bluuebunny/crossref_metadata_2025_split huggingface dataset:
Human Monocytes and Monocyte-Like Cells Are Highly Sensitive to Plitidepsin-Induced Cell Death In In Vitro Assays
Authors: Morande Pablo E , Zanetti Samanta , Borge Mercedes , Nannini Paula , Jancic Carolina , Bezares Raimundo , Galmarini Carlos M. , Gamberale Romina , Giordano Mirta 
Published: October 2019

Abstract
Abstract 4942
Plitidepsin (Aplidin®) is a marine-derived cyclic depsipeptide with strong cytotoxic effects against a variety of cancer cell types. It is currently in Phase II/III clinical trials for solid and hematologic malignancies. We evaluated the cytotoxic effects of Plitidepsin on lymphoid and monocyte cells from healthy donors and analysed the mechanisms involved. We found that monocyte cells showed a markedly higher sensitivity to Plitidepsin compared to lymphoid subsets as determined by incorporation of 7-AAD and flow cytometry analysis. Plitidepsin induced a dose- and time-dependent cell death on freshly isolated monocytes (7-AAD+ cells at 24 h: control: 3±1% versus Plitidepsin 10 nM: 47±5%, mean±SEM, n=6) and on macrophages (5±2 vs 49±3, n=5) and dendritic cells (4±1 vs 21±4, n=5) differentiated in vitro. By contrast, resting or mitogen-activated lymphocytes were not affected by Plitidepsin at 10 nM. The mechanisms of induced cell death in monocytes involved the early exposure of phosphatidylserine to the outer leaflet of plasma membrane, activation of caspase-3 and subsequent PARP fragmentation, indicative of death via apoptosis. Incubation of monocytes with Plitidepsin (10 or 100 nM) for 30 minuntes induced ROS production. On the other hand, the cytotoxic effect of Plitidepsin was significantly diminished when monocytes were pre-incubated with the antioxidant reagent Ebselen, which acts as an scavenger of peroxynitrite. We also determined Plitidepsin activity on malignant cells from chronic myelomonocytic leukemia (CMML) and chronic lymphocytic leukemia (CLL) patients. Monocyte-like cells obtained from 3 CMML samples resulted sensitive to Plitidepsin, though to a different extent (viable CD14+ cells at 24 h: control 100%, incubated with Plitidepsin 10 nM: sample 1: 79%, sample 2: 82%, sample 3: 29%). Finally we evaluated the effects of Plitidepsin on nurse-like cells (NLC) from CLL patients. These cells represent a subset differentiated from monocytes that favours leukemic cell progression through pro-survival signals. NLC were very sensitive to Plitidepsin and, more importantly, their death indirectly decreased neoplasic clone viability (7-AAD+ leukemic B cells co-cultured with control NLC: 36±10 versus 7-AAD+ leukemic B cells co-cultured with NLC exposed to Plitidepsin 100 nM for 24 h: 47±6, n=5, p<0.05). Myeloid cells are present in tumor microenvironment and actively affect the malignant cells, both directly and indirectly via the suppression of host immunity. Given their relevance in supporting tumor growth, the sensitivity of monocyte-like cells to Plitidepsin may contribute to its antitumoral effects.

Disclosures:
Galmarini: PharmaMar: Employment, Equity Ownership. Giordano: PharmaMar: Research Funding.
Function and Clonality of T-Cells In Myelodysplasia
Authors: Davison Glenda , Novitzky Nicolas , Abdulla Rygana 
Published: October 2019

Abstract
Abstract 4972

Objective:
Myelodysplastic (MDS) syndromes are clonal disorders arising from haematopoietic multi-potential stem cells. Research has implied that cells of the immune system play a role in the disease process. The aim of this study was to examine the clonality, response to mitogens, effects of lymphocytes on bone marrow clonogenic progenitors and proliferative capacity of selected marrow progenitors in a well defined cohort of MDS patients.

Methodology:
Seventeen patients with MDS were accrued. The clonality of the T-cells was studied using the TCR Vβ repertoire kit. In order to assess in vitro function, the lymphocytes were activated using phorbol 12-myristate13 acetate (PMA) and thereafter analysed for expression of the activation antigen CD69 using standard flow cytometry. The results were compared to age matched controls. The effect of the autologous blood mononuclear cell populations on clonogenic growth was studied by culturing Li- selected CD34+ cells with both PMA activated and non-activated autologous lymphocytes using a non-contact double layer culture technique.

Results:
T-cells did not demonstrate monoclonality, although skewing of the T-cell repertoire was observed. The median percent of CD3+ T-cells expressing CD69 after activation was 30% (9.5 – 95.4%). This was reduced in comparison to age matched controls (p=0.025). The effect of the lymphocytes on clonogenic cell growth was heterogeneous however median values demonstrated a cell dose response in the colony numbers directly related to cultured lymphocyte numbers (p=0.04 compared to no lymphocytes added). In addition, the median CFU-GM scores were higher when cultured with PMA activated lymphocytes (p=0.05). This pattern was not significantly different to normal controls.

Background:
Our investigations suggest that in this cohort of MDS patients the T-cells were not clonal but had reduced activation capacity when stimulated with the mitogen PMA. However, their ability to stimulate colony growth of autologous bone marrow CD34+ cells was preserved. Furthermore, selected CD34+ MDS cells proliferated well, with clone numbers not different from control. These observations imply that the previously reported abnormal T-cell responses and poor clonogenic growth may be the result of complex interactions between T-cells, the malignant clone and accessory cells in the bone marrow stroma. Further study examining each of these cell populations is required to better understand the mechanism of marrow failure and progression to leukaemia.

Disclosures:
No relevant conflicts of interest to declare.
EVALUATION of the Intra-LABORATORY Reproducibility of Percentage of Blast Counting IN MYELODYSPLASIA
Authors: Wuilleme Soraya , Eveillard Marion , Barille Sophie , Accard Francoise , Lodé Laurence , Avet-Loiseau Hervé , Garrand Richard 
Published: October 2019

Abstract
Abstract 4981

Background.
In the two International Prognostic Scoring Systems of Myelodysplastic Syndrom (MDS), the percentage of BLAST cells in the Bone Marrow is the most important parameter implicated in score: either directly in the IPSS (International Prognostic Scoring System) (1) or indirectly in the WPSS (Who classification-based prognostic scoring system) (2). In these two systems, as in the recommendations of the WHO 2001 (3), the morphological criteria defining blasts cells compared to promyelocyte cells are not specified.
In 2005, the IWGM-MDS (International Working Group on Morphology of myelodysplastic syndrome) (4-6) has established morphological criteria defining Blasts cells.
PURPOSE OF THE WORK.
Our objective in this study is to evaluate the reproducibility, among five observers of our laboratory, of the counting of the marrow blasts of 73 myelodysplasia.

Materials and Methods.
This study was conducted in several stages. 1st step was to test the implementation of the MDS-Foundation (http://www.mds- foundation.org/virtualmicroscopy) by 5 observers. The 2nd step was to perform correlation test of the selected cells from RAEB (selected and stained in our laboratory conditions), for the 5 observers. Finally Step 3 was to assess the correlation of blast percentage of 73 bone marrow smears from MDS patients, selected due to the presence of an excess of blast cells in the first reading on the bone aspiration.

Results.
Our results show that the correlation on counting blasts is generally satisfactory (percentage agreement: Test 1 = 86% and test 2 = 94%), while the concordance on the counting blasts of bone marow smears of 73 MDS patients and concordance on the WHO classification seems less satisfactory (agreement 3/5 observers. = 95% but agreed to 4–5/5 observers = 64%). These results can be explained partly by the inter-observer variability and by the variability of some parameters specific to smear marrow (poor quality smears, the staining and/or poverty of smears).

Conclusion.
The evaluation of the blasts in the MDS must be achieved: (i) at least 500 cells counted (ii) by at least two different observers (iii) by a third observer in discordant case. Despite these recommendations, the assessment of the blasts in myelodysplasia is difficult to achieve in many cases.

