bluuebunny/crossref_metadata_2025_split huggingface dataset:
A New Prognostic Index for Multiple Myeloma Patients in the Era of Novel Agents
Authors: Cho Hyungwoo , Suh Cheolwon , Yoon Dok Hyun , Park Ji Hyun , Kim Shin , Lee Sang-wook , Park Chan-Sik , Huh Jooryung , Lee Kyoungmin , Kang Eun Hee , Park Jung Sun , Park Chan-Jeoung , Kim Jin Seok , Kim Kihyun 
Published: October 2019

Abstract
Introduction
Various prognostic factors for multiple myeloma (MM) have been identified, and combinations of these factors were suggested for staging classification of MM patients. The international staging system (ISS) is a widely accepted prognostic system for MM patients. However, since ISS was established using data collected from 1981 through 2002, only a small portion of patients who received the novel agents (e.g., thalidomide, bortezomib, and lenalidomide) were included. Therefore its validity is being challenged in the era of novel agents. We aimed to develop an alternative prognostic index for MM patients based on easily obtained laboratory measures in the era of novel agents.
Patients and methods
Data were collected from February 2001 through June 2012 at Asan Medical Center. 470 patients were newly diagnosed as symptomatic MM and 156 of them were treated with novel agents including thalidomide, lenalidomide and bortezomib. We analyzed these 156 patients who received novel agent therapy at least once, using prospectively collected data. A new prognostic model was developed based on significant potential prognostic factors in univariate and multivariate analysis, which was validated in external validation data set from two tertiary hospitals in Korea.
Results
With a median follow-up duration of 27.9 months (range, 0.2-122 months), the median overall survival in the whole population was 28.7 months. The median overall survival (OS) of patients according to ISS stage 1, 2 and 3 were 122.2, 43.9 and 30.0 months, respectively. There were no significant differences in median OS between ISS stage 1 and 2 patients and 2 and 3 patients (Figure 1). Furthermore, ISS was not statistically significant in the multivariate analysis. We incorporated serum beta2-microglobulin (Sβ2M), serum albumin and serum lactate dehydrogenase (LDH) as independent risk factors to develop a new prognostic model. Using these three factors, we stratified the patients into three groups; Group 1, Sβ2M < 5.5 mg/L plus serum albumin >= 3.5 g/dL plus serum LDH normal; Group 2, neither group 1 nor 3; Group 3, Sβ2M >= 5.5 mg/L plus serum albumin < 3.5 g/dL plus serum LDH > normal. We found that there were significant differences in median OS among these three groups (122.2, 41.6 and 5.6 months for group 1, 2 and 3, respectively) (Figure 2). This new prognostic index was further validated in the external cohorts (Median OS was 75.2, 36.3 and 23.1 months for group 1, 2 and 3, respectively) (Figure 3).
Conclusion
Our study suggests that this new prognostic index based on easily obtainable variables (Sβ2M, serum albumin, serum LDH) might be a simple and useful tool to predict prognosis for patients with MM in the era of novel agents, which warrants further validation.

Disclosures
No relevant conflicts of interest to declare.
DOCK2 Interacts with FLT3 and Modulates the Survival of FLT3-Expressing Leukemia Cells
Authors: Wu Min , Hamaker Max , Li Li , Small Donald , Duffield Amy S. 
Published: October 2019

Abstract
The FMS-like tyrosine kinase-3 (FLT3) receptor gene is the most commonly mutated gene in acute myeloid leukemia (AML), and patients carrying FLT3/ITD mutations have a poor prognosis. Despite continuing progress in the development of more effective FLT3 inhibitors, long-term success in inhibition of FLT3 activity in AML patients is still elusive. In order to achieve a better understanding of FLT3 biology and more effective strategies for the inhibition of FLT3 activity, a screen was performed on leukemia cell lines to search for FLT3-interacting proteins. One of the proteins identified in the screen was dedicator of cytokinesis 2 (DOCK2).
The DOCK family of proteins acts as guanine nucleotide exchange factors (GEFs) for Rho family of GTPases, which includes Rac GTPases. DOCK2 expression is limited to hematopoietic cells, and is known to regulate several crucial processes, including lymphocyte migration, activation and differentiation of T cells, and cell-cell adhesion and bone marrow homing of various immune cells.
We first verified that DOCK2 is expressed in primary AML samples from patients, and co-immunoprecipitation experiments showed that DOCK2 interacts with both wild-type FLT3 and FTL3/ITD in these cells. Co-immunoprecipitation experiments using leukemia cell lines demonstrated that DOCK2 interacts with FLT3, FLT3/ITD, FLT3/D835Y and FLT3/D835H, and that it predominantly interacts with the unphosphorylated form of FLT3. Knock-down of DOCK2 by shRNA did not significantly affect the growth of cell lines that lack expression of FLT3, but greatly reduced growth of cell lines expressing amplified wild type FLT3 (Sem K2), FLT3/D835H (HB11;19) and FLT3/ITD (MV4;11). Accordingly, colony formation assays revealed that cell lines with elevated expression of wild type or mutant FLT3 produced fewer, smaller and more compact colonies when the expression of DOCK2 was decreased, while colonies from cell lines lacking FLT3 expression showed no significant difference in response to the knock-down of DOCK2. Furthermore, an Annexin V binding assay indicated that reduction in DOCK2 expression level greatly sensitized cells with elevated FLT3 activity (MV4;11 and Sem K2) to cytarabine, resulting in increased apoptosis, but no significant sensitization was observed in cell lines that lack FLT3 expression.
These findings demonstrate that DOCK2 interacts with FLT3 in leukemia cell lines, and suggest that this interaction has important roles in regulating the survival of leukemia cells with elevated FLT3 activity, both alone and in combination with conventional anti-leukemic agents. Therefore, DOCK2 is a potential therapeutic target for AML treatment, and better understanding of the interaction between DOCK2 and FLT3 may contribute to the development of novel strategies to effectively inhibit FLT3 activity in AML patients.

Disclosures
No relevant conflicts of interest to declare.
Frequency and Prognosis of JAK2 V617F, Calr, MPL and ASXL1 Mutations in Primary Myelofibrosis
Authors: Sorigue Marc , Cabezón Marta , Garcia Olga , Velez Patricia , Marce Silvia , Boqué Concha , Xicoy Blanca , Fernandez Cristalina , Xandri Marisol , Casanova Enric , Gallardo David , Milla Fuensanta , Ribera Josep-Maria , Feliu Evarist , Zamora Lurdes 
Published: October 2019

Abstract
INTRODUCTION: Several mutations have been described in patients with BCR-ABL1 –negative chronic myeloproliferative neoplasms, including primary myelofibrosis (PMF). The most frequent mutation is JAK2 V617F, followed by calreticulin exon 9 (CALR), MPL exon 10 and ASXL1 exon 12. Currently, less than 10% of patients lack molecular marker. CALR and ASXL1 mutations have been consistently found to have favorable and unfavorable prognostic implications, respectively.
The aim of this study was to describe the frequency and prognostic impact of JAK2 V617F, CALR, MPL and ASXL1 mutations in patients diagnosed with PMF in 6 Spanish hospitals.
METHODS: To detect the presence of JAK2 V617F mutation, an allele-specific PCR using TaqMan probes was used. Screening for insertions and deletions in CALR gene was performed with 6-FAM labeled primers spanning exon 9 and CALR mutations were described by Sanger sequencing. Sanger sequencing was also used to detect MPL exon 10 and ASXL1 exon 12 mutations.
RESULTS: Sixty-eight patients were included in the study. All of them were screened for JAK2 and CALR mutations. Forty-five of them (66%) were positive for the JAK2 V617F mutation, while 11/68 (16%) were positive for CALR mutations. Of the 11 CALR mutations, 10 were JAK2 wild-type. MPL exon 10 mutation analysis was only performed in JAK2 wild-type patients and was positive in 4/23 patients (17%), and all of them were CALR wild-type. At the time of submission, ASXL1 exon 12 has been assessed in 18 patients (analysis in the rest of them is currently ongoing). ASXL1 mutations have been found in 3/18 (17%) patients, two of them also with a CALR mutation and the other one with the JAK2 V617F mutation. All three cases were indel mutations. Overall, no mutation was detected in 9/68 (13%) patients. JAK2 V617F, CALR and MPL mutations had no prognostic impact on overall survival. The effect of ASXL1 mutation on prognosis (with and without CALR mutation) will be assessed once all samples have been sequenced.
CONCLUSION: JAK2 V617F, CALR and MPL mutations were found in our series of PMF patients in the same proportion found in larger series. ASXL1 has so far been found in a smaller percentage but the entire series of patients will need to be sequenced before reaching definitive conclusions. Studying these genes, only 13% of patients with PMF did not have a clonal marker. None of the studied mutations had prognostic significance.
ACKNOWLEDGMENTS: The authors would like to thank Diana Dominguez for her excellent technical assistance and to the grant 2014 SGR225 (GRE), Generalitat de Catalunya.

