bluuebunny/crossref_metadata_2025 huggingface dataset:
MicroRNA Changes Occur in Multiple Myeloma Cells in the Context of Bone Marrow Milieu.
Authors: Roccaro Roccaro M. , Sacco Antonio , Azab Abdel Kareem , Azab Feda , Ngo Hai , Melhem Molly R , Quong Phong , Runnels Judith M. , Anderson Kenneth C. , Ghobrial Irene
Published: October 2019

Abstract 1785

Poster Board I-811

Background

We and Others have previously demonstrated that primary multiple myeloma (MM) cells are characterized by a specific microRNA (miRNA) signature compared to the related normal plasmacell counterpart; and that miRNAs play a crucial role in regulating MM pathogenesis. Nevertheless, miRNA changes that occur in MM cells in the context of the bone marrow microenvironment have not been previously examined. Therefore, characterization of miRNA profiling of MM cells in conjunction with bone marrow stromal cells (BMSCs) is important to better understand the underlying molecular changes that lead to initiation and progression of this disease.

Methods

We performed miRNA-expression-profiling of MM cell lines (MM.1S; RPMI8226) that were co-cultured with primary BMSCs obtained from 5 MM patients, using liquid phase Luminex microbead miRNA profiling (Luminex, Austin, TX). The expression patterns of unfiltered data were performed using unsupervised hierarchical clustering of samples, based on centroid linkage and 1-correlation distance metric, using dChip (www.dchip.org). To further define those miRNAs differentially expressed between groups (patients vs normal), the data were filtered on significance of differences using ANOVA test, (P < 0.05). Microbead-miRNA profiling data were validated data by stem-loop qRT-PCR. To identify specific predicted miRNA-targeted mRNAs, TargetScan, PicTar, and miRanda algorithms were used.

Results

miRNA profiling of MM cells cultured with primary BMSCs (MM+BMSC system) differs from MM cells which were not grown in contact with primary BMSCs (MM cells alone). Specifically, we observed increased expression of miRNA-450, -432*, -299-5p, -409-3p, -29b, -542-5p, -184, -517*, -218, 128b, -142-5p and -211 (P<0.05) in MM cells obtained from the MM+BMSC system, compared to MM cells alone. Stem-loop qRT-PCR was performed on matched samples and showed expression patterns similar to those observed in miRNA analysis. Using algorithms commonly used to predict human miRNA gene targets (miRanda; TargetScan; PicTar), predicted targets of the increased miRNAs included negative regulators of NFkB, PI3K/Akt/mTOR, and MAPK/ERK signaling pathways, such as PTEN, KSR2, TWEAK, and DUSP; as well as tumor suppressors (MCC, TSSC1, TUSC1, FBW7, RHOBTB), pro-apoptotic factors and cyclin-dependent kinases inhibitors.

These data demonstrate that bone marrow stromal cells exert a modulatory effect on miRNA profiling in MM cells, which results in promoting MM cell growth and reducing MM cell survival.

Disclosures

Ghobrial: Millennium : Honoraria, Research Funding, Speakers Bureau; Celgene: Consultancy, Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau.
The Combination of Arsenic Trioxide and Interferon-Alpha Eradicates Leukemia Initiating Cells in TAX-Driven Murine Adult T Cell Leukemia/Lymphoma.
Authors: Bazarbachi Ali , Hajj Hiba El , El-Sabban Marwan , Hasegawa Hideki , Zaatari Ghazi , Saab Shahrazad , Janin Anne , Mahfouz Rami , Nasr Rihab , Kfoury Youmna , Nicot Christophe , Hermine Olivier , Hall William , de The Hugues
Published: October 2019

Abstract 2714

Poster Board II-690

Adult T cell leukemia (ATL) is one of the rare human cancers initiated by a transforming retrovirus, HTLV-I. After many years of controversy, it is now accepted that the viral transactivator protein Tax plays a critical role in initiating the leukemic process, because Tax transgenics develop a disease with striking ATL features. Long-term prognosis of ATL patients remains extremely poor, but we recently reported that the combination of As2O3, interferon-a (IFN), and zidovudine yielded unprecedented response rates. Indeed, in 10 chronic ATL patients, a 100% response rate was observed, including 7 complete remissions (CR), 2 CR but with more than 5% circulating atypical lymphocytes, and one partial response. We demonstrate that the As2O3/IFN combination, previously shown to degrade Tax, cures Tax-driven murine ATLs. Unexpectedly, this combination immediately abrogates leukemia transplantation into secondary recipients, while the primary tumor continues to grow and only exhausts much later. Leukemia initiating cell (LIC) clearance is reversed by proteasome inhibition, demonstrating that LICs are addicted to continuous Tax expression. Most anticancer treatments fail to target LIC. These results establish that As2O3/IFN/zidovudine acts through Tax targeting and predict a favorable long-term outcome for responsive patients. Thus, oncogene degradation can selectively target LICs, explaining this very recent success of a similar regimen in patients.

Disclosures:

No relevant conflicts of interest to declare.
Characterization of Bortezomib (Velcade ™) Resistance in a New Set of Human Cell Lines Obtained by Stepwise Selection.
Authors: Cai Jie , Lian Xian Jin , von Roretz Christopher , Shustik Chaim , Gallouzi Imed , Gros Philippe
Published: October 2019

Abstract 2741

Poster Board II-717

Bortezomib (Velcade ™, PS-341, BTZ) is a boronate dipeptide that reversibly inhibits the 26S proteasome, which is essential for the breakdown of ubiquitinated proteins and the regulation of normal cellular homeostasis. The activity of BTZ in treatment of newly diagnosed and refractory/relapsed multiple myeloma may be limited by the development of chemoresistance, the mechanisms of which are poorly understood. To investigate the molecular basis of Bortezomib resistance, BTZ-resistant (BTZr) cell lines were generated by stepwise selection procedures from HeLa, CCRF-CEM, and 4 multiple myeloma cells lines (8226S, U266, H929, and MM.1S), respectively. These BTZ-selected cell lines displayed varying degrees of elevated resistance (2 to 50 fold) to clinically relevant concentrations of BTZ. In addition, while most of the BTZr cells showed cross resistance to several other proteasome inhibitors (PIs), including MG-132 and Epoxomicin, they remained as sensitive to other chemotherapeutic drugs, such as anthracyclines, vinkalkaloids and etoposide, as their parental cells. The proteasome activity profiles are distinct among the cell lines. All parental cell lines displayed varying levels of chymotrypsin-like (CT-L) activity, which is the primary target of BTZ. Most BTZr lines showed markedly decreased CT-L activity, with a few exceptions. Moreover, the observed CT-L activity in all cell lines can be inhibited directly by BTZ and other PIs. In contrast, very low levels of caspase-like or post-glutamyl peptide hydrolyzing (PGPH) proteasome activity were detected in all cell lines. BTZ resistance in HeLa/BTZ cells was closely associated with increased resistance to PI-induced apoptosis, as shown by reduced number of Annexin V-stained cells and by delayed activation/cleavage of apoptosis proteins, such as Caspase-3 and Poly(ADP-ribose) Polymerase (PARP). Furthermore, the resistance to BTZ affected the mechanisms of cell stress responses. As for the parental HeLa cells, HeLa/BTZr cells retained the ability to form, in response to PI treatment, pro-survival foci in the cytoplasm known as stress granules (SGs). However, the drug concentrations required to induce SG formation in HeLa/BTZr cells are much higher (∼4 fold) than those for the parental HeLa, suggesting the development of stress-coping mechanisms in these BTZr cells. Gene expression profiling studies are in progress to identify transcriptomes individually or generally associated with BTZ resistance in these cell lines. Further characterization of these phenotypically similar, yet mechanistically distinct BTZr cell lines may elucidate diverse mechanisms of drug resistance to Bortezomib and other proteasome inhibitors.

Disclosures:

No relevant conflicts of interest to declare.
Targeting Axl Kinase in Hematological Malignancies.
Authors: Gourley Eric S. , Liu Xiao-hui , Hewitt William , Walker Rusty , Warner Steven L. , Vankayalapati Hariprasad , Bearss David J.
Published: October 2019

Abstract 2758

Poster Board II-734

Axl kinase, a member of the TAM family and also known as UFO, ARK, and Tyro7, is a receptor tyrosine kinase implicated in tumorigenesis. Overexpression of Axl is associated with increased cellular transformation, cell survival, proliferation, migration, angiogenesis, and adhesion. The oncogenic potential of Axl was first discovered in chronic myelogenous leukemia (CML) and has been shown to play a role in the development of acute myelogenous leukemia (AML) and myelodysplasia. Binding of Growth Arrest Specific Gene-6 (GAS6), a vitamin K-dependent protein and a known ligand for Axl, leads to subsequent phosphorylation events of downstream effecter molecules such as mitogen-activated protein (MAP) kinase and phosphatidyl-inositol (PI)-3 kinase/AKT pathways which are critical in oncogenic transformation. Furthermore, target validation studies of in vivo cancer models show that inhibition of Axl expression by RNA interference blocked tumor growth in those models. Taken together, this information makes Axl kinase an exciting target for small molecule drug discovery. Effectively utilizing SuperGen's proprietary CLIMB“technology, a series of small molecule inhibitors were rapidly discovered and developed for potency and selectivity against Axl. Lead candidate compounds demonstrate low nano-molar IC50 values in an Axl kinase biochemical assay, and focused selectivity for Axl when screened in kinase panels. Likewise, these compounds showed low and sub micro-molar activity from an anti-proliferative leukemia/lymphoma cell-based panel. Similarly, treatment of leukemia cells with these compounds in combination with known chemotherapeutic agents produced results that implicate combinatorial therapies as potentially beneficial treatments for patients suffering from hematological malignancies. In mechanistic, target validation assays, EC50 data from immunoprecipitated western blots of transiently transfected liquid tumor cells showed nano-molar inhibition of phospho-Axl when detecting with an anti-phospho-tyrosine antibody. A western blot panel of leukemia and lymphoma cell lines showed overexpression of Axl in several of these cell lines, further validating Axl as a potential therapeutic target in hematological cancers. Functional cell-based assays using SuperGen's lead compounds also potently inhibited phospho-AKT (S473). We propose that SuperGen's small molecule inhibitors against Axl kinase represent a new class of compounds with potent and selective activity in hematological malignancies.

Disclosures:

No relevant conflicts of interest to declare.
Inhibition of Cytokine Signaling through Activation of Jak2 Ubiquitination by WP1130.
Authors: Kapuria Vaibhav , Peterson Luke F , Bornmann William , Talpaz Moshe , Donato Nicholas J
Published: October 2019

Abstract 2907

Poster Board II-883

Constitutive activation of Jak2 through mutations or cytokine stimulation leads to abnormal expansion of the myeloid lineage and plays a key role in multiple myeloproliferative disorders. Kinase-specific inhibitors that prevent substrate phosphorylation can provide effective therapy for these disorders. However, this approach is sometimes inadequate as additional mutations and target-specific modifications can alter the kinase in a sufficient fashion to reduce kinase inhibitor affinity. Therefore additional approaches and molecules may be needed to reduce kinase activity through alternate mechanisms. In this report we describe a novel small molecule-mediated approach to block the Jak2/Stat signaling pathway. Using a previously identified small molecule with Jak2 inhibitory activity, we performed structure-activity relationship studies of AG490 derivatives in a cell-based analysis for inhibition of cytokine-stimulated Stat activation. This effort lead to the synthesis of WP1130 which suppressed Jak2 driven Stat activation with nM to μM potency. However, WP1130 did not act as a direct Jak2 kinase inhibitor. Rather, WP1130 treatment induced a rapid, time and dose-dependent trafficking of Jak2 from the detergent soluble to insoluble cellular fraction. We observed a robust induction of protein ubiquitination in WP1130 treated cells which was not directly or indirectly due to proteosome inhibition. Further, WP1130-mediated ubiquitination was unique with respect to other compounds as it did not reduce association of Jak2 with chaperones, did not induce ER stress or increase reactive oxygen species to activate protein ubiquitination. Jak2 was poly-ubiquitinated, primarily with lysine63-linked polymers, after WP1130 treatment and ubiquitination correlated with Jak2 mobilization without its proteolysis. Sub-cellular fractionation and analysis using confocal microscopy suggested rapid trafficking of Jak2 into cellular structures called aggresomes, in complex with Hsp90, Hsp70 and HDAC6. Lysates from WP1130 treated cells had diminished capacity to hydrolyze ubiquitinated substrates, suggesting the involved of reduced deubiquitinase activity as a mediator of Jak2 ubiquitination in WP1130-treated cells. Together, these results suggest that WP1130 enhances a resident ubiquitination cascade that results in Jak2 ubiquitination and its displacement from its peri-membranous, signaling competent sites. The results also suggest that Jak2 is able to undergo K63-linked poly-ubiquitination and may be exploitable as an alternate approach to interrupt Jak/Stat signaling.

Disclosures:

No relevant conflicts of interest to declare.
Procoagulants and Subclinical Atherosclerosis in Young Adults: The CARDIA Study.
Authors: Green David , Foiles Nancy , Chan Cheeling , Kang Joseph , Schreiner Pamela , Liu Kiang
Published: October 2019

Abstract 2973

Poster Board II-948

Background:

Elevated levels of procoagulants (FVII, FVIII, vWF) have been associated with cardiovascular disease, but whether they play a role in atherogenesis is unclear. Study of these factors in young adulthood, prior to clinically manifested disease, and years later when subclinical atherosclerosis has developed, might clarify these associations. The Coronary Artery Risk Development in Young Adults (CARDIA) Study provides an opportunity to examine this issue.

Methods:

Assays of FVII, FVIII, and vWF were performed in 1255 participants ages 23-37 (Year 7,Y7) and repeated at ages 38-50 (Year 20,Y20). Coronary artery calcification (CAC) prevalence and mean intimal-medial thickness (IMT) in the common carotid (CC) and internal carotid (IC) were measured at Y20. Prospective (Y7 clotting factors, Y20 CAC & IMT) and cross-sectional (Y20 clotting factors, Y20 CAC & IMT) analyses were performed. We also grouped participants according to whether they had one or more procoagulants in the highest tertile at Y7, Y20, or both, and evaluated their associations with subclinical disease.

Results:

Y7 levels of procoagulants (%), mean(SD) were: FVII 76(18), FVIII 102(38), and vWF 108(47). At Y20, all had increased by 40% to 55%, and CAC>0 was present in 20% of participants. After adjustment for age, the prospective analyses showed a trend of progressively greater CC thickness from the lowest to the highest tertile of FVII activity in the total group (P=0.007) and in whites (P=0.002) and men (P=0.015). Associations of FVII with IC thickness (0.82 mm to 0.84 mm) and the prevalence of CAC (18.6% to 23%) were weaker (P-trend, 0.1-0.3). Higher FVIII levels were associated with greater IC thickness in the total group, in whites, and in women (highest vs lowest tertile, P<0.05 for each comparison). No associations were seen with vWF. Cross-sectional analyses confirmed the association between FVII and carotid thickening, most strongly with the CC (P=0.002). Most associations were attenuated by multivariable adjustment (BMI, smoking, education, blood pressure, cholesterol, & CRP). Participants with FVII values in the highest tertile at Y7, Y20, or both had a higher prevalence of CAC and greater carotid thickening than those with values in the lowest tertile (P<0.05). Such associations were not observed with FVIII or vWF.

Conclusion:

FVII is a marker but not an independent risk factor for future atherosclerosis in young adults; FVIII is associated with IC thickening, and no associations were observed for vWF.

Disclosures:

No relevant conflicts of interest to declare.
Coating of Tumor Cells by Platelets Confers Expression of Immunoregulatory Molecules Which Impair NK Cell Anti-Tumor Reactivity.
Authors: Placke Theresa , Kopp Hans-Georg , Kanz Lothar , Salih Helmut R
Published: October 2019

Abstract 2993

Poster Board II-970

NK cells play an important role in cancer immunosurveillance and may prevent tumor progression and metastasis due to their ability to mediate direct cellular cytotoxicity and by releasing immunoregulatory cytokines which shape adaptive immune responses. Their reactivity is governed by various activating and inhibitory molecules expressed on target cells and reciprocal interactions with other hematopoietic cells like dendritic cells. Platelets contribute to tumor immune escape, metastasis, and angiogenesis (e.g. Jin et al Nature Med. 2006). In mice, thrombocytopenia inhibits metastasis, and this is reversed by NK cell depletion suggesting that platelets are an important additional player in NK cell-tumor interaction. However, the knowledge regarding the molecular mechanisms by which platelets influence NK cells is fragmentary at best. We found recently that platelet release soluble factors, most notably TGF-β, upon interaction with tumor cells which mediates NK cell silencing through downregulation of the activating immunoreceptor NKG2D (Kopp et al., Cancer Res 2009, in press). However, immunoregulatory molecules residing in the platelet membrane may also modulate NK cell anti-tumor responses. We report here that presence of platelets causes coating of tumor cells, and this markedly reduces NK cell lysis of tumor cells. This is mediated by conferment of “pseudoexpression” of platelet-expressed immunoregulatory molecules to tumor cells which are absent on the tumor cells alone. Among those immunregulatory molecules we identified various ligands for NK cell receptors like MHC class I, GITR ligand or CD62P. To establish the functional significance of tumor cell pseudoexpression of platelet molecules we employed functional analyses of tumor cells and NK cells with or without coating by autologous platelets. The impaired anti-tumor reactivity of NK cells against coated tumor cells was restored by blocking MHC class I on the coating platelets, while isotype control had no effect. Moreover, coating of tumor cells by platelets was validated by ex vivo analyses of primary leukemic cells from patients which also revealed substantial coating by platelets and confered expression of NK cell-modulating molecules. Our data indicate that platelets enable a molecular mimicry of tumor cells, which enables tumor cells to escape NK cell-mediated tumor immunosurveillance.

Disclosures:

No relevant conflicts of interest to declare.
Astrocyte-Plexus Is Formed in the Neovascularization Induced by CD11b+ Cells From An Ischemic Hind-Limb Mouse Model.
Authors: Kim Jeong A. , Hong Oak Kee , Hong Jong Wook , Broxmeyer Hal E.
Published: October 2019

Abstract 3043

Poster Board II-1019

Angiogenic sprouting needs to navigate through tissues to establish a vascular branching pattern. Such angiogenic guidance is mainly inferred by nonrandom angiogenic sprouting in the developing central nervous system (CNS). In mammalian retina, a vascular plexus initially forms, superimposed on a preexisting astrocyte-plexus, and retinal astrocytes begin to express vascular endothelial growth factor (VEGF-A), which stimulates endothelial cells to sprout radially from the optic nerve head into the retinal periphery. However, there has been little investigation of the role of astrocyte-plexus in new vessel formation of other ischemic sites besides retina. We recently found that infiltrated CD11b+ cells in ischemic muscles differentiate into endothelial-like cells in vitro, and that direct-injection of muscle-derived CD11b+ cells enhances recovery of blood perfusion in ischemic hind-limbs of C57BL/6 mice. To study if the astrocytes are related to the neovascularization process in ischemic limbs, infiltrated CD11b+ myeloid-cells in ischemic muscles were isolated 4 days after femoral artery dissection, and then seeded at 0.5×105/cm2 onto Matrigel and cultured in EGM-2 medium. Under these conditions, we tested the following hypotheses: (1) cultured cells initially form an astrocyte-network in vitro, and (2) astrocytes are involved in the formation of vascular structure. Seven days after the cells were embedded in Matrigel, spindle shaped cells grew, spread-out radially from cell clusters to the periphery, and established a mesh-like network expressing glial fibrillary acidic protein (GFAP), a marker for astrocyte. These spindle shaped cells were also positive for pericyte markers: desmin, NG-2, and platelet-derived growth factor receptor (PDGFR)-β. Interestingly, proliferating endothelial cells were closely associated with astrocytes by extension of endothelial cell filopodia on astrocytes. This observation is consistent that astrocyte scaffold guides extension of endothelial cell filopodia. Two weeks after cells were embedded in Matrigel, CD11b+ cells expanded and were layered in the same way to form vessel wall-like structures consisting of hundreds of cells. Spindle shaped GFAP-positive cells gradually expressed smooth muscle-actin at the bottom of the culture plate, migrated to the vessel wall-like structures, and covered the surface of the walls like pericytes. Taken together with the novel finding of astrocyte-like cell differentiation from CD11b+ cells from Matrigel culture, mobilization of CD11b+ myeloid-cells to regions of muscle ischemia would appear to play an important role in neovascularization after ischemic injury.

Disclosures

No relevant conflicts of interest to declare.
Vascular Endothelial-Cadherin Targeted Alpha-Particle Mediated Vascular Killing and Remodeling Augments Subsequent Chemotherapeutic Efficacy.
Authors: Escorcia Freddy E , Henke Erik , McDevitt Michael R. , Villa Carlos , Smith-Jones Peter , Blasberg Ronald , Benezra Robert , Scheinberg David A.
Published: October 2019

Abstract 3054

Poster Board II-1030

Cancer therapies targeting tumor endothelium have shown promising results when combined with chemotherapeutics. Vascular endothelial (VE) cadherin is an endothelial cell specific molecule that is expressed constitutively throughout the vasculature and participates in the formation of adherens junctions between adjacent endothelial cells. The monoclonal antibody E4G10 specifically binds to a cryptic epitope exposed only on the monomeric unengaged form of VE-Cadherin found in neovasculature. We chemically conjugated E4G10 with the cytotoxic, short-range αa-particle emitter, 225Ac, and show that the agent dampens tumor growth as a single agent and this effect is enhanced when given in combination with chemotherapy in a xenograft mouse model. The therapeutic efficacy of the combination therapy is sequence dependent and most pronounced when 225Ac-E4G10 is administered prior to the chemotherapy. Immunohistochemical and immunofluorescence studies for endothelial cells (Meca32), vascular basement membrane (Collagen IV), and pericytes (alpha-smooth muscle actin (alpha-SMA), nerve/glial antigen 2 (NG2) ) showed that the vasculature of 225Ac-E4G10 treated tumors was depleted and the remaining vessels appeared normalized as evidenced by morphological changes, increased pericyte density and coverage. Fluorescence studies with Hoechst 33342 dye i.v. injected showed more homogenous distribution of the dye within 225Ac-E4G10 treated tumors, suggesting improved perfusion, and biodistribution studies with i.v. injected 111In-Diethylene triamine pentaacetic acid (DTPA), a freely diffusible small molecule, showed significantly (p=0.03) increased passive tumor accumulation of 111In-DTPA in 225Ac-E4G10-treated tumors when compared to controls. Quantitative autoradiographic studies on tumor sections also showed both stronger and more homogenous signal distribution in 225Ac-E4G10-treated tumors. These results show that 225Ac-E4G10 treatment leads to ablation and remodeling of the tumor vasculature allowing improved delivery of subsequent small molecules, such as chemotherapeutics, and thereby improves therapeutic outcome.

Disclosures

McDevitt: Actinium Pharma Inc.: Patents & Royalties. Scheinberg:Actinium Pharma Inc.: Patents & Royalties.
The Frequencies of ETV6-RUNX1 Fusion and Hyperdiploidy (>50 chromosomes) in Children with Acute Lymphoblastic Leukemia Are Lower in Far East Than the West..
Authors: Liang Der-Cherng , Shih Lee-Yung , Yang Chao-Ping , Hung Iou-Jih , Liu Hsi-Che , Jaing Tang-Her , Yeh Ting-Chi , Liang Sung-Tzu , Chang Chia-Ling , Lee En-Hui , Liu Jo-Chuan , Yao Hsien-Wen , Chang Wan-Hui
Published: October 2019

Abstract 3064

Poster Board III-1

Both ETV6-RUNX1 (TEL-AML1)fusion and hyperdiploidy (>50 chromosomes) of lymphoblasts are favorable outcome predictors in childhood acute lymphoblastic leukemia (ALL). In 433 children with ALL diagnosed at our hospitals between 1997 and 2007 in Taiwan, the frequency of ETV6-RUNX1 fusion was 15.8%, and the frequency of hyperdiploidy (>50 chromosomes) was 14.1%, both were lower than those of the West. While ETV6-RUNX1 fusion had borderline favorable impact on outcome (p=0.053-0.061), hyperdiploidy showed significant favorable impact on event-free survival (91.1% vs 76.6 %, p= 0.016) in our patients. A meta-analysis from literature enrolled reports in which the case numbers and frequency of ETV6-RUNX1 fusion or hyperdiploidy in childhood ALL were described. It revealed that the frequency of ETV6-RUNX1 fusion in childhood ALL in Far East (Japan, Korea, Hong Kong, Chinese in Singapore, and Taiwan) was 14.2% (127/893, range 10-17%), significantly lower than 21.8% (152/697, range 19-27%) in the West (USA, Germany, Italy, France and Chile) (p < 0.0001). The frequency of hyperdiploidy in Japan and Taiwan was 15.2% (140/921, range 13-20%), significantly lower than 31.6% in the West (977/3,158, range 19-34%) (USA, UK and Germany) (p < 0.0001). So far as we know, there were several articles, including ours, addressing that the frequency of ETV6-RUNX1 fusion in childhood ALL was lower in a Far East country. This is the first meta-analysis to demonstrate that the frequency of ETV6-RUNX1 fusion in childhood ALL in Far East was lower than that in the West. There was no report on that the frequency of hyperdiploidy in Far East was lower than that in the West. This is also the first meta-analysis to demonstrate that the frequency of hyperdiploidy in childhood ALL in Far East is significantly lower than that in the West. The nature of these differences, probably due to racial, needs further study. In Far East, with both a lower frequency of ETV6-RUNX1 fusion, and a lower frequency of hyperdiploidy, it warrants renewed effort to cure a higher proportion of children with ALL.

Disclosures

No relevant conflicts of interest to declare.
Total Body Irradiation Causes Residual Bone Marrow Injury by Induction of Persistent Oxidative Stress in Murine Hematopoietic Stem Cells.
Authors: Wang Yong , Liu Lingbo , Pazhanisamy Senthil Kumar , Meng Aimin , Zhou Daohong
Published: October 2019

Abstract 3209

Poster Board III-146

Ionizing radiation (IR) and/or chemotherapy cause not only acute tissue injury but also have late effects including long-term bone marrow (BM) suppression. The induction of residual BM injury is primarily attributable to induction of hematopoietic stem cell (HSC) senescence. However, neither the molecular mechanisms by which IR and/or chemotherapy induce HSC senescence have been clearly defined, nor has an effective treatment been developed to ameliorate the injury, which were investigated in the present study using a total body irradiation (TBI) mouse model. The results showed that exposure of mice to 6.5 Gy TBI induced a persistent increase in reactive oxygen species (ROS) production in HSCs only for up to 8 weeks, primarily via up-regulation of NADPH oxidase 4 (NOX4). This finding provides the foremost direct evidence demonstrating that in vivo exposure to IR causes persistent oxidative stress selectively in a specific population of BM hematopoietic cells (HSCs). The induction of chronic oxidative stress in HSCs was associated with sustained increases in oxidative DNA damage, DNA double strand breaks, inhibition of HSC clonogenic function, and induction of HSC senescence but not apoptosis. Treatment of the irradiated mice with N-acetyl-cysteine (NAC) after TBI significantly attenuated IR-induced inhibition of HSC clonogenic function and reduction of HSC long-term engraftment after transplantation. These findings suggest that selective induction of chronic oxidative stress in HSCs by TBI leads to induction of HSC senescence and residual BM injury and that antioxidant therapy may be used as an effective strategy to mitigate IR- and chemotherapy-induced residual BM injury.

Disclosures

No relevant conflicts of interest to declare.
Expression of FLT3 Internal Tandem Duplication in Pediatric Patients with Acute Myeloid Leukemia and Its Correlation with Multidrug Resistance.
Authors: Qiao Zhenhua , Zhao Jiangning , Xu Lianrong , Lu Quanyi , Niu Xiaoqing , Zhang Peng , Wang Zhao
Published: October 2019