Disclosures:
No relevant conflicts of interest to declare.
Severe Lenalidomide-Induced Hypersensitivity Pneumonitis
Authors: Dy Irene A. , Gamboa Eric O. , Frankel Etta 
Published: October 2019

Abstract
Abstract 4982
Lenalidomide is an immunomodulatory agent that stimulates T-cell activation, enhances natural killer cells and inhibits cytokines, as tumor necrosis factor-alpha. It is FDA-approved for use in both transfusion-dependent myelodysplastic syndrome, especially 5q deletion syndrome and refractory and relapsed multiple myeloma. The known side effects are myelosuppression, diarrhea/constipation, fever, fatigue, rash and thromboembolism, while pulmonary complications are rarely reported. We present a patient with myelodysplastic syndrome on lenalidomide with acute pulmonary toxicity.
A 63-year-old Caucasian man was diagnosed with low-intermediate risk myelodysplastic syndrome with 5q deletion after presenting with anemia and thrombocytopenia. He was started on lenalidomide at 21mg daily dose but after 5 days began to develop dyspnea on exertion, cough and fever. He was admitted for treatment of community acquired pneumonia. Physical examination revealed a middle aged man in respiratory distress. Oxygen saturation on room air was 86%. Chest examination revealed dry crackles. Heart sounds were regular without murmurs or gallops. Laboratory tests revealed hemoglobin of 8.6g/dl, normal white count at 10,000 and platelets of 7,000/uL. BNP and echocardiogram were normal and chest radiography was initially unremarkable.
He was started on antibiotics but developed respiratory failure requiring endotracheal intubation and mechanical ventilation. CT scan of the chest showed diffuse interstitial reticulonodular opacities most prominent at the apices with scattered ground glass attenuation. Cultures from blood, sputum, urine and stool and brochoalveolar lavage did not reveal an infectious source. Broad spectrum antibiotics and antifungals were withheld. Clinical and radiologic improvement was noted after withdrawal of lenalidomide and initiation of steroids.
He was persistently transfusion-dependent hence lenalidomide was reintroduced. Within 6 days of lenalidomide, respiratory symptoms recurred. He later improved after withdrawal of the offending drug.
Our myelodysplastic patient developed acute respiratory failure after lenalidomide introduction which improved with steroids and drug withdrawal. He had symptom recurrence after reintroduction of lenalidomide. The exposure history, clinical and radiological examination, positive challenge test with exclusion of other etiologies support the diagnosis of lenalidomide-induced hypersensitivity pneumonitis. This is the third reported case of lenalidomide-induced hypersensitivity pneumonitis and it is likely that this entity is underdiagnosed due to concurrent steroid use especially in myeloma patients. Clinicians should be aware of this potential complication in patients who present with fever, hypoxemia and diffuse pulmonary infiltrates unresponsive to broad spectrum antibiotics as this disease is reversible if diagnosed early.

Disclosures:
No relevant conflicts of interest to declare.
Comparison of Survival and Clinical Features In Different Age Cohorts of Patients with Multiple Myeloma: A Single Center Experience
Authors: Jeong Jae Ho , Yoon Shinkyo , Kim Dalyong , Kim Jeong Eun , Yoon Dok Hyun , Kim Shin , Suh Cheolwon 
Published: October 2019

Abstract
Abstract 4997

Purpose
This analysis was aimed to compare clinical features in different age cohorts of multiple myeloma (MM) patients and to identify the impact of the clinical characteristics on survival in the patients in a single center.

Patients and Methods
We retrospectively analyzed data from 513 patients between Sep 1995 and Feb 2010 in the Asan Medical Center. Correlation of baseline characteristics with survival was made by the following age strata: <45, 45–54, 55–64, and ≥65 years of age.

Results
Median observed overall survival (OS) in the entire cohort was 3.20 years (95% confidence interval: 2.71–3.68). Observed survival decreased steadily with ages from 4.56 years in patients younger than age 45 years to 2.50 years in patients ≥65 years (P=0.003). The patients in the oldest group (age 65 years or older) were more likely to have International Staging System (ISS) stage II or III than others (P=0.007). There were no significant differences in hypercalcemia (≥10 mg/dL), elevated serum creatinine (≥2 mg/dL), anemia (Hb <10 g/dL) and osteolytic bone lesion, so called CRAB among the different age cohorts. Prognostic factors including elevated C-reactive protein (CRP) (≥ 0.8mg/dL), lactate dehydrogenase (LDH), bone marrow plasma cell infiltration also did not differ significantly among 10-year age cohorts. However, hypoalbuminemia (<3.5 g/dL) and high serum β2-microglobulin (≥3.5 mg/dL) level were more frequent in patients ≥65 years of age (P=0.001 and P=0.046, respectively). Furthermore, higher proportion of patients underwent ASCT in the younger 10-year age cohorts (71.4%, 53.7%, 43.4%, 2.5% in < 45, 45–54, 55–64, and ≥ 65 years of age, respectively, P<0.001).