Disclosures
No relevant conflicts of interest to declare.
Calreticulin Gene Mutations in Suspicion of Myeloproliferative Neoplasms: A Single Institution Experience
Authors: Ben Lassoued Amin , Beaufils Nathalie , Bonnefoy Caroline , Gabert Jean 
Published: October 2019

Abstract
Recently, Klampfl et al. and Nangalia et al. reported recurrent somatic mutations in the calreticulin (CALR) gene in patients with essential thrombocythemia or primary myelofibrosis with nonmutated JAK2 and MPL. All CALR mutations described to date are either deletions or insertions, and occur in exon 9 and result in a frameshift leading to a loss of KDEL sequence (endoplasmic reticulum retention motif) and multiple calcium biding sites with a new basic (instead of acidic) C terminal region.
The aim of this work is to know how far the CALR mutations can fill the molecular diagnostic gap for myeloproliferative neoplasms (MPNs) left by JAK2 and MPL mutations.
The study was approved by the local ethics committee.
Our study population consists of 155 patients with platelet counts > 500 G/L sampled between 2012 and 2013 for suspicion of MPNs. Only JAK2 V617F negative patients were included.
Analyses were performed using genomic DNA isolated from peripheral blood. Patients were screened for CALR mutations using a High Resolution Melting (HRM) and then the precise mutations were characterized by Sanger sequencing which is a new approach for this test.
A total of 21 (13.5%) patients with CALR mutations were detected with 8 distinct variants.
Among CALR mutations, type 1 (52 basepair deletion, c.1092_1143del) and type 2 (5 basepair insertion, c.1154_1155insTTGTC) were the most common (respectively 7 (33%) and 7 (33%) cases). Three patients had 2 concurrent mutations (insertion+substitution / Deletion+substitution / deletion +insertion). We found a deletion that has not been reported so far to our knowledge (c.1125_1147del). Sequencing is ongoing for 4 patients.
Compared with negative patients, CALR-mutated patients demonstrated markedly higher platelet counts (median count: 820 vs 543 G/L).

Disclosures
No relevant conflicts of interest to declare.
Human BDH2 As a Predictor for Leukemia Progression in Myelodysplastic Syndrome
Authors: Yang Wen-Chi , Lin Sheng-Fung 
Published: October 2019

