bluuebunny/crossref_metadata_2025_split huggingface dataset:
High BCR-ABL/GUSIS Levels at Diagnosis Are Associated with Unfavorable Responses to Standard Dose Imatinib
Authors: Vigneri Paolo , Stagno Fabio , Stella Stefania , Cupri Alessandra , Forte Stefano , Massimino Michele , Antolino Agostino , Caracciolo Clementina , Nocilli Laura , Impera Stefana , Musolino Caterina , Turri Diamante , Russo Mario , Tomaselli Carmela Anna Maria , Rizzo Michele , Musso Maurizio , Morabito Fortunato , Levato Luciano , Manzella Livia , Mueller Martin C , Hochhaus Andreas , Di Raimondo Francesco 
Published: October 2019

Abstract

Background
The approval of second-generation tyrosine kinase inhibitors (TKIs) for the first line treatment of Chronic Myeloid Leukemia (CML) has generated a need for early molecular parameters associated with inadequate responses to Imatinib Mesylate (IM).
Objective
We correlated quantitative determination of BCR-ABL transcripts at diagnosis with the outcome (defined according to the 2013 European Leukemia Net recommendations) of 272 newly diagnosed CML patients receiving IM 400 mg/die.
Methods
BCR-ABL transcripts were measured from peripheral blood samples drawn at diagnosis before patients received any pharmacological treatment using Real-Time Quantitative PCR (RQ-PCR). All molecular determinations were performed twice (in triplicates) on the same sample using either ABL or glucuronidase-beta (GUS) as reference genes. BCR-ABL values were then reported on the international scale (IS).
Results
With a median follow-up of 60 months, 65.4% of patients achieved an optimal response, 5.6% presented a response currently defined as "warning", 22.4% failed IM treatment and 6.6% switched to a different tyrosine kinase inhibitor because of intolerance to the drug. We recorded 19 deaths (6.9%), 7 (2.5%) attributable to disease progression. We applied Receiver Operating Characteristic (ROC) curves to define BCR-ABL/GUSIS expression levels that would separate patients likely (i.e. below the threshold) or unlikely (i.e. above the threshold) to achieve multiple endpoints, namely: optimal response (OR), failure-free survival (FFS), event-free survival (EFS), transformation-free survival (TFS) and overall survival (OS). Employing the specific threshold calculated for each endpoint we found that high BCR-ABL/GUSIS levels at diagnosis were associated with inferior probabilities of OR (p<0.001), FFS (p<0.001) and EFS (p<0.001). Elevated BCR-ABL/GUSIS levels were also associated with higher rates of disease transformation to the accelerated phase or blast crisis (p=0.029) but not with OS (p=0.132).
Conclusions
High BCR-ABL transcripts at diagnosis measured by RQ-PCR employing GUS as a reference gene allow the identification of CML patients unlikely to benefit from standard dose IM that should be considered for alternative forms of treatment.

Disclosures
Hochhaus: Novartis: Honoraria, Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; ARIAD: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding.
HH-002 Effectively Suppresses Myeloproliferative Neoplasm Phenotypes in JAK2V617F Transgenic Mice
Authors: Ho Wan-Ting , Zhao Wanke , Hallenbeck Paul , Rong Frank , Zhao Zhizhuang Joe 
Published: October 2019

Abstract
Ph-negative myeloproliferative neoplasms (MPNs) represent a group of conditions characterized by chronic increases in some or all of the blood cells (platelets, white blood cells, and red blood cells). A major molecular defect in MPNs is JAK2V617F, a gain-of-function mutant form of tyrosine kinase JAK2 found in the majority of MPN patients. In earlier studies, we generated JAK2V617F transgenic mice by using the vav promoter which drives gene expression in all hematopoietic cells. These mice develop MPN-like phenotypes in a transgene dose-dependent manner. The objective of this study is to use this animal MPN model to test the efficacy of HH-002, a derivative of natural plant alkaloid homoharringtonine.
Treatment of JAK2V617F transgenic mice with a daily dose of 1 mg/kg HH-002 through subcutaneous injection reduced blood cell counts to the normal range or slightly below the normal level. HH-002 causes a preferential reduction of myeloid cells in JAK2V617F transgenic mice since percentages of lymphocytes (CD3e+ T cells and CD19+ B cells) were increased (despite somewhat decreases in absolute numbers) while the percentage of Gr-1+/CD11b+ granulocytes sharply declined. HH-002 also effectively reduced the spleen size of JAK2V617F transgenic mice and prevented development of myelofibrosis. With a JAK2V617F bone marrow transplant mouse model, HH-002 showed a similar effect. Although it did not increase the ratio of JAK2V617F-negative cells to JAK2V617F-positive, it stopped further expansion of JAK2V617F-containing malignant cells in JAK2V617F bone marrow recipient mice. In vitro experiments with primary hematopoietic cells demonstrated that HH-002 potently inhibited formation of erythroid and myeloid colonies with an IC50 value of 1-3 nM. However, it does not show a preferential inhibition of JAK2V617F-containing cells. This is not unexpected since homoharringtonine is known to inhibit protein synthesis. Taken together, HH-002 is a promising candidate for development of therapeutic drugs to treat MPNs and related diseases. Combination drug therapies with JAK2 inhibitors need to be explored.