Abstract 4427

FLT3 Internal Tandem Duplication is a kinase mutation frequently observed in cases of adult acute myeloid leukemia (AML). This study was aimed to investigate the expression of FLT3 internal tandem duplication (FLT3-ITD) in pediatric patients with acute myeloid leukemia (AML) and to analyse the clinical features of patients with mutations and the relation of FLT3-ITD with multidrug resistance gene 1 (mdr1). Samples were analysised from eighty-one new diagnosed pediatric patients with AML. The diagnosis was based on French-American-British (FAB) criteria. Immunophenotyping was done by flow cytometry. For each patient, several clinical and biological characteristics were analyzed (age, WHO performance status, WBC count at diagnosis, bone marrow blasts, FAB subtypes, immunologic phenotype, and cytogenetics). The expressions of FIT3-ITD and mdr1 gene in bone marrow samples was determined by RT-PCR. patients were treated with uniform chemotherapy. Statistical analysis were carried by SAS. The results indicated that the FLT3-ITDs were detected in 8 out of 81 pediatric patients (9.88%) and all mutations detected were hybrid, while less frequently this mutation was detected in adult patients. Although they were irrelevant with sex and immunophenotypes, the mutations seemed predominant in older pediatric patients. The average age is 14-year-old in the patients with mutations, but 9-year-old in those without mutation (P=0.039). The leukocyte counts and bone marrow blast cell counts in pediatric patients with FLT3-ITD at diagnosis were higher than those in pediatric patients without FLT3-ITD (median of 58.4×10∧9/L versus 19.49 ×10∧9/L, p = 0.001 and median of 78.5% versus 54.4%,p = 0.041 respectively). The normal chromosomes were found in most pediatric patients with FLT3-ITD, although no significane in the stastistics. The patients with FLT3-ITD had lower induction remission rate (only 25%), but the patients without FLT3-ITD had higher remission rate (76.1%). According results detected by RT-PCR, the mdr1 gene was found in 27 pediatric patients, but only 3 out of 8 pediatric patients with FLT3-ITD were detected to express both FLT3-ITD and mdr1. It is concluded that the FLT3-ITD is a frequent mutation in pediatric patients with AML, especially in the elder pediatric patients. It is related with higher leukocyte counts and bone marrow blast cell counts, which may due to the continual activation of Flt3 tyrosine kinase. The prognosis of these patients with mutations is worse and the induction remission rate is lowe. But the FLT3-ITD not relates with the mdr1, which suggests that the low induction rate is not resulted from the multidrug resistance, at least not from the P-glycoprotein (Pgp) activity. So the common MDR modulators may be uneffective for therapy of the patients with FLT3-ITD.

Disclosures:

No relevant conflicts of interest to declare.
Severe Early-Onset Inflammatory Bowel Disease Caused by IL10 Receptor Deficiency Can Be Cured by Allogeneic Hematopoietic Stem Cell Transplantation.
Authors: Boztug Kaan , Kotlarz Daniel , Glocker Erik , Gertz Mike E , Schäffer Alejandro A , Noyan Fatih , Perro Mario , Diestelhorst Jana , Allroth Anna , Murugan Dhaarini , Hätscher Nadine , Pfeifer Dietmar , Sykora Karl-Walter , Sauer Martin , Kreipe Hans , Lacher Martin , Nustede Rainer , Woellner Cristina , Baumann Ulrich , Salzer Ulrich , Koletzko Sibylle , Shah Neil , Segal Anthony , Sauerbrey Axel , Buderus Stefan , Snapper Scott , Grimbacher Bodo , Klein Christoph
Published: October 2019

Abstract 713

In spite of recent genome-wide association studies, the molecular pathophysiology of many human auto-inflammatory diseases such as enterocolitis remains largely unknown.

Here, we discover the first fully penetrant monogenetic defect causing inflammatory bowel disease (IBD) in humans. Using homozygosity mapping and candidate gene sequencing, we identified three distinct, homozygous mutations in IL10RA, encoding the IL10R1 protein, and IL10RB, encoding the IL10R2 protein, in patients with severe and refractory enterocolitis. IL10R1 is a specific receptor for IL10, whereas IL10R2 is a shared cytokine receptor unit for IL10, IL22, IL26, and IFNλ. The striking similarity of the clinical phenotype between patients with IL10RA and IL10RB deficiency, respectively, suggests that defective IL10-mediated signaling, and not IL22, IL26, or IFNλ dependent effects, is the critical reason for disease.

Deleterious missense mutations in IL10RA abrogate IL10-induced signaling, as shown by deficient phosphorylation of STAT3 at the residue tyrosine 705 in primary patient cells and in HeLa cells engineered to express mutant IL10R1. Mutations in IL10RB introduced a premature stop codon. Defective expression of IL10R2 on the cell surface and deficient STAT3 signaling could be reconstituted by lentiviral gene transfer. As a consequence of defective STAT3 signal transduction in response to IL10 stimulation, IL10R-deficient primary cells showed increased secretion of TNFαa and various other proinflammatory cytokines unresponsive to IL10-dependent negative feedback regulation (TGFβ1, IL1αa, IL1β, IL2, IL6, IL6sR, RANTES, MCP1, MIP1αa, and MIP1β). We are currently assessing whether other IBD patients with early-onset IBD show defects in IL10-mediated signal transduction.

In view of the therapy refractory course of disease and the critical role of IL10 signaling on cells of the hematopoietic system, we have successfully treated two IBD patients with IL10 receptor deficiency by hematopoietic stem cell transplantation (HSCT) without overt side effects. This proof-of-principle suggests that allogeneic HSCT may represent a novel therapeutic approach to treat defined subgroups of IBD patients.

In summary, our results suggest that IL10 receptor defects constitute monogenetic causes for severe, early-onset IBD patients, proving that a lack of IL10-mediated negative feedback signaling perturbs homeostasis of the intestinal immune system.

Disclosures:

No relevant conflicts of interest to declare.
Discontinuation of Imatinib Therapy After Achieving a Molecular Response in Chronic Myeloid Leukemia Patients.
Authors: Mahon Francois-Xavier , Rea Delphine , Guilhot Francois , Huguet Francoise , Nicolini Franck Emmanuel , Legros Laurence , Charbonnier Aude , Guerci Agnes , Varet Bruno R. , Etienne Gabriel , Aton Emilie , Reiffers Josy , Rousselot Philippe
Published: October 2019

Abstract 859

Background

Imatinib (IM) has greatly improved survival rates in chronic myeloid leukemia* (*CML). However, all patients (pts) must continue treatment for an unknown period of time.

A pilot study of the first pts who discontinued IM therapy was previously reported (Rousselot et al. Blood 2007;109:58–60). The multicentre study Stop Imatinib (STIM) was initiated in July 2007 in order to evaluate the persistence of complete molecular remission (CMR) after stopping IM, and to determine the factors that could be associated with CMR persistence.