Abstract
Objective: Myelodysplastic syndrome (MDS) is a disorder of hematopoietic stem cells. In MDS patients, leukemia progression is associated with iron overload. Hydroxybutyrate dehydrogenase type 2 (BDH2) catalyzes the production of 2,5-dihydroxybenzoic acid (DHBA), an iron-binding component.
Methods: The bone marrow (BM) of 187 MDS patients, 119 de novo acute myeloid leukemia (AML) patients, and 43 lymphoma patients (stage I-III) with normal BM were analyzed. BDH2mRNA expressions from BM and BDH2-KD THP1 were analyzed using quantitative reverse-transcription polymerase chain reaction (qRT-PCR). Cell cycle and cell maturation were determined by flowcytometry and surface markers in RNA interference–mediated-knockdown of BDH2 (BDH2-KD) THP1, an acute leukemia cell line. Immunohistochemistry was performed to detect BDH2 protein expression in the bone marrow.
Results: Elevated BDH2mRNA expression was observed in bone marrow in MDS patients (P=0.009) and was related to ferritin levels (P=0.049). The higher BDH2 expression (15%) group showed a greater risk for leukemia progression than the lower expression group (3.18%) (P=0.017). Leukemia-free-survival was longer in patients with low BDH2mRNA expression (P=0.028) and patients with ferritin level <1000 ng/mL. Additionally, cell cycle analysis, surface markers, and special stain studies indicated that BDH2-KD induced differentiation and decreased the growth rate of THP1 cells, which was associated with the retardation of cell cycle.
Conclusions: We identified new molecular mechanisms of BDH2. BDH2 is involved in cell cycle arrest and inhibition of differentiation in malignant cells. Further, high BDH2 expression in MDS patients suggest that it could act as a poor prognostic factor.

Disclosures
No relevant conflicts of interest to declare.
Evaluation of Azacitidine Clinical Efficacy in Patients with Myelodysplasia Syndrome Dudina G.a., Golenkov a.K., Kataeva E.V., Mitina T.a. Moscow Regional Clinical Research Institute
Authors: Golenkov Anatoly 
Published: October 2019

Introduction: The anti-tumor effect of azacitidine in patients with myelodysplasia syndrome is based on cytotoxicity against pathologically changed hemopoetic cells of bone marrow and DNA hypomethylation.
Objective: evaluation of clinical efficacy of azacitidine in patients with myeloddysplasia syndrome.
Materials and methods: we have monitored 7 patients with myelodysplasia syndrome and refractory anaemia with excessive blast count, type II. IPSS with a high risk of acute leucosis development was found in 4 patients; intermediate risk was reported in 3 patients. Mean age – 62 years. Chromosomal aberrations (multiple clone transformations) were found in 4 patients during cytogenetic analysis. Normal karyotype was reported in 3 patients. Treatment regimen: azacitidine 75 mg/m² daily s/c injections, Days 1 - 7. Totally, 3 – 7 induction cycles were conducted, median (Me) – 4 cycles. Further maintenance treatment with 6-8 weeks interval was conducted in 5 patients. The mean index of haemoglobin level decrease was used to evaluate the hemopoetic ability in non-selective group; it was calculated as the ratio of the haemoglobin level at the time of discharge minus haemoglobin level at the time of the next hospitalization to the duration of the interval between treatment courses. The number of blood transfusions during the interval between treatment courses was constant: 2-3 transfusions of RBCs.
Results: 4 azacitidine treatment courses resulted in decrease of blast cells number in bone marrow from 16.4 ± 0.2 % to 5.4% ± 0.7 % (р<0.05). Mean index of haemoglobin level decrease after the first hospitalization was 1.9 g/l per day and 0.3 g/l per day after the 4-th treatment course (р<0.05) The following results were reported using standard Cheson criteria: full response – 1 patient, hematologic improvement – 2 patients, bone marrow remission – 3 patients. Mean duration of response 8.5 months. Treatment complications: pleural empyema, abscess of left lung lower lobe after the 2-nd treatment cycle – 1 patient; broncho-pulmonary aspergillosis after 3-rd treatment cycle – 1 patient. In other patients drug toxicity that could possibly affect the interval between courses or decrease the drug dose was not reported. Delayed treatment results: 100% of patients survived the 12 – months period.
Conclusions: in patients with myelodysplasia syndrome and refractory anaemia with excessive blast count, type II treatment with azacitidine (median treatment course duration – 4 months) resulted in full response in 14% of cases (1 patient), bone marrow remission was reported in 43% of cases (3 patients). Use of original method of haemoglobin level decrease index determination enables evaluation of the cumulative azacitidine dose effect on hemopoesis recovery after 4 treatment cycles.