Disclosures
Hallenbeck: Hangzhou Bensheng Pharmaceutical Corp. Ltd.: Employment, Equity Ownership. Rong:Hangzhou Bensheng Pharmaceutical Corp. Ltd.: Employment, Equity Ownership. Zhao:Hangzhou Bensheng Pharmaceutical Corp. Ltd.: Research Funding.
A Patient-Derived EZH2 Mutant Induces MDS-like Diseases with Derepressed ABCG2 Expression in Mice
Authors: Kawabata Kimihito Cojin , Inoue Daichi , Kitaura Jiro , Harada Yuka , Goyama Susumu , Harada Hironori , Aburatani Hiroyuki , Kitamura Toshio 
Published: October 2019

Abstract
A histone H3 Lysine 27 (H3K27)-methyltransferase, enhancer of zeste homolog 2(EZH2) is known as a tumor-associated gene. Physiological role of EZH2 is an enzymatic component of polycomb repressive complex 2 (PRC2) to inhibit expression of target genes. While EZH2 plays oncogenic roles by repressing the expression of tumor suppressors in solid tumors and some lymphomas, it plays rather tumor-suppressive roles in myeloid malignancies. We have generated a short-form EZH2 that lacks the catalytic SET domain (EZH2-dSET). Using this EZH2 mutant we could produce serially transplantable MDS-like diseases. Microarray analysis using the MDS-like bone marrow cells enabled us to identify novel targets of EZH2 in MDS tumorigenesis, including ATP-binding cassette (ABC) transporters. Derepression of Abcg2 via decreased H3K27-trimethylation was confirmed. Retroviral transduction of EZH2-dSET to MDS-like cell lines increased surface ABCG2-high populations and those cells functioned to exclude anticancer drugs as expected. Intriguingly, with Abcg2 expression alone, primary bone marrow cells could produce an MDS-like cytopenic disease in our BMT model. In our clinical specimens, ABCG2 high expressions were observed in MDS samples but not in de novo AML and CML samples. In two MDS patients, ABCG2 expression decreased along with leukemic transformation. Interestingly, two out of 33 MDS patients with extremely high expression of ABCG2 harbored the same U2AF1 mutation (Q157P). In addition, somatic mutations of EZH2 and those of either U2AF1 or SRSF2 were mutually exclusive in all investigated cases. Interestingly, U2AF1 mutants (S34F and Q157P) reduced EZH2 expression, leading the derepression of ABCG2 via decreased H3K27-trimethylation. These results indicate a link between U2AF1 mutations and ABCG2 expression via disrupted EZH2. In conclusion, different mechanisms are supposed to converge at dysregulated EZH2 in MDS. And a short form of EZH2 upregulates ABCG2 expression resulting in MDS advancing to secondary leukemia. Thus, either mutations affecting the EZH2 function or mutations of EZH2 itself could play an important role in MDS, and one of the downstream targets of EZH2 suppression in MDS pathogenesis is aberrant expression of ABCG2.

Disclosures
No relevant conflicts of interest to declare.
No Mutations of SF3B1, U2AF1, and SRSF2 spliceosomal Gene but Polymorphisms Associated with the Worse Prognosis in Multiple Myeloma
Authors: Kang Min-Gu , Kim Hye-Ran , Lee Jun Hyung , Ganapathy Thashma Pemmanda , Yang Seung-Hyeon , Hoque Moinul , Park Ra-Young , Kee Seung Jung , Kim Soo Hyun , Shin Jong-Hee , Suh Soon-Pal , Shin Myung-Geun 
Published: October 2019

