bluuebunny/crossref_metadata_2025_split huggingface dataset:
KIT D816V Mutation Positive Bone Marrow Mesenchymal Stem Cells in Indolent Systemic Mastocytosis Are Associated with Disease Progression
Authors: Garcia-Montero Andres C , Jara-Acevedo Maria , Alvarez-Twose Ivan , Teodosio Cristina , Sanchez-Muñoz Laura , Muñoz-Gonzalez Javier Ignacio , Mayado Andrea , Matito Almudena , Caldas Carolina , Morgado Jose M , Escribano Luis , Orfao Alberto 
Published: October 2019

Abstract
PURPOSE: Multilineageinvolvement of bone marrow (BM) hematopoiesis by the somatic KIT D816V mutation is present in a subset of adult indolent systemic mastocytosis (ISM) patients in association with a poorer prognosis. Here we investigated the potential involvement of BM mesenchymal stem cells (MSC) from ISM patients by the KIT D816V mutation and its potential impact on disease progression and outcome.
METHODS: The KIT D816V mutation was investigated in highly-purified BM MSC and other BM cell populations from 83 ISM patients followed for a median of 116 months. MC clonality was further evaluated in female patients by the pattern of inactivation of the X chromosome (XCIP).
RESULTS: KIT D816V-mutated MSC were detected in 22/83 (27%) ISM patients. All MSC-mutated patients had multilineage KIT mutation (100% vs. 30%, p=0.0001) and they more frequently showed involvement of lymphoid plus myeloid BM cells (59% vs 22%; P =.03) and a polyclonal XCIP of the KIT- mutated BM MC (64% vs 0%; P =0.01) vs other multilineage ISM cases. Moreover, presence of KIT D816V-mutated MSC was associated with more advanced disease features of ISM, a greater rate of disease progression (50% vs 17%; P =.04) and a shorter progression-free survival at 10, 20 and 30 years (P ≤.003).
CONCLUSION: Overall, these results support the notion that ISM patients with mutated MSC may have acquired the KIT mutation in a common pluripotent progenitor cell, prior to differentiation into MSC and hematopoietic precursor cells, before the X-chromosome inactivation process occurs. From a clinical point of view, acquisition of the KIT mutation in an earlier BM precursor cell confers a significantly greater risk for disease progression and a poorer outcome.

Disclosures
No relevant conflicts of interest to declare.
TOP2A Knockdown Resensitizes Carfilzomib-Resistant Hmcls to Carfilzomib
Authors: Reale Antonia , Khong Tiffany T. , Mithraprabhu Sridurga , Savvidou Ioanna , Ramachandran Malarmathy , Spencer Andrew 
Published: October 2019

Abstract
Introduction/background: Multiple myeloma (MM) remains incurable despite the introduction of novel therapeutic agents. Microarray-based technologies were adopted in our study to determine if a genetic signature associated with resistance to carfilzomib, a second-generation proteasome inhibitor already in use in clinical settings, could be identified.
Materials and Methods: 18 genetically heterogeneous human myeloma cell lines (HMCLs) were treated with carfilzomib and a cell viability profile was assessed categorizing the HMCLs as sensitive, intermediate or resistant to carfilzomib. Following categorization gene expression profiling was performed and validated with q-RT-PCR and knockdown assays.
Results: 29 genes were differentially regulated between the sensitive and resistant cell lines. Top genes based on intensity values and biological significance were: LOC731314, TSPAN13, APH1B, TSPYL5, COX7B2, PCSK1N, LRRC38, TCIRG1, TOP2A, ADM2, ITM2A, TSPAN13, STOM, UBE2C, SNHG8. Gene ontology (GO) enrichment analysis identified two pathways that were significantly different between the resistant and sensitive HMCLs; pathogenic escherichia coli infection (p=0.002) and lysosome (p=0.006). Eight GO terms were enriched: 4 related to biological processes and 4 related to cellular components.
TOP2A, an enzyme that controls and alters the topologic states of DNA during transcription and is involved in cell cycle and proliferation, was identified to be overexpressed in resistant HMCLs. It functions as the target for several anticancer agents and a variety of mutations in this gene have been associated with the development of drug resistance. TOP2A may be used as a predictive factor for patient selection for specific protocols or as independent prognostic marker in solid tumors. TOP2A was also overexpressed in the 'proliferation cluster' associated with greater proliferation rate and poor outcome in newly diagnosed MM patients. Suppression of TOP2A by siRNA in carfilzomib-resistant HMCLs significantly resensitised the cell lines to carfilzomib.
Conclusion: Our results suggest that TOP2A is overexpressed in carfilzomib-resistant HMCLs indicating a possible role as a predictive marker of response to carfilzomib in MM.