Conclusion
No significant differences in the myeloma related organ or tissue impairment and prognostic markers including CRP, LDH and bone marrow infiltration of myeloma cells were noted among different age groups. However, survival significantly declined in the higher age group, which might be related to higher ISS stage associated with hypoalbuminemia, high serum β2-microglobulin level and lower proportion of patients receiving ASCT.

Disclosures:
No relevant conflicts of interest to declare.
Comorbid Diabetes Does Not Impact the Survival of Patients with Multiple Myeloma
Authors: Hsiao Liang-Tsai , Chiou Tzeon-Jye , Hong Ying-Chung , Liu Chun-Yu , Yu Yuan-Bin , Teng Hao-Wei , Yang Sheng-Hsiang , Yang Ching-Fen , Gau Jyh-Pyng , Liu Jin-Hwang , Hsu Hui-Chi , Chen Po-Min , Tzeng Cheng-Hwai 
Published: October 2019

Abstract
Abstract 5003

Background & aims.
For patients with multiple myeloma (MM), comorbid type II diabetes mellitus (DM) has leaded additional consideration and complications. Preceding DM in MM patients may have lead to end-organ damage involving cardiovascular, renal and nervous systems, and the use of glucocorticoids - the mainstay of anti-myeloma agents - further impairs glycemic control. However, it is not well studied at present whether comorbid DM itself will influence clinical features and prognosis of MM patients. Methods. Patients who were newly diagnosed in Taipei Veterans General Hospital between 1996 and 2007 were enrolled. Data of clinical features, comorbidities, and laboratory tests at diagnosis, treatment modalities, and survival at last follow up were collected. MM patients with or without DM was compared in terms of clinical/laboratory tests and outcome. Results. There were 389 MM patients (M/F=288/101, 74% vs. 26%) with median age of 71 years, with the features summarized in the Table. The immunophenotypes consisted of IgG (194, 49.9%), IgA (113, 29%), and light chain diseases (LCD) (67, 17.2%). Clinical stages included Durie-Salmon (DS) stage I/II/III = 7.2%/20.1%/72.7% and International Staging System (ISS) stage I/II/III = 14.5%/29.3%/56.2%. Serum Cr > 2.0 mg/dL at diagnosis was noted in 34%. Sixty patients (15.4% of 389) had DM preceding the diagnosis of MM. There was no statistical difference of median age, gender distribution, DS stages, and renal impairment when comparing MM patients with or without comorbid DM.MM patients with comorbid DM were associated with a lower prevalence of IgA-type diseases (with vs. without DM= 17% vs. 32%, P=0.094), an elevated serum beta-2 microglobulin >5 mg/L (with vs. without DM = 75% vs. 52%, P=.006), and ISS-stage III disease (with vs. without DM = 75% vs. 53%, P=.005). In terms of anti-myeloma therapy, the proportion of MM patients with or without DM who once received thalidomide, bortezomib and high-dose chemotherapy plus stem cell transplantation (HDT and SCT), and bisphosphonates were similar. The median overall survival (OS) of all patients was 20.5 months. The OS was well correlated with ISS stages (I vs. II vs. III = 51.2 vs. 27.2 vs. 13.4 months, P<0.001) but was not with comorbid DM. Conclusions. Comorbid DM was present in up to 15% of MM patients, and was more frequently associated with elevated serum beta-2 microglobulin (>5 mg/L) and ISS-stage III diseases; however, comorbid DM did not significantly influence the OS. Therefore, comorbid DM itself should not be considered as a contraindication of aggressive anti-myeloma treatments at present.

Disclosures:
No relevant conflicts of interest to declare.
Effect of Imids on Gene Expression Profiles of Fresh Human Myeloma Cells
Authors: Tagoug Ines , Plesa Adriana , Vendrell Julie , Dumontet Charles 
Published: October 2019

Abstract
Abstract 5013
Immunomodulatory drugs represent a major therapeutic advance in the treatment of patients with multiple myeloma. While these agents appear to exert various effects on the microenvironment, including effect on immune cells and angiogenesis, a direct effect on the tumor cells themselves is also likely. To describe and compare the effect of the three clinically available agents (thalidomide, lenalidomide, pomalidomide) we analyzed the gene expression profiles of fresh human myeloma cells exposed to thalidomide, lenalidomide or pomalidomide, using high density DNA arrays. Fresh human myeloma samples were obtained from bone marrow aspirates of patients with myeloma, and myeloma cells were immunopurified using anti CD138 magnetic beads. Purified myeloma cells (1.106 cells/ml) were incubated for 24 hours in RPMI 1640 medium supplemented with 10% fetal calf serum under each of the four following conditions: 1) DMSO; 2) thalidomide 40 microM; 3) lenalidomide 1 microM; 4) pomalidomide 100 nM. These levels are achievable in the plasma of MM pts. Pangenomic array experiments were performed usingWhole Human Genome 4 × 44K Agilent one-color microarrays. Data were normalized using the quantile normalization method. Samples were analysed for differentially expressed genes, taking into account both the level of significance and the fold-change. Ten evaluable samples were processed. Exposure to thalidomide, lenalidomide and pomalidomide induced differential expression of 36, 50 and 75 genes, respectively, in comparison to DMSO-exposed controls, the total list including 101 genes. Twelve of these were found to be differentially expressed after exposure to all of the three agents, including trophoblast glycoprotein, WAS protein family member 1, dickkopf homolog 1, pentraxin-related gene, CD28, interleukin 12B, tissue factor pathway inhibitor 2, phospholipase A2, dehydrogenase/reductase (SDR family) member 9, hypothetical LOC145788 and betacellulin. These commonly altered genes could be mechanistically involved in themultiple activities of these agents in multiple myeloma or may represent epiphenoma mechanistically unrelated to drug-induced cell death. Genes differentially expressed between the treatment with each of these agents could be indicative of the different and non-overlapping actions these agents have in multiple myeloma. An example of this is the recent demonstration that pomalidomide is clinically active in lenalidomide refractory patients. These results suggest that exposure to IMIDs induce various intracellular signalization pathways in myeloma cells which might be involved in the cytotoxic activity of these compounds.

Disclosures:
Dumontet: Celgene: Research Funding.
Frontline Management of Myeloma with Bortezemib and Dexamethasone - Focus on Renal Failure - A Study From India
Authors: Aggarwal Shyam , Jauhri Mayank , Negi Anita , Kohli Sangeeta , Minhas Sachin 
Published: October 2019

Abstract
Abstract 5045

Introduction:
First line treatment of myeloma has witnessed significant changes and improvement in the last one decade. Bortezemib – proteasome inhibitor and lenalidomide – IMID, in combination with other drugs offers the best chances to achieve complete remission and or very good partial remission. These two agents also seem to have overcome the barrier of poor prognostic factors including 4:14 translocation, 13p deletion, renal failure.

Material and methods:
We analysed 21 patients of multiple myeloma treated at our hospital in 2008–2009 with bortezemib and dexamethasone. Median age at presentation was 62 years. There were sixteen males and five females. IgG Kappa monoclonal spike was seen in fourteen cases. Light chains alone were seen in urine in 4 cases. Sixteen patients had renal failure. Bone involvement was seen in fifteen patients, anemia in sixteen cases. B2 microglobulin levels were >2.5mg/L in fifteen cases. As per ISS staging, thirteen patients were stage 3, six were stage 2 and two were stage 1. All patients were treated with bortezemib 1.3 mg per sq.mt and dexamethasone 40 mg day 1, 4, 8, 11. Cycles were repeated every three weeks. Four such cycles were given. Hemodialysis was done as indicated.

Results:
Complete remission (negative IFE) was seen in five, VGPR in five, PR in five, stable disease in four and progressive disease in two cases. Improvement in renal failure and independence from dialysis was seen in twelve of sixteen cases. Out of 21 cases, eight developed peripheral neuropathy – four mild to moderate and four severe painful neuropathy. In the latter four bortezemib was discontinued. Five patients developed thrombocytopenia requiring dose reduction for bortezemib. Four patients developed herpes zoster.

Conclusions:
Bortezemib and dexamethsone combination is effective and safe in frontline management of myeloma patients especially with renal failure. Acyclovir prophylaxis should be considered for all patients and peripheral neuropathy should be closely monitored.

Disclosures:
No relevant conflicts of interest to declare.