bluuebunny/crossref_metadata_2025_split huggingface dataset:
Therapeutic Granulocyte Transfusions in Neutropenic Patients with Invasive Pulmonary Aspergillosis,
Authors: Drognitz Kathrin , Lubbert Michael , Illerhaus Gerald , Gartner Frank , Bertz Hartmut 
Published: October 2019

Abstract
Abstract 3372

Background:
Granulocyte transfusions (GTX) are used as an additional therapeutic option in patients with severe neutropenia following chemotherapy constituting an increased risk for life-threatening bacterial and fungal infections. We hypothesized that interventional GTX would provide a clinical benefit for neutropenic patients with invasive pulmonary aspergillosis (IPA). Methods: We reviewed the clinical outcome of 44 patients with severe neutropenia (46 cases) and underlying hematological malignancies suffering from IPA unresponsive to standard antifungal therapy. They received a total of 181 human recombinant granulocyte colony-stimulating factor (rh G-CSF) stimulated GTX at Freiburg University medical center from 1996 – 2009. Neutropenias were caused by conventional chemotherapy (n=38), conditioning chemotherapy for allogeneic (n=4) and autologous (n=2) hematopoietic cell transplantation (HCT), aplastic anemia (n=1) and by intense immunosuppressive (n=1). Donors were exclusively relatives and acquaintances of the recipients. IPA was diagnosed by clