bluuebunny/crossref_metadata_2025_split huggingface dataset:
Genome-Wide Profiling of Epigenetic and Transcription Factor Regulation In Human Macrophage Differentiation
Authors: Pham Thu-Hang , Lichtinger Monika , Benner Chris , Pape Sabine , Schwarzfischer Lucia , Klug Maja , Gogol-Doering Andreas , Hu Yuhui , Andreesen Reinhard , Chen Wei , Rehli Michael 
Published: October 2019

Abstract
Abstract 3875
The differentiation of human macrophages is accompanied by distinctive phenotypical changes and generally proceeds in the absence of proliferation. The molecular events governing this process are still poorly understood. Using ChIP-Seq technology we studied epigenetic changes as well as alterations in transcription factor occupancy during human monocyte differentiation and correlated these events with gene expression levels in hematopoietic cell types. We show that putative enhancer regions marked by histone H3 lysine4 monomethylation (H3K4me1) at different developmental stages (human progenitor cells, peripheral blood monocytes and in vitro differentiated macrophages) are enriched in distinct sets of transcription factor motifs corresponding to lineage-determining factors. Cell stage-specific histone methylation at promoter-distal sites corresponds with increased mRNA expression levels of neighboring genes. We generated global DNA-binding maps in monocytes and macrophages for two transcription factors (PU.1 and C/EBPβ) with a well established role in monocyte/macrophage differentiation. Comparison of human binding sites with corresponding mouse data revealed a surprisingly low level of conservation (∼10-15%) of PU.1-or C/EBPβ -bound sites between man and mouse, despite a highly conserved binding preference for both transcription factors. During monocytic differentiation, human macrophages primarily gained additional binding sites for both transcription factors (as well as promoter-distal H3K4me1). Interestingly, only neighboring genes with multiple binding events showed significantly increased, macrophage-specific mRNA expression as compared to monocytic as well as lymphocytic cell types. Human macrophage-specific H3K4me1-marked regions as well as macrophage-specific PU.1- and C/EBP-bound sites were characterized by overlapping sets of novel sequence motifs, suggesting that the combinatorial interaction of corresponding DNA-binding factors with PU.1 and C/EBPβ may be required for the establishment of human macrophage-specific enhancers. These data provide novel insights into PU.1 and C/EBPβ mediated gene regulation during human macrophage differentiation.

Disclosures:
No relevant conflicts of interest to declare.
Early Interim 18f-FDG PET In Hodgkin's Lymphoma: Evaluation on 304 Patients
Authors: Zinzani Pier Luigi , Rigacci Luigi , Stefoni Vittorio , Broccoli Alessandro , Puccini Benedetta , Gandolfi Letizia , Fanti Stefano , Castagnoli Antonio , Pellegrini Cinzia , Montini Gian Carlo , Quirini Federica , Antognoli Giulia , Vaggelli Luca , Derenzini Enrico , Argnani Lisa , Ambrosini Valentina , Agostinelli Claudio , Pileri Stefano A. , Bosi Alberto , Baccarani Michele 
Published: October 2019

Abstract
Abstract 3879

Purpose.
The use of early (interim) positron emission tomography (PET) restaging during front-line therapy in Hodgkin's lymphoma (HL) has considerably increased in clinical practice as an early recognition of treatment failure allows patients to be addressed to more intensive treatment regimens.

Patients and Methods.
Between June 1997 and June 2009, 304 newly-diagnosed Hodgkin's lymphoma patients (147 early-stage and 157 advanced-stage) were treated with the ABVD regimen at two Italian institutions. Patients underwent to a PET staging and restaging at baseline, after 2 cycles of therapy and at the end of the treatment.

Results.
53 patients showed a positive interim PET and only 13/53 (24.5%) achieved a complete response (CR), whereas 251 patients showed a negative PET and 231/251 (92%) remained in CR. Comparison between interim PET-positive and interim PET-negative patients indicated a significant association between PET findings and 9-year progression-free survival (p=0.0000) and 9-year overall survival (p=0.0000), with a median follow-up of 31 months.
Among the early-stage patients, 19 had a positive interim PET and only 4 (21%) achieved a CR; among the 128 negative interim PET patients, 122 (97.6%) obtained a CR. In the advanced-stage subset, 34 patients showed a persistently positive PET (with only 9/34, 26.4% in CR), whereas 123 showed a negative interim PET, with 109 (88.6%) remaining in CR.

Conclusions.
Our results confirm the role of early PET as a significant step forward for the management of both early and advanced-stage HL patients, offering the potential for an immediate switch to high-dose treatments, if required.

Disclosures:
No relevant conflicts of interest to declare.
Direct Sensing of Toll-Like Receptor Agonists by Dendritic Cell Progenitors Leads to Their Homing to Inflamed Lymph Nodes Via Regulation of CXCR4 and CCR7
Authors: Schmid Michael A , Baumjohann Dior , Manz Markus G 
Published: October 2019

Abstract
Abstract 3895
Dendritic cells (DCs), the key antigen-presenting cell population, continuously need to be regenerated from bone marrow (BM) hematopoietic stem and progenitor cells. Common dendritic progenitors (CDP) were previously shown to efficiently generate DCs in lymphoid and non-lymphoid tissues. How the dissemination of bone marrow (BM) DC-progenitors to peripheral tissues is regulated upon demand remains elusive to date. Acute microbial infections are sensed via Toll-like receptors (TLR). Recent studies showed that stem and progenitor cells express TLRs. We found that CDPs in the BM of mice express relative high levels of Tlr2, Tlr4 and Tlr9, and hypothesized that these might be involved in regulating CDP migration. CDPs in steady-state expressed high levels of Cxcr4, but no, or low Ccr7. Upon direct stimulation with the respective TLR-agonists in vitro, CDPs rapidly down-regulated Cxcr4 and up-regulated Ccr7 mRNA and protein. CDPs that were stimulated with TLR-agonists for only 2 h preferentially homed to the lymph nodes (LN) in expense of BM in steady-state recipients. When TLR-agonists were injected subcutaneously, CDPs gave rise to increased numbers of plasmacytoid DCs, classical DCs, and DCs with a skin-derived migratory phenotype in inflamed LNs on day 4. This was not due to increased proliferative activity. Injecting the CXCR4 antagonist AMD3100 demonstrated that the retention of CDPs in the BM depends on CXCR4. Furthermore, CCR7 was important for the engraftment of CDP-derived DCs into LNs in steady-state and during inflammation. In conclusion, DC progenitors in the bone marrow are capable to directly sense TLR-agonists via their cognate receptors in systemic infections. This results in differential expression of chemokine receptors and consecutive migration of DC-progenitors to inflamed LNs. This mechanism helps to restore DC subsets during ongoing immune responses and to return to DC homeostasis once the inflammation ceases.

Disclosures:
No relevant conflicts of interest to declare.
Lestaurtinib (CEP701) Inhibits Proliferation and Activates Apoptosis In Hodgkin Lymphoma Cells through Inhibition of the JAK/STAT Signaling Pathway
Authors: Navarro Alfons , Díaz Tania , Martinez Antonio , Gaya Anna , Monzó Mariano 
Published: October 2019

Abstract
Abstract 3919

Background:
The constitutive activation of the JAK/STAT pathway plays an important role in the pathogenesis and proliferation of Hodgkin Lymphoma (HL). Although somatic activating point mutations in the JAK2 gene have been reported in myeloproliferative disorders (MPD), they are rarely described in HL, where JAK2 amplification is associated with mutations of regulator genes such as SOCS-1, constitutive activation of STAT proteins or miRNA deregulation. Recently, many JAK2 inhibitors, including Lestaurtinib (CEP701), have been reported to have clinical efficacy in MPD. CEP701 is a multitargeted tyrosine kinase inhibitor that potently inhibits FLT3 at nanomolar concentrations. Recent studies in MPD have further shown that CEP701 inhibitory activity is not limited to FLT3 and can suppress JAK2/STAT5 signaling through JAK2 inhibition. As a first step towards elucidating the potential role of CEP701 in HL therapy, we have analyzed its efficacy in vitro.

Methods:
Four HL cell lines, L-428, L-1236, HDMYZ and L-540, were assayed for proliferation, apoptosis and levels of proteins in the JAK2/STAT pathway (pJAK2, JAK2, pSTAT5, STAT5, Bcl-xL) after CEP701 treatment. 100,000 cells were plated in a 96-well plate in 100 ml culture medium with CEP701 or DMSO (vehicle control) at concentrations of 30–300 nM. After 1 or 24 hours of incubation with CEP701, the levels of the proteins and of FLT3 were analyzed by Western blot. Proliferation was analyzed with CellTiter 96 AQueous One Solution Cell Proliferation Assay (MTS) and apoptosis by CaspaseGlo 3/7 after 48 hours of treatment.