Methods

Inclusion criteria were IM treatment duration of at least 3 years and sustained CMR. Sustained CMR was defined as BCR-ABL/ABL levels below a detection threshold corresponding to a 5-log reduction (undetectable signal using RQ-PCR) for at least 2 years. Molecular relapse, defined as RQ-PCR positivity, was taken into account if confirmed in two successive assessments. In cases of molecular relapse, pts were re-challenged with IM at 400 mg daily.

Results

From the pilot study, 8 among 15 patients are still in CMR with a median follow up of 42 months (range 37-49). The number of patients enrolled in the STIM study was 69. 34 patients had received interferon alpha (IFNa) prior to IM and 35 pts were de novo. Median follow-up (range) was 17 months (6-24). 37 pts relapsed (loss of CMR) within the first 6 months and two patient relapsed after more than 6 months (M7 ,M18). At M12, the probability of remaining in CMR was 45% (95% CI: 33-56%). For previously treated with IFN (n=34) this probability was 44% (95% CI: 27-59%) versus 46% (95% CI: 29-61%) for de novo pts (p=0.93, overall). All patients in molecular relapse were sensitive again after imatinib re-challenge (decreasing BCR-ABL level, achievement CMR again). Male pts had a better probability of survival without molecular relapse (p=0.02) and a trend was observed for the low Sokal risk group (p = 0.06). Peripheral NK cells counts prior to IM discontinuation were significantly lower in relapse pts (mainly cytotoxic cells CD56dim) as compared to the non relapse pts(p=0.005).

Conclusions

We have confirmed that CMR can be sustained after discontinuation of imatinib with a long follow-up, particularly in male patients and in pts with cytotoxic NK cells in their peripheral blood. Using stringent criteria, it is possible to stop treatment in patients with sustained CMR, even in those treated with IM as a single agent.

(ClinicalTrials.gov number, NCT00478985 [ClinicalTrials.gov])

Disclosures:

Mahon: Amgen: Honoraria; Novartis Pharma: Consultancy, Honoraria, Research Funding; Alexion: Consultancy, Honoraria. Guilhot:Novartis Pharma: Consultancy, Honoraria; BMS: Consultancy, Honoraria.
R-HCVAD/R-MTX-Arac (R-HCVAD) Overcomes Risk Features Associated with Inferior Outcomes In the Treatment of Newly Diagnosed, High-Risk Diffuse Large B-Cell Lymphoma (DLBCL).
Authors: Mato Anthony R , Feldman Tatyana , Zielonka Tania , Bhattacharyya Pritish K. , Campaiola Alexandria , Shah Rina P , Rosario Maria , Bejot Coleen , Stives Susan , Curtin Mary , Vicioso Rosalba , Goldberg Stuart L , Pecora Andrew L , Goy Andre
Published: October 2019

Abstract 1782

Background:

Chemo immunotherapy (R-CHOP) has improved outcomes of both GC (germinal center) and ABC (activated B-cell) subtypes of DLBCL. However outcomes in DLBCL patients treated with R-CHOP with ABC subtype (vs. GC) and/or with poor-risk features (High IPI, high Ki-67) remain inferior. These patients might benefit from more dose-intensive or high-dose therapy approaches. In our practice at The John Theurer Cancer Center, we have employed a risk-adaptive strategy with R-HCVAD to treat patients with DLBCL with aggressive features.

Methods:

Utilizing Kaplan Meier (KM) survival and Cox regression analyses, we conducted a retrospective cohort study to describe the outcome of patients treated with R-HCVAD in the 1st-line setting with the following high-risk features: high Ki-67 (MIB-1), high IPI, multiple extra-nodal (EN) sites, bulky disease or immunohistochemistry (IHC) staining patterns (GC vs. non GC by Hans’ model). The primary study endpoints were PFS and OS. The proportional hazards assumption was met for this analysis.

Results:

45 patients (median age 57, range 34–71 yrs) with newly diagnosed DLBCL treated with R-HCVAD (median 6 cycles, range 1–8) were available for this analysis, representing 1010 total months of follow up at-risk. Baseline characteristics included: stage III-IV (90%), IPI ≥ 3 (52%), median Ki-67 (80%, range 10–100%), median EN sites (2), non-GC subtype (34%), bone marrow (BM) involvement (38%), EBER positive (14%), HIV negative (100%). With 17 months (range 9–64 months) median follow up, median OS and PFS (graph) are not yet reached. 2-yr PFS and OS were 80% (95% CI 61–91%) and 78% (95% CI 61–88%) respectively. In Cox regression analysis, survival outcomes were not adversely affected by: patient age > 60 (HR .8, p=.18), LDH > ULN (HR 2.3, p=.3), non-GC IHC pattern (HR .5, p=.5), BM involvement (HR 1.9, p=.4), Ki-67 ≥ 80% (HR 1.7, p=.6) or EN sites ≥ 2 (HR 4.7, p=.15).

Conclusions:

This analysis represents the largest reported cohort of DLBCL patients treated with R-HCVAD. These data suggest that R-HCVAD can overcome traditional poor-risk features such as high IPI, high Ki-67 and non-GC IHC pattern. Future work will focus on identifying molecular markers for failure in DLBCL patients treated with dose-intensive regimens. A randomized trial comparing R-HCVAD to R-CHOP in selected high-risk patients is warranted.

Disclosures:

No relevant conflicts of interest to declare.
Interim Results of a Phase 1 Trial of An Oral Histone Deacetylase Inhibitor Belinostat In Patients with Lymphoid Malignancies.
Authors: Zain Jasmine M. , Foss Francine M , de Bono Johann S. , Narwal Ameet , Neylon Ellen , Blumenschein George , Lassen U. , Knoblauch Poul , Diefenbach Catherine S. , O'Connor Owen A.
Published: October 2019

Abstract 1787

Background:

Belinostat (Bel) is a histone deacetylase inhibitor with broad preclinical activity. A phase I study of oral Bel in patients (pts) with solid tumors found a recommended dose for day (d) 1–14, q3w, of 750 mg QD, with allowance for intra-patient dose escalation if limited toxicity. The current study was initiated to assess if the same dose could be utilized in pts with lymphoma.