Disclosures
No relevant conflicts of interest to declare.
In Vivo Response to Cytarabine Chemotherapy Uncovers the Role of the Oxidative and Energetic Metabolism in the Chemoresistance of Human Primary AML Stem Cells
Authors: Aroua Nesrine , Sarry Jean-Emmanuel , De Toni Fabienne , Carson Robin , Vergez Francois , Saland Estelle , Sugita Mayumi , Carroll Martin , Recher Christian , Danet Gwenn 
Published: October 2019

Abstract
Resistant cells to conventional chemotherapy - which targets DNA synthesis - are thought to be rare and enriched in quiescent leukemic stem cells (LSCs) in acute myeloid leukemia (AML). In order to develop an improved understanding of chemotherapy resistance in AML, we analyzed the response to cytarabine (Ara-C) in our xenograft model with 20 primary AML patient specimens from two clinical sites. After demonstrating AML engraftment, mice are treated with Ara-C given IP daily for 5 days as a single agent at 60 mg/kg daily, which correlates with human dosing. In all mice treated with this regimen, there was a 4- to 46-fold cytoreductive effect at 1 week post-treatment. While in vivo Ara-C treatment induces changes in CD34pos CD38pos/neg phenotypes of 50% of AML-engrafted mice, this treatment does not induce any significant changes in cell cycle distribution and does not enrich in G0 quiescent cells in AML-engrafted mice. Surprisingly we observed that Ara-C induces a decrease or no change in the frequency of SL-ICs (control, 1:2,500-21,900 versus Ara-C, 1:4,500-34,200), suggesting that Ara-C would not spare LSCs in vivo and equally kills both cycling and quiescent cells. Furthermore, transcriptomic analysis of Ara-C residual leukemic cells (RLCs) from three different patient-derived xenografts confirmed that RLCs are not enriched in stem cell marker genes at 1 week post-treatment. However, our gene signature of the in vivo AML resistance is enriched in genes involved in inflammatory, immune and stress response. When tested in three independent AML cohorts (Verhaak et al. 2009; TGCA, 2011; Metzeler et al. 2011), this signature is associated with an unfavorable prognosis in patients treated with intensive chemotherapy. Finally we have shown that RLCs are PIMOpos cells with both high ROS content and active mitochondrial mass and membrane potential in AraC-treated xenografted mice compared to control mice. Accordingly, AML cells exhibiting high OXPHOS energetic phenotype are resistant to Ara-C chemotherapy in vivo while low OXPHOS AML cells are sensitive to AraC in NSG mice. Altogether these results suggest a novel model of cytarabine chemotherapy resistance in AML based on the oxidative and mitochondrial energy metabolism in vivo.

Disclosures
No relevant conflicts of interest to declare.
A Point-of-Care Platform for Hematopoietic Stem Cell Gene Therapy
Authors: Adair Jennifer E , Haworth Kevin G , Sauvageau Guy , Heimfeld Shelly , Hocum Jonah D , Trobridge Grant D. , Kiem Hans-Peter 
Published: October 2019

Abstract
Hematopoietic stem and progenitor cell (HSPC) gene therapy holds great promise but requires highly technical and dedicated facilities. Current state-of-the-art requires ex vivo HSPC gene transfer in dedicated Good Manufacturing Practices (GMP) facility infrastructure, limiting treatment to highly developed countries. A patient point-of-care strategy would therefore make HSPC gene therapy available to patients worldwide. We developed a short, semi-automated, mostly-closed platform for ex vivo isolation and lentivirus (LV) gene modification of CD34+ HSPCs from either bone marrow or mobilized peripheral blood sources using the CliniMACS Prodigy® (Figure 1). Experiments were performed with a biosafety cabinet and personal protective equipment to simulate anticipated conditions in clinical facilities of underdeveloped countries. A total of 7 custom programs were developed for bone marrow or mobilized peripheral blood CD34+ cell isolation and LV transduction. Addition of a pyrimidoindole derivative, UM 729, permitted efficient transduction of CD34+ HSPCs in <18 hours. Complete semi-automated production took <34 hours from collection to infusion of the gene modified cell product, requiring very little operator hands-on time. Autologous, LV gene-modified CD34+ HSPCs from two nonhuman primates produced using this platform engrafted and supported multilineage hematopoietic repopulation after myeloablative total body irradiation at 1020 cGy. Total cell doses achieved were 27 x 106 and 5.4 x 106 CD34+ cells/kg body weight, respectively. We observed stable, persistent and polyclonal gene marking in peripheral blood granulocytes up to 40% and lymphocytes up to 15% within 4 months after infusion. Neither animal displayed evidence for increased toxicity, including potential contamination from the cell products. We then validated processing of human bone marrow and mobilized apheresis products from healthy adult donors. We demonstrate efficient isolation of human CD34+ HSPCs and up to 60% transduction efficiency with a clinical-grade LV currently being tested in a phase I clinical trial for treatment of HIV-associated lymphoma. Products tested met current FDA approved specifications for infusion. Xenotransplantation of these products into immunodeficient mice resulted in polyclonal engraftment over 12 weeks. These data demonstrate preclinical safety and feasibility of a patient point-of-care strategy for ex vivo LV gene transfer into HSPCs. This platform represents a major advance in global portability of LV mediated HSPC gene therapy.
Figure 1. Figure 1.