Disclosures
No relevant conflicts of interest to declare.
The Role BAFF and BAFF Receptors in Macrophage Induced Drug Resistance of Multiple Myeloma
Authors: Chen Jing , He Donghua , Guo Xing , Yang Li , Li Yi , Chen Qingxiao , He Jingsong , Yang Yang , Zhang Enfan , Wu WenJun , Han Xiaoyan , Zheng Gaofeng , Zhao Yi , Huang He , Cai Zhen 
Published: October 2019

Abstract
Background: B-cell–activating factor (BAFF) is a member of TNF family that critical for maintenance of B-cell development and homeostasis. BAFF also modulates the proliferation, survival and drug resistance of multiple meyloma (MM) cells. BAFF can be secreted by neutrophils, monocytes,dentritic cells and macrophages. Our previous study showed that, macrophages protect MM cells from drug-induced apoptosis by a cell-to-cell interaction between MM cells and macrophages. We supposed that the interaction of BAFF and BAFF receptors plays a role in macrophages induced MM resistance.
Methods: First, we detected the expression levels of BAFF and its three receptors in MM cells were detected by semiquantitative real time-polymerase chain reaction (qPCR) and flow-cytometry. The concentration of BAFF in the culture supernatants of MM cell lines was detected by ELISA. Second, we collect peripheral blood monocytes form healthy donors. Monocytes were induced into macrophages by culturing with M-CSF for 7 days.BAFF expression level in macrophages was detected by qPCR, flow-cytometry and ELISA. Then MM cells were sole-cultured or coculture with macrophages for the indicated time (usually 24 h), after that bortezomib was added to the culture system. Cell viability and apoptosis of MM cells were verified by MTT and flow cytometry.At last,the recombinant human BAFF were added to MM cells, MTT and flow cytometry were used for detection of cell viability and apoptosis of MM cells.
Results: Two receptors of BAFF, transmembrane activator and CAML interactor (TACI) and B-cell maturation antigen (BCMA), are highly expressed in various MM cell lines evidenced by real-time PCR and flow cytometry. The expression of BAFF in PBMC-induced macrophages are heterogeneous. We verify macrophage-mediated MM drug resistance by directly coculturing MM cells (ARP-1, 8226 and OPM2) with PBMC-derived macrophages from healthy donors. Functional studies show that recombinant human BAFF rescues RPMI8226 myeloma cells from dexamethasone-induced apoptosis.
Conclusions: Our data showed that macrophage might induce drug resistance of MM cells by the interaction of BAFF and BAFF receptors. Further study will focused on the mechanism of interaction between BAFF and BAFF receptors.

Disclosures
No relevant conflicts of interest to declare.
Antitumor Activity of a Novel Proteasome Inhibitor BSc2118 Against Multiple Myeloma
Authors: Zang Meirong , Liu Lanting , Li Xin , Li Wei , Kuckelkorn Ulrike , Zhou Wen , Qiu Lugui , Jin Fengyan 
Published: October 2019

