. Of 72 patients 71 % were classified as lymphoplasmacytic lymphoma, 29 % as lymphoplasmocytoid subtype by central pathology review. The median age was 61.5 years (60 years for R-CHOP and 62 years in the CHOP arm); 24% of the patients had an IPI score >2. The overall response (OR) rate was significantly improved by combining Rituximab with CHOP compared to CHOP alone with an OR of 94% (11% CR, 83% PR) for R-CHOP and an OR of 69% (3% CR, 67% PR) for CHOP (p=0.012). Furthermore, patients treated with R-CHOP showed a significantly prolonged time to treatment failure (TTF) with an estimated median TTF of 22 month in the CHOP arm, whereas the median was not reached in the immunochemotherapy arm after a maximum follow up of nearly 4 years (p=0.0057). There was no major difference of the toxicity in both treatment groups. In summary, these data indicate a beneficial effect of combined immunochemotherapy (R-CHOP) as compared to CHOP alone with regards to initial cytoreduction as well as TTF in patients with LP-IC in a prospective randomized trial, establishing R-CHOP as an highly attractive regimen for first line treatment of this distinct lymphoma subtype.
The Impact of Anemia on Mortality, Cognition and Function in a Racially Diverse Community-Dwelling Elderly Population.
Authors: Denny Susan , Kuchibhatla Maragatha , Cohen Harvey Jay
Published: October 2019

The WHO definition of anemia, Hgb<12g/dL (W) and <13g/dL (M) is largely based on studies in younger populations. As our population is aging it is important to explore the impact of varying Hgb levels on mortality and function in representative populations of older persons. The Duke Established Populations for Epidemiologic Studies for the Elderly (EPESE) is a random household sample, over sampled for African Americans (AA), initiated in 1986. In 1992 we obtained Hgb levels for 1744 subjects age >71 and evaluated functional and cognitive status at baseline and 4 years later, and mortality for 8 years. Using WHO criteria, the prevalence of anemia was 24%. It was age related; 17% (65–74), 25% (75–79), 32% (>80) p<.0001, but not gender related. There was a strong racial difference, 33.6% AA, 13.8% Caucasians (p=<.0001) with an odds ratio, adjusted for age, education, renal insufficiency and comorbidity of 3.0 (95%CI 2.3 – 3.9) Baseline functional status was worse in anemic subjects (p<.0001) but cognition was not (p=.06). The risk ratio for 8 year mortality was 1.7 (95% CI 1.5, 2.0) for anemic subjects (p=.0001). This did not differ by gender nor, despite the increased prevalence, by race (RR AA: caucasian = 0.85; 95% CI 0.6–1.2). Assessing survival by Hgb level, a nonlinear relationship was seen, with highest % survival between 13–14 g/dL for women (p<.0001) and 14–15g/dL for men (p=.09). Using the WHO standard as reference, controlling for baseline characteristics, women with Hgb 10–11 g/dL had higher mortality (RR 2.1, 95% CI. 1.5 –3.1) but with 11–12 g/dL did not. Similarly men with Hgb 10–11 g/dL had increased mortality (RR2.0, 95% CI 1.1, 3.7). In women there was a progressively greater decline in function at 4 years with each decrement of Hgb level from 16 to 10g/dL, but not in men. Both women and men with mild anemia (11–12 g/dL and 10–11g/dL respectively) had greater decreases in cognition than those at higher levels (p=.01). Thus in an elderly community based population, anemia is significantly more common in AA independent of disease status, and is independently associated with increased mortality over eight years for both races and genders, and with declines in function and cognition most prominently in women. Further investigation of the etiology of these relationships is warranted.