Disclosures
No relevant conflicts of interest to declare.
An HSP90 Inhibitor, PU-H71, Antagonizes Stroma-Induced Pro-Survival Effects in CLL through Its Inhibition of Multi-Component B-Cell Receptor Signaling Pathway
Authors: Guo Ailin , Lu Pin , Zhen Chaojie , Chiosis Gabriela , Wang Yue Lynn 
Published: October 2019

Abstract
Background: Chronic lymphocytic leukemia (CLL) is characterized by the accumulation of B-cells in the hematopoietic system. The B-cell receptor (BCR) plays an essential role in the pathogenesis of CLL and many components of the BCR signaling pathway are known clients of HSP90. HSP90 is a highly conserved molecular chaperone that ensures the proper folding and stabilization of its client proteins. In this study, we investigated whether PU-H71 a novel purine-scaffold HSP90 Inhibitor, has anti-tumor activity in CLL by destabilizing BCR signaling pathway constituents.
Design: Fresh CLL cells were isolated and cultured ex vivo with or without stromal co-culture. Molecular and cellular events were studied in PU-H71-treated and control CLL cells.
Results: Immunoblotting revealed that a significantly higher amount of HSP90 is present in CLL cells than in peripheral blood mononuclear cells (PBMC), suggesting the chaperone is pathogenically relevant. We found that PU-H71 caused the death of CLL cells in a dose and time dependent manner while the viability of either PBMC or normal B lymphocytes were not affected. PU-H71 induced apoptosis resulting in CLL cell death as it caused mitochondrial cytochrome C release and a decrease in the abundance of several anti-apoptotic proteins. Interestingly, PU-H71 has the ability to counteract the pro-survival effects of the stroma and caused apoptosis in CLL cells co-cultured with stroma. To gain mechanistic insights into how PU-H71 acts, we examined the BCR signaling pathway. We found that the amounts of several key components of the pathway were reduced by PU-H71 treatment. This occurred even in the presence of stromal co-culture. The results suggest that PU-H71 antagonizes the function of HSP90 leading to the destabilization of the BCR signaling transducers. A chemical pull-down experiment revealed the co-existence of the BCR components and HSP90 in the same complex, suggesting these BCR constituents are indeed clients of HSP90 in CLL cells. Further, specific genetic knock-down of the signal transducers by siRNA confirmed their key roles in mediating the survival of CLL cells.
Conclusions: PU-H71 antagonizes stroma-induced pro-survival effects in CLL through its inhibition of the B-cell receptor signaling pathway. Our results suggest that PU-H71 may serve as a useful therapy against CLL and is worth further clinical development.

Disclosures
No relevant conflicts of interest to declare.
Risk of Early Mortality in Patients with Newly Diagnosed Multiple Myeloma
Authors: Liu Chia-Jen , Hsu Pei , Lin Ting-Wei , Gau Jyh-Pyng , Hsiao Liang-Tsai , Tzeng Cheng-Hwai , Chen Po-Min , Yu Yuan-Bin , Chiou Tzeon-Jye , Liu Yao-Chung , Liu Jing-Hwang 
Published: October 2019