Abstract
Background: Although the first generation of proteasome inhibitor bortezomib is very effective, the development of resistance limits it long-term utility. In this study, we explored the efficiency and molecular mechanisms of the novel, irreversible proteasome inhibitor BSc2118, particularly, the reversal of bortezomib resistance.
Materials and Methods: Human MM cell lines (MM.1S, MM.1R, RPMI-8226, U266, NCI-H929) were treated with BSc2118 at various concentrations for 48h, and assessment for cell viability by CCK-8 assay. MM.1S and MM.1R cells were treated with BSc2118 for 24 hours, and cell cycle and apoptosis analysis were conducted by flow cytometry. Associated molecules were detected by qRT-PCR and western blot. Chymotrypsin-like proteasome activity assay was performed by using the 20S proteasome assay kit. Ubiquitinated proteins were isolated and determined with ubiquitin enrichment kit.
Results: Our results revealed that treatment of MM cell lines with BSc2118 inhibits the chymotrypsin-like proteasome activity and induces accumulation of ubiquitinated proteins. BSc2118 inhibits MM cell growth and induces MM apoptosis via induction of G2/M phase arrest, activation of cleaved caspase-3, caspase-8 and caspase-9 and PARP, increasing p53, p21 and E2F1, and inhibition of autophagy in MM.1S, MM.1R and RPMI-8226 cell lines. In addition, BSc2118 dramatically inhibits cytokines mRNA, such as IL-6, VEGF and bFGF in both myeloma cells line and primary bone marrow stromal cells from myeloma patients. More importantly, BSc2118 could overcome bortezomib resistance in vitro by using primary CD138 positive plasma cells from bortezomib-resistant myeloma patients and bortezomib resistance cell line ANBL-6 (ANBL-6.BR), most likely as the consequence of inhibition of autophagy flux which is responsible for bortezomib resistance.
Conclusion: Our study revealed BSc2118, a novel irreversible proteasome inhibitor, exerts anti-MM effect, mainly through activation of caspase pathway and inhibition of basal autophagy. It is of great importance that BSc2118 could overcome bortezomib resistance via inhibition of autophagy flux. A head to head of BSc2118 versus Bortezomib is performing in human plasmacytoma xenograft tumor model to evaluate drug safety, anti-tumor efficiency, in particular, reversal of bortezomib resistance. Our preclinical study supports clinical evaluation of BSc2118, particularly, overcomes bortezomib resistance, as a potential MM therapy.

Disclosures
No relevant conflicts of interest to declare.
Effectiveness of Lenalidomide and Dexamethasone for the Treatment of Extramedullary Plasmacytoma in Patients with Multiple Myeloma: Report of Two Cases
Authors: Masárová Katarína , Stefanikova Zdenka , Mistrik Martin , Batorova Angelika 
Published: October 2019

Abstract
Extramedullary plasmacytoma (EMP) can occur at the time of primary diagnosis of multiple myeloma (MM), during the disease, or as solitary plasmacytoma without bone marrow involvement. The presence of EMP at any time in the course of disease is associated with significantly shorter overall survival and progression-free survival. We report a series of two patients with EMP who were successfully treated with lenalidomide and dexamethasone.
The first patient was diagnosed in 2006 with EMP in the spine without presence of MM at the time of diagnosis. Reconstruction surgery followed by high dose dexamethasone treatment led to stabilization of patient’s condition. Patient received zoledronic acid for prevention of skeletal fractures. In 2011 the patient was diagnosed with MM and two new EMP lesions in the spine were discovered. The patient received bortezomib and achieved a partial remission. In 2012 a significant growth of EMP in retroperitoneum was observed. Tumor was surgically removed, plasmacytoma was confirmed histologically. Patient was treated with lenalidomide and dexamethasone with subsequent autologous peripheral blood stem cell transplantation (APBSCT) and achieved complete remission.
Patient Two had advanced stage of MM at the time of diagnosis, without presence of EMP. She was treated with induction therapy followed by APBSCT. EMP in the area of clivus, diagnosed three years after initial diagnosis of MM was successfully managed with conventional chemotherapy and radiotherapy. In 2010 another EMP in the left sphenoid sinus progressed during relapse of MM. Treatment with lenalidomide and dexametazone led to remission of MM and disparition of EMP. Both patients are in complete remission at the time of submitting this abstract.
Our clinical experience strongly suggests that lenalidomide with dexamethasone is an effective agent for treating EMP in patients with MM. The treatment was well tolerated with manageable side effects.

Disclosures
No relevant conflicts of interest to declare.