Abstract
Recently, a striking example of the effects of acquired somatic mutations in splicing factors such as SF3B1, U2AF1, and SRSF2 has been described. The DNA sequences from abnormal blood cells of patients with hematologic malignancies has shown that a high proportion of these diseases are associated with somatic mutations of spliceosomal proteins. However, little is known about the aberrant splicing pathways in multiple myeloma (MM) patients. Therefore, this study investigated the presence and prognostic implication of splicing gene aberration in MM patients. Also, we planned functional experiments in cultured normal and Hela cell line to determine the effects of the common spliceosome mutations on cellular growth rate, cell count, and apoptosis. In total, 87 MM patients and 100 healthy controls were examined for somatic mutations in SF3B1, U2AF1, and SRSF2 by direct sequencing. Also, the clinical and laboratory characteristics of MM patients were reviewed. Regarding to the initial findings in recent reports, this study expressed the wild-type and the mutant (Q157P) U2AF1 in Hela cells and examined their proliferation.
MM patients did not show mutation in SF3B1, U2AF1,and SRSF2 splicing genes. However, the patients displayed 39198T>C or 39198T>T/C polymorphism (70.1 %) in exon 18 of SF3B1, 8345T>T/G polymorphism (13.8 %) in exon 2 of U2AF1, and 5399C>T or 5399C>C/T polymorphism (100.0 %) in exon 1 of SRSF2 (Fig. 1). In the entire cohort, the number of patients with one polymorphism, two polymorphisms, and three polymorphisms was counted up to 24.1 %, 67.8 %, and 8.0 %, respectively. Meanwhile, in 100 normal controls, polymorphism of SF3B1 exon 18 and U2AF1 exon 2 was counted up to 82.0 % and 10.0 %, respectively. Free light chain (FLC) kappa/lambda (K/L) ratio and chromosome analysis showed significant difference according to polymorphism status (p=0.002) while other clinical characteristics did not. The patient with polymorphisms in both SF3B1 and U2AF1 gene had unfavorable overall survival (p =0.067) and disease-free survival (p=0.001), compared to patients without polymorphism.
Our results show no recurrent SF3B1 and U2AF1 mutations in MM patients. Instead, polymorphisms were found in SF3B1 or U2AF1, which were significantly implicated in worse prognosis.
Figure 1. Figure 1.

Disclosures
No relevant conflicts of interest to declare.
Examining the Impact of Altered Protein Expression and Ubiquitination Levels on the Development of Resistance to Proteasome Inhibitors Using Proteomics Analysis
Authors: Dowling Paul , Ting Kay Reen , Meiller Justine , Kelly Shane , Clynes Martin , O'Gorman Peter 
Published: October 2019

Abstract
The range of new proteasome inhibitors currently in clinical or preclinical development hold tremendous promise for the treatment of haematological malignancies, in particular multiple myeloma, both in frontline and relapsed or refractory settings. However the development of resistance to proteasome inhibitors is a significant problem. To facilitate the analysis of this phenotype, a carfilzomib-resistant cell line was established and the effects on altered protein expression and ubiquitination levels determined. Label-free quantitative mass spectrometry was performed on whole cell extracts from the resistant model together with ubiquitin remnant motif antibody K-ε-GG enriched peptides for modification analysis. A significant number of proteins with altered expression and ubiquitination levels were positively identified. In particular, the proteasome maturation protein (POMP) was found to have a significantly up-regulated abundance level in the resistant phenotype, a protein essential for the assembly of standard proteasomes and immunoproteasomes (Figure 1). Abnormal ubiquitination levels were also detected on proliferating cell nuclear antigen (PCNA), significantly on Lysine-164 which is involved in DNA-repair pathways. Identifying the relevant biological pathways affected by proteasome inhibitors and the changes which mediate drug resistance may allow for optimal therapeutic use and potential new drug targets. POMP and PCNA may be important target molecules to help overcome the establishment of resistance to proteasome inhibitors in the treatment of multiple myeloma. Further work could examine broadening the therapeutic application of proteasome inhibitors that is currently confined to multiple myeloma and identifying the key proteins involved in common resistance mechanisms.

Disclosures
No relevant conflicts of interest to declare.
Cord Blood Unit Cryopreservation: Positioning Segments for Potency Assessment
Authors: Shoulars Kevin , Gentry Tracy , Noldner Pamela , Page Kristin M , Kurtzberg Joanne 
Published: October 2019

Abstract
Introduction: Banked, unrelated umbilical cord blood units (CBU) are a rich source of hematopoietic stem cells for hematopoietic reconstitution. Potency assays on attached segments have been used for CBU release from the bank to the transplant center. CBUs have historically been manufactured and cryopreserved in bags with three segments (made from the tubing used to fill the bag) arranged across the top of the cassette. Prior to cryopreservation, the CBU is overwrapped, placed in a metal cassette and frozen in a control-rate freezer and submerged under liquid nitrogen for long-term storage. After cryopreservation, segments can be removed from the cassette, without disturbing the bag, for HLA typing, potency assays, or other testing. Last year, in an effort to increase the number of segments available for testing, we changed the configuration of the cryobag to create a fourth segment. To fit in the cassette, the segments were arranged with two segments across the top and two segments bent at 90 degrees to fit between the 20% and 80% portions of the bag. When performing our annual stability testing, we realized that the two distal segments, positioned between the 20% and 80% compartments yielded lower potency results than the segments positioned across the top of the cassette. We investigated this finding as described below.
Method: Three large fresh CBUs were processed separately on a Sepax cell processor (Biosafe, Geneva, Switzerland) and evenly divided into two cryopreservation bags. For each CBU, one bag was created with three segments and the other bag with four segments. The cryopreservative, dimethylsulfoxide in dextran, was added at a final concentration of 10% to both bags prior to cryopreservation. Segments were positioned either across the top (3 segments) or in the bent configuration (4 segments) for each of the paired CBUs. All units were cryopreserved and stored in a Thermogenesis Bioarchive. After two weeks in storage, the CBUs were tested. The bags and segments were assayed for ALDHbr, viable CD34, CD45, glycophorin A and viability (7-AAD) along with colony forming units (CFUs), the potency assay used by our bank.
Results: The segments configured across the top of the bag correlated with results from the cryobag for ALDHbr (as a percent of viable CD45+, Table 1), CD34+ (as a percent of viable CD45+) and CFU. In contrast, segments placed between the 20% and 80% compartments were significantly lower than the bag for both ALDHbr (as a percent of viable CD45+) and CFU, although CD34+ (as a percent of viable CD45+) was equivalent.
Conclusions: Segments attached to CBUs can predict potency of the CBU by various parameters. However, proper placement of the segments within the cryopreservation cassette must be performed to ensure that potency measured on the segments reflects potency measured on the cord blood unit.

Disclosures
No relevant conflicts of interest to declare.
Intrathecal Rituximab for EBV-Associated Post-Transplant Lymphoproliferative Disorder with Central Nervous System Involvement Unresponsive to Intravenous Rituximab-Based Treatments: A Prospective Study
Authors: Liu Qifa , Wu Meiqing , Sun Jing , Zhang Yu , Huang Fen , Zhou Hongsheng , Fan Zhiping , Xuan Li , Yu Guopan , Guo Xutao , Dai Min , Feng Ru 
Published: October 2019

Abstract

Backgroud:
Although the introduction of rituximab as a first-line treatment has improved outcome of post-transplant lymphoproliferative disorder (PTLD), PTLD with central nervous system involvement (CNS-PTLD) still has a dismal prognosis because of low penetrance across the blood-brain barrier. In this prospective study, we reported intrathecal rituximab was efficacy and safety in the patients with CNS-PTLD who had failed to respond to the intravenous rituximab-based treatments.
Methods:
From June 2009 to November 2013, 32 cases of EBV-associated PTLD were recorded in the Southern Medical University Institute of Hematology. Fourteen patients diagnosed with CNS-PTLD were enrolled in this prospective study. For the patients who failed to response to the initial intravenous rituximab-based treatments, sequential dose-escalation schedule of intrathecal rituximab (initial dose of 20mg, increased by 10mg each week and maximum dose of 50 mg) was administrated weekly.
Results:
Three patients were responsive and 11 were unresponsive to the initial treatments within one week after the treatments. For the 11 patients who failed to respond to the initial treatments, 9 patients received intrathecal rituximab within 7-11 days and 2 patients refused the treatment. After two cycles of rituximab-based treatments, 10 patients achieved complete response (CR), 2 patients were partial response and 2 patients were non-response. With a median follow up of 664 (range 18 to 1545) days after the diagnosis of CNS-PTLD, 7 patients survived and 7 died. The causes of death included PTLD progressing (n=3), PTLD relapse (n=1), GVHD (n=1), CMV pneumonia (n=1) and pseudomonas aeruginosa sepsis (n=1). The 3-year probability of OS was 45.7% ±14.7% in CNS-PTLD, which was no significant difference as compared to PTLD without CNS involvement(55.6% ±11.7%, P=0.706).
Conclusion:
Intrathecal rituximab might be an effective and safe method for CNS-PTLD in the allo-HSCT recipients who were unresponsive to the intravenous rituximab-based treatments.

Disclosures
No relevant conflicts of interest to declare.
To Treat or Not to Treat: Metabolomics Reveals Biomarkers for Treatment Indication in Chronic Lymphocytic Leukaemia Patients
Authors: Piszcz Jaroslaw , Armitage Emily Grace , Alessia Ferrarini , Ruperez Francisco Javier , Kulczynska Agnieszka , Bolkun Lukasz , Kloczko Janusz , Kretowski Adam , Ciborowski Michal , Barbas Coral 
Published: October 2019

Abstract
In chronic lymphocytic leukaemia (CLL) clinical course of patients is heterogeneous. Some present an aggressive disease onset and require immediate therapy, while others remain without treatment for years. Current disease staging systems developed by Rai and Binet may be useful in forecasting patient survival time, but do not discriminate between stable and progressive forms of the disease in the early stages. Recently ample attention has been directed towards identifying new disease prognostic markers capable of predicting clinical aggressiveness at diagnosis. In this research we reached for LC-MS metabolic fingerprinting method to analyse serum of stable (n=51) and progressive (n=42) CLL patients and controls (n=45) with aim to discover metabolic indicators of disease status. A panel of markers discriminating aggressive from indolent patients was discovered. Ten of them were selected for validation on larger population (45 controls and 92 CLL) with an independent analytical technique. Linoleamide (p=0.002) in addition to various acylcarnitines (p=0.001-0.000001) showed to be significant markers of CLL in its aggressive form. Acetylcarnitine (p=0.05) and hexannoylcarnitine (p=0.005) were also distinguishable markers of indolent subjects. Forming a panel of selected acylcarnitines and fatty acid amides, it was possible to reach a highly specific and sensitive diagnostic approach (AUC=0.766).

Disclosures
No relevant conflicts of interest to declare.
Outcome of Patients with Hodgkin Lymphoma transformed from Chronic Lymphocytic Leukemia
Authors: Mocikova Heidi , Markova Jana , Gaherova Lubica , Smolej Lukas , Simkovic Martin , Kozak Tomas 
Published: October 2019

Abstract
Background. Hodgkin lymphoma (HL) that develops in the course of chronic lymphocytic leukemia (CLL) is a rare event. Optimal management of these patients (pts) is not defined. We analyzed outcome of 10 pts treated for Hodgkin´s variant (HV) of Richter syndrome.
Patients and methods. Overall 10 pts (8 males) with CLL and subsequent HV were treated between 1996 and 2015. Median age at diagnosis of HV was 68 (range 54 - 83) years. Prognostic markers of CLL at diagnosis: unmutated IGHV (5 pts), del(17p) or TP53 mutations (0 pt), del(11q) (2 pts), deletion of 13q14 region (4 pts), trisomy 12 (2 pts), ZAP 70 pos. (6 pts), CD38 pos.(4pts). The diagnosis of HV included various subtypes of HL: nodular sclerosis (3pts), mixed cellularity (3pts), lymphocyte rich (1pt), not othervise specified (3pts). EBV status of pts at diagnosis of HV of Richter syndrome: 6 pos., 1 negat. nad 3 not done. Initial and second-line treatments of CLL consisted of fludarabine- based regimens combined with rituximab or alemtuzumab and 3 pts received chlorambucil. Treatment of HV of Richter syndrome included: ABVD (7 pts) in combination with rituximab (2 pts), COPP (3 pts) and involved field radiotherapy of 30 Gy after chemotherapy (2 pts). Further treatment was required in 6 pts due to insufficient response with persistent CLL: rituximab alone (2 pts), R-bendamustine (1pt), R-DHAP (1 pt), cyclophosphamide and dexamethasone (1 pt) and gemcitabine (1 pt). None of these patients underwent autologous or allogeneic stem cell transplantation.
Results. The median time from CLL diagnosis to development of HV was 8.4 (range 2.6 - 16.5) years. Median overall survival from diagosis of CLL was 17.3 years and median survival after diagnosis of HV was 3.5 years. Out of 10 pts 5 are alive :3 in complete remission (CR), 1in partial remission of CLL (HL in CR), 2 in progression of CLL (HL in CR). Five pts died (3 lymphoma progressions, 1 second cancer, 1 unknown causality).
Conclusion. Our results indicate a long period for developing of HV of Richter syndrome and its subsequent poor outcome. Current HL based chemotherapy is not sufficient in HV of Richter syndrome and new treatment approaches should be considered.

Disclosures
No relevant conflicts of interest to declare.