Results:
The proliferation analysis showed an effective dose-dependent inhibition of cell growth in the 4 HL cell lines after treatment with increasing concentrations of CEP701. At 48h, in comparison to cells treated with DMSO alone (normalized to 100%), in cells treated with 100nM of CEP701, we observed a marked inhibition of 35% in L-428, 55% in L-1236, 15% in HDMYZ and 77% in L-540. Moreover, apoptosis increased by 38%, 31%, 21% and 25%, respectively. The protein analysis showed that after one hour, CEP701 inhibited phosphorylation of JAK2 (pJAK2) and its downstream target STAT5 (pSTAT5) in a dose-dependent manner, with no changes in the non-phosphorylated proteins. The downstream target Bcl-xL also decreased.

Conclusions:
Taken together, these data demonstrate that growth inhibition and apoptosis activation by CEP701 in HL cells correlates with the inhibition of the JAK2/STAT5-dependent signal transduction pathway. Here we present the first biological evidence that Lestaurtinib could be a promising new agent in the treatment of patients with HL.
Supported by a FIS grant (PS09/00547).

Disclosures:
No relevant conflicts of interest to declare.
Mimetic Peptide to CDK Inhibitor p16 INK4a Induces Cell Death In Mantle Cell Lymphoma Cells: A New Strategy to Eradicate Minimal Residual Disease
Authors: Liu Hongtao , Koval Greg , Malnassy Greg , Ihonor Pamela , Liu Hui , Luo Roger , Wickrema Amittha , Dowdy Steven , Thirman Michael , van Besien Koen , Stock Wendy 
Published: October 2019

Abstract
Abstract 3921
Autologous stem cell transplantation (ASCT) is an effective treatment strategy for Mantle Cell Lymphoma (MCL) that has been shown to improve disease free survival. However, data from recent trials suggest that the presence of minimal residual disease (MRD) contributes to relapse following current intensive treatment strategies, including ASCT. Thus, we sought to test a highly selective approach to eradication of MRD in stem cell collections from MCL patients using targeted small peptides that disrupt the aberrant cyclin/CDKs interactions that are involved in the pathogenesis of MCL. In the current study, the efficacy of a mimetic peptide to the CDK inhibitor p16INK4a to eliminate MRD was tested. Using (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) (MTS) cell viability assays, we demonstrated that a transducible TAT-p16 mimetic peptide was able to induce cell death in the MCL cell line, Jeko-1, in a dose and time-dependent fashion (see Figure). A TAT-p16 mimetic was able to induce cell death in approximately 60% of Jeko-1 cells after 4 hours of incubation, and approximately 80% of Jeko-1 cells at 24 hours. In contrast, the same dose of TAT-p16 mimetic had no effect on cell survival when it was incubated with CD34+ rich apheresis samples from healthy donors. Furthermore, CFU-GM, BFU-E colony assays showed that there was no toxicity of the TAT-p16 mimetic when compared to untreated cells or cells treated with a control, scrambled TAT-p16 peptide. TAT-p16 mimetic did not impair erythroblast differentiation but induced mild apoptotic cell death in the erythroblast cells. The strategy was further tested using a CD34+ enriched apheresis sample collected from a patient with MCL with documented MRD who had been treated on a clinical trial of ASCT for MCL. Using Real-time quantitative PCR for IgH and BCL1 gene rearrangements, we demonstrated that the TAT-p16 mimetic was able to reduce MRD level by 40% compared with the scrambled TAT-p16 peptide. These data suggest that the use of a peptide-mimetic to p16 selectively and effectively can reduce MRD in MCL cells and in an apheresis (stem cell) sample from a patient with MCL. Treatment with the peptide allowed differentiation of CD34+ progenitor cells. A combinational approach using additional targeted cell cycle regulatory peptides that block p21CIP1/WAF1 and cyclin D2 is being explored to optimize the efficacy of this purging technique. Thus, this novel strategy may be an effective, selective and non-toxic method for eradication of MRD in MCL.

Disclosures:
No relevant conflicts of interest to declare.
A Small Molecule PLK1 Inhibitor, MLN0905, Induces An Anti-Tumor Response In Human Models of Diffuse Large B-Cell Lymphoma
Authors: Shi Judy , Lasky Kerri , Shinde Vashali , Duffey Matt O , Stringer Bradley , Qian Mark , Liao Debra , Liu Ray , Rao Youlan , Vos Tricia J , D'Amore Natalie Roy , Hyer Marc L 
Published: October 2019

Abstract
Abstract 3922
Diffuse large B-cell lymphoma (DLBCL) is the most common of the non-Hodgkin lymphomas, accounting for up to 30 percent of newly diagnosed cases. Current treatment options for this disease are effective, but not always curative; therefore experimental therapies are being investigated. Diffuse large B-cell lymphoma cells typically over-express the serine/threonine mitotic kinase PLK1, which plays a key role in mitotic cell cycle progression, and has been linked to poor patient prognosis. We have discovered a potent and selective small molecule inhibitor of PLK1, MLN0905, which inhibits cell proliferation in a broad range of human tumor cells including DLBCL lines. We explored the pharmacokinetic, pharmacodynamic, and anti-tumor properties on MLN0905 in mouse models harboring human DLBCL disease. MLN0905 has drug-like pharmacokinetic properties and an acceptable toxicity profile making it a good clinical candidate. In human xenograft tumor tissue, MLN0905 modulates the pharmacodynamic biomarker phospho-Histone H3 (in a dose dependent fashion), enabling us to track pathway modulation in vivo. The anti-tumor activity of MLN0905 was evaluated in three human subcutaneous xenograft models OCI-LY10, OCI-LY19, and PHTX-22L (primary lymphoma). In each model, MLN0905 yielded significant anti-tumor activity on both a continuous (daily) and intermittent dosing schedule, underscoring dosing flexibility. The anti-tumor activity of MLN0905 was also evaluated in a disseminated xenograft (OCI-LY19) setting to better mimic DLBCL in humans. In this disseminated model, MLN0905 induced a highly significant survival advantage. Finally, MLN0905 was combined with Rituximab in the disseminated OCI-LY19 model. Combining Rituximab and MLN0905 provided both a synergistic anti-tumor effect and a synergistic survival advantage. Our findings indicate for the first time that PLK1 inhibition leads to pharmacodynamic pH3 modulation and significant anti-tumor activity in multiple models of DLBCL. These data strongly suggest evaluating PLK1 inhibitors as DLBCL anti-cancer agents in the clinic.

Disclosures:
Shi: Millennium: The Takeda Oncology Company: Employment. Lasky:Millennium: The Takeda Oncology Company: Employment. Shinde:Millennium: The Takeda Oncology Company: Employment. Duffey:Millennium: The Takeda Oncology Company: Employment. Stringer:Millennium: The Takeda Oncology Company: Employment. Qian:Millennium: The Takeda Oncology Company: Employment. Liao:Millennium: The Takeda Oncology Company: Employment. Liu:Millennium: The Takeda Oncology Company: Employment. Rao:Millennium: The Takeda Oncology Company: Employment. Vos:Millennium: The Takeda Oncology Company: Employment. D'Amore:Millennium: The Takeda Oncology Company: Employment. Hyer:Millennium: The Takeda Oncology Company: Employment.
Rituximab Plus Chlorambucil Compared with Chlorambucil and Prednisone In Patients with Untreated Follicular Lymphoma
Authors: Bassi Simona , Sammassimo Simona , Gigli Federica , Pruneri Giancarlo , Bertazzoni Paola , Quarna Jessica , Cocorocchio Emilia , Laszlo Daniele , Negri Mara , Lionetti Maria , Vanazzi Anna , Alietti Alessandra , Agazzi Alberto , Gardellini Angelo , Andreola Giovanna , Preda Lorenzo , Pastano Rocco , Martinelli Giovanni 
Published: October 2019

Abstract
Abstract 3956

Background:
Recently in large randomized studies, the addition of Rituximab to standard chemotherapy in the first-line treatment of follicular lymphoma (FL) demonstrated improvements in long-term outcome in FL.

Methods:
We compared the outcome of 111 naive FL patients (pts) treated at our institution from 1995 to 2009 with a single alkylating agent in combination or not with Rituximab: 58 pts received Chlorambucil plus Rituximab (R-Chl) and 53 pts Chlorambucil and prednisone (Chl+PDN). The 2 schedules included an induction phase, where Chlorambucil was given at 6mg/mq for 6 consecutive weeks in both groups and the Rituximab in 4 weekly administrations. The maintenance phase was longer in the Chl+PDN group: Chlorambucil was administered 6mg/mq daily 14 days each month for a total of 12-months of treatment versus 14 days with monthly-Rituximab for 4 consecutive months in the R-Chl group.

Results:
The demographic and prognostic factors are reported in Table 1.
At the end of treatment the ORR was 98% in the R-Chl pts and 77% in the Chl+PDN group, with a percentage of complete response of 79% and 55% respectively. No significant incidence of adverse events were reported and only one HBsAg positive patient in R-Chl group discontinued the therapy. One case of myelodysplastic syndrome was described in a Chl+PDN relapsed patient. With a median follow up of 34 months, the OS and the EFS in R-Chl pts is 95% and 76% respectively. Otherwise, with a median observation time of 82 months in the Chl+PDN group 73% are alive and only 28% of pts maintained the response. The median time to subsequent therapy (TTST) is 21 and 15 months respectively in the 2 groups.

Conclusions:
The addition of Rituximab to alkylating in first-line treatment in previously untreated FL pts improves significantly the EFS and prolongs the time to next antilymphoma therapy compared with alkylating alone. Because of its low toxicity profile, our combination with Rituximab and Chlorambucil could be considered a first-line therapy, especially in FL patients not eligible for aggressive chemotherapy regimens.

Disclosures:
No relevant conflicts of interest to declare.
Anti-Leukemic Property of Sulphoraphane In Precursor B Cell Acute Lymphoblastic Leukemia
Authors: Suppipat Koramit , Zhu Xiao , Park Chun Shik , Lacorazza H. Daniel 
Published: October 2019

Abstract
Abstract 3974
Acute lymphoblastic leukemia (ALL) is the most common form of hematologic malignancy in children. In spite of significant advances achieved in the treatment of childhood ALL, one fifth of these patients still relapse after the standard treatment. Moreover, relapse ALL is the second most common cause of cancer-related deaths in children. The development of novel therapies is prevented by a limited understanding of the exact pathobiology. There are emerging evidences that the transcription factor KLF4 has a tumor suppressor property in ALL. Recently, a gene expression classifier study in pediatric precursor B-cell ALL (pre-B ALL) showed that KLF4 expression was significantly reduced in high risk ALL patients with positive MRD after induction. This finding suggests a possible role of this cell cycle inhibitor on the development, expansion and drug-resistant of leukemic cells. Several studies demonstrate that overexpression of KLF4 in normal B cells and BCR transformed B cells show increased apoptosis and reduced proliferation. Furthermore, we recently described that KLF4 inhibits proliferation of naïve lymphocytes by activating p21 (Yamada, et al, 2009). Sulphoraphane (SF; 4-methylsulfonylbutyl isothiocyanate) is a dietary compound derived from Cruciferae vegetables with anti-carcinogenic activity in colon cancer by upregulating KLF4 and p21 among other genes. Thus, we hypothesized that SF could also exhibit anti-leukemic activity in human-derived acute lymphoblastic leukemia cells via the activation of KLF4. The pre-B ALL cell lines (Nalm6, REH, RS-4, SUP-B15) and an EBV transformed B cell line were treated with different concentrations of SF (0-40 μM) for 24–48 hours. Then, cell number was estimated using an ATP-based viability method. Flow cytometric analysis of ANNEXIN-V/7-AAD binding as well as CFSE dilution was used to measure apoptosis and proliferation respectively. We found that SF induced cytotoxicity in Nalm-6, REH and RS-4 cell lines in a dose and time dependent manner. This cytotoxic effect was less pronounced in EBV-transformed B cell line. SF treatment led to increased ANNEXIN-V and 7-AAD positive cells (82% apoptotic cells in SF-treated versus 9% in DMSO control). Further, SF-treated cells displayed significantly less proliferation in comparison to DMSO controls thus suggesting that SF inhibits cellular proliferation. Preliminary data also suggest that SF-mediated apoptosis is caused by upregulation of KLF4. In conclusion, Sulphoraphane exhibits an anti-leukemic property by inducing apoptosis and abrogating proliferation in pre-B ALL cell lines. Thus, sulphoraphane could potentially be used as an adjunctive therapy in a subgroup of pre-B ALL patients who have decreased expression of KLF4.

Disclosures:
No relevant conflicts of interest to declare.