Objectives:

safety and efficacy assessments of oral Bel in cohorts of 3–6 pts (A 750; B 1000; C 1250; D 1500, E 1750; F 2000), treated d 1–14, q3w in pts with relapsed/refractory non-Hodgkin lymphoma (NHL) or Hodgkin's disease (HD) with evaluable disease and acceptable organ functions.

Results:

21 pts, sex (9 F, 12 M) - median age 48 (range 21 – 81), prior regimens, median 5 (range 1 – 13), 12 had BM transplants, including 3 pt with allogeneic have been enrolled. Diagnoses included Mantle Cell lymphoma (MCL; 5 pts), HD (7 pts), other NHL (9 pts).

Median number of Belinostat cycles 3 (range 1–29) 1 pt remains on trial. In 21pts evaluable for tox the most frequent adverse events (regardless of attribution or grade) were: Diarrhea and fatigue (each in 18pts), vomiting (14 pts), cough and constipation (each in 12 pts), anxiety and decreased appetite (each in 11pts). Non-hem gr 3/4 events: 6 pts had gr 3 diarrhea – no dose dependency, – (1 grade 3 diarrhea at 1500-mg dose resulting in hospitalization - DLT) Gr 3 Abdominal pain; cholelithiasis; DVT; hyperthyroidism (each seen in 1 pt;), only gr 4 event was respiratory distress (1 pt in cohort C). Gr 4 Thrombocytopenia (2 pts - Cohort C and E) gr 3 Thrombocytopenia (4 pts - Cohorts, D, E, F) Gr 4 anemia (1 pt - cohort B) Gr 3 anemia (2 pts - Cohort C and D) Leucopenia grade 3 or 4 was not seen-. In 16 pts evaluable for efficacy, stable disease have been noted in 12 pts, including 4- pts with MCL, 4 pts with NHL and 4 with HD. 1 pt with HD had CR after C2 and was treated with additional 2 cycles.

Conclusions:

Oral Bel can be administered safely with a d 1–14, q3w schedule in pts with lymphoma at a daily dose higher than what has been established for pts with solid tumors. Protocol defined DLTs have been encountered in the dose range 750 – 2000mg in pts with lymphoma. Cohort D at 1500 mg was expanded due to 1 DLT (diarrhea). Cohort E at 1750 mg was tolerated without DLT and cohort F 2000 mg is currently being investigated. Final evaluation will include additional pts and possible dose escalation. The safety profile and early tumor shrinkage noted in MCL and HD warrants continued evaluation of Bel, especially in combination with other active compounds.

Disclosures:

Foss: Allos Therapeutics, Inc.: Consultancy, Speaker. Neylon:Allos Therapeutics, Inc.: Honoraria. Knoblauch:TopoTarget: Employment. O'Connor:Allos Therapeutics, Inc.: Research Funding.
Targeting Chronic Lymphocytic Leukemia with Cord Blood NK Cells In NSG Model
Authors: Xing Dongxia , Decker William , Li Sufang , Robinson Simon , Shah Nina , Yang Hong , Champlin Richard E. , Parmar Simrit , Gribben John G. , Bollard Catherine M , Shpall Elizabeth
Published: October 2019

Abstract 2453

The incompatibility between donor killer cell immunoglobulin-like receptors (KIRs) and their corresponding ligands has been reported to reduce the risk of relapse after haploidentical and human leukocyte antigen (HLA) mismatched hematopoietic stem cell transplantation in patients with acute myeloid leukemia. We tested this KIR-ligand mismatch hypothesis in the context of allogeneic cord blood NK cells as an adoptive transfer of lymphocytes treating residual chronic lymphocytic leukemia (CLL). As a model for targeting malignant B cells in CLL, we examined allogeneic cord blood NK cell function in NOD scid gamma (NSG) mice, which carry the null interleukin-2 receptor gamma chain mutation, as the mice developed leukemia. Positively selected CD56+ cord blood NK cells were expanded ex vivo with interleukin-2 for 14 days. CLL cells were established in the NSG model by infusion of CLL cells obtained from patients. The leukemia that develops in NSG mice resembles human CLL, with a proliferating CD19+CD23+CD5+ B-cell population detected in the bone marrow, spleen, lymph nodes, and peripheral blood. Subsequently, expanded cord blood NK cells (5 × 106 per mouse) were intravenously infused into NSG-CLL mice. The NK cells that were infused into the CLL mice were typed for HLA and KIR (four main KIRs: KIR2DL2, KIR2DL3, KIR3DL1, and KIR2DL1). The CLL patients' samples that had been used in the NSG models were genotyped for KIR ligands (HLA-C group or HLA-Bw4 group and HLA-A3). In the six pairs of cord blood NK and CLL cells typed, all were HLA mismatched. Five pairs were KIR-ligand mismatched; these mice showed robust NK cell–mediated killing of CLL cells 7 days after NK cell infusion. Of interest, although no KIR-ligand mismatch was seen between the cord blood NK cells and CLL cells in one pair, we still observed NK cell–mediated killing of CLL cells in the mice. In this instance, NK cell–mediated cell killing could have been attributed to possible lower expression of HLA ligands by leukemic cells. Overall survival was significantly improved in CLL-NSG mice that had received cord blood NK cell treatment compared with overall survival in untreated mice (Kaplan-Meier analysis, p < 0.03). These results showed that adoptive transfer of allogeneic cord blood NK cells can be effective in killing CLL, and will be explored in the clinic.

Disclosures:

Gribben: Roche: Consultancy; Celgene: Consultancy; GSK: Honoraria; Napp: Honoraria.