Abstract

Background
The overall survival of patients with multiple myeloma has been improved greatly over the last two decades with the advances of treatment. Several studies reported that this improvement in survival has been ascribed to the broader use of novel drugs and autologous tandem transplantation. However, there were still a certain portion of myeloma patients died early after diagnosis. We therefore aim to investigate the risk factors of early mortality (death within 60 days after diagnosis) in patients with multiple myeloma.
Patients and Methods
We included in this study 451 consecutive patients with multiple myeloma, newly diagnosed at an Asian tertiary medical center between January 1, 2002 and April 30, 2015. A total 57 subjects who developed early mortality were identified. Risk factors for early mortality in myeloma patients were collected and analyzed.
Results
Compared with non-early mortality myeloma patients, early mortality patients had higher probability of being male, primary plasma cell leukemia, low platelet count, low serum albumin, high corrected serum calcium, high serum creatinine, high LDH, high serum β2-microglobulin, poor performance status, and high ISS stage. With multivariate analysis, we found that male (adjusted OR 2.93, 95% CI 1.17-7.31), serum albumin < 3.5g/dl (adjusted OR 2.76, 95% CI 1.17-6.52), corrected serum calcium ≥ 12mg/dl (adjusted OR 3.56, 95% CI 1.47-8.63) and LDH ≥ 250U/L (adjusted OR 3.30, 95% CI 1.62-6.74) were significant risk factors of early mortality. Pneumonia represented as the leading cause of early mortality in myeloma patients (n = 18, 31.5%), followed by renal failure (n = 7, 12.2%).
Conclusion
Early mortality rate is high (12.6%) in patients with multiple myeloma. Patients of male gender, low serum albumin, high corrected serum calcium and LDH are at risk of early mortality. More than one third myeloma patients (21 out of 57) who developed early mortality are died of infection. Identifying the risk group and providing prompt intervention, such as prophylaxis antibiotics, may reduce the incidence rate of early mortality and improve the life expectancy of myeloma patients.

Disclosures
No relevant conflicts of interest to declare.
A Bioinformatics Approach Revealed a Differential Expression of RKIP-Related Genes in Pre-Multiple Myeloma and Multiple Myeloma: Clinical Implications
Authors: Shvartsur Anna , Chatterjee Devasis , Chen Haiming , Berenson James , Garban Hermes , Bonavida Benjamin 
Published: October 2019

Abstract
Introduction: Multiple myeloma (MM) is a clonal plasma-cell neoplastic disorder arising from an indolent premalignant disease known as monoclonal gammopathy of undetermined significance (MGUS) that can evolve to asymptomatic pre-MM and later to symptomatic MM. MM is a biologically complex and a heterogeneous disease. The metastatic suppressor/anti-resistance factor, Raf-1 kinase inhibitor protein (RKIP), is poorly expressed in the majority of cancers. In contrast, inactive phosphorylated RKIP is overexpressed in MM and assisting in cell proliferation and resistance. Active RKIP inhibits the RAF/MEK/ERK1/2 and NF-κB pathways.
Rationale: A molecular identification of stage-specific pre-MM and MM biomarkers will support a more precise diagnosis/prognosis, an effective therapeutic regime, and will assist in the identification of novel therapeutic molecular targets.
Objective: To identify RKIP-related gene products in pre-MM and MM as novel biomarkers and therapeutic targets.
Hypothesis: We hypothesized that there would be differential molecular signatures between the pre-MM and MM disease-related to RKIP-regulated genes.
Methodology: We examined transcriptomic datasets on Oncomine platform (Life Technologies) for pre-MM and MM.
Results: Several gene products were found to be commonly overexpressed (e.g., RKIP, Bcl-2 and DR5); and underexpressed (e.g., Bcl-6 and TNFR-II) in both pre-MM and MM, suggesting a plausible distinctive general molecular signature of the disease. Importantly, a significant inverse correlation of differentially expressed pro-apoptotic genes was observed in pre-MM: overexpression of Fas and TNF -α and underexpression of YY1; versus expression of anti-apoptotic genes in MM: overexpression of YY1 and underexpression of Fas and TNF -α.
Conclusion and Future Directions: The findings suggested differential molecular signatures between pre-MM and MM. We are currently analyzing the relationship between RKIP and both overexpressed and underexpressed gene products to identify molecular pathways with cross-talks for the regulation of RKIP.
Clinical Implication: The proposed findings might lead to a more precise diagnosis/prognosis of MM and its disease stages, and can identify novel molecular therapeutic targets for pre-MM and MM.

Disclosures
No relevant conflicts of interest to declare.
Efficacy and Safety of Bortezomib Based Regimens Retreatment in Relapsed Multiple Myeloma Patients: Results from a Retrospective Study.
Authors: Wu Wenjun , Zheng Gaofeng , Han Xiaoyan , Zhao Yi , He Donghua , Yang Yang , He Jingsong , Zheng Weiyan , Sun Jie , Shi Jimin , Wei Guoqing , Huang He , Cai Zhen 
Published: October 2019

Abstract
Objective: To evaluate the efficacy and safety of bortezomib retreatment in relapsed multiple myeloma (MM) patients, who previously responded to bortezomib. Methods: This retrospective observational study included data from 45 patients and evaluated the efficacy and safety of bortezomib based retreatment in these patients, who had achieved at least a partial response (PR) on initial bortezomib therapy in our hospital from May 2006 to May 2013. Results: The overall response rate (ORR) was 71.2%, among them 9% patients achieved CR, 11.1% patients achieved very good partial response (VGPR), 51.1% patients achieved PR. All patients were divided into 3 groups according to the response of initial bortezomib therapy, including CR group, VGPR group and PR group. After bortezomib retreatment, the ORR of the 3 groups was 76.9%, 75% and 62.5%, respectively. According to the response of bortezomib retreatment, the patients were divided into 2 groups: group 1 who at least achieved PR, group 2 who showed no response. The median progression-free survival (PFS) after bortezomib retreatment for group 1 and 2 was 9( 95% confidence interval 7.947~10.051) and 10 (95% confidence interval 8.381∼11.619) months, respectively (P>0.05), while the median overall survival (OS) after bortezomib retreatment was 71 (95% confidence interval 66.694∼75.306)) and 37 (95% confidence interval 1-28) months, respectively (P<0.05). In patients with bortezomib retreatment had different degrees of adverse events (AE) , the most AE for grade 1~2. The most common grade ≥3 AE was thrombocytopenia, neutropenia and anemia. The incidence rate of grade ≥3 AE peripheral neuropathy bortezomib was 15%. Conclusion: Bortezomib based regimens retreatment was effective and tolerable in relapsed MM patients, who had achieved at least a partial response (PR) on initial therapy. The incidence rate of AE was not significantly increased.

Disclosures
No relevant conflicts of interest to declare.
CD38 Protects Tumor Cells from NK Lysis
Authors: Wang Anlai , Yang Guang , Song Zhili L , Adrian Francisco J 
Published: October 2019

Abstract
Expression of the type II glycoprotein CD38 in hematological malignancies is abnormally high compared to that in normal cells of hematopoietic origin. Across all malignancies, the highest CD38 expression levels are detected on the surface of multiple myeloma (MM) cells. The high CD38 level has prompted the development of anti-CD38 antibody-based therapies, like daratumumab and isatuximab (SAR650984), that are promising therapeutics for the treatment of patients with MM. Cellular functions associated with CD38 include co-receptor mediated signaling, cell adhesion and ecto-enzymatic conversion of NAD to modulate calcium signaling and adenosine synthesis; but other than allowing antibody-mediated engagement of innate immunity, the role of CD38 in MM is not fully understood.
In order to investigate a potential immune-protective role for CD38, we evaluated a panel of 15MM and 6 diffuse large B-cell lymphoma (DLBCL) cell lines with varying levels of CD38 for their susceptibility to NK-mediated lysis. Briefly, calcein AM pulsed target cells were incubated with NK-92.CD16 NK cells (E:T=3:1) for 1 hr and cell lysis measured as a function of the fluorescent calcein released into the supernatant. In an initial experiment, we observed that the percentage of cell lysis for all lines with >100,000 CD38s per cell was low, below 10%. The cell lines with <100,000 CD38s per cell were more susceptible to NKs, with a median percentage cell lysis of around 25%. The observed levels of cell lysis were not correlated with the levels of released IFNγ or TNFα.
The differential lytic activity against the target cells led to the hypothesis that CD38 might have a role in protecting the tumor cells from NK-mediated lysis. To investigate this, we first knocked down CD38 in SUDHL-8 cells using shRNA, decreasing the levels of CD38 from 246,000 to 51,000 per cell. This resulted in an increase in the percentage of lysed cells from 25% to more than 50%, likely reflecting a loss of protection against the NK cells. We next overexpressed CD38 in U266 cells, increasing the CD38 levels from 9,000 to 327,000 per cell. As a result, the percentage of lysis decreased from more than 60% to around 30%, indicating that U266 cells are now more resistant to the lysis by NK cells. The protection that CD38 seems to confer to CD38-overexpressing U266 cells is completely lost by addition of isatuximab which triggers CD16-mediated lysis (antibody dependent cellular toxicity, ADCC).
Whether the protective function of CD38 described here is a direct or an indirect protection mechanism is currently under investigation.

Disclosures
Off Label Use: isatuximab (SAR650984, anti-CD38 antibody). Song:Sanofi: Employment. Adrian:Sanofi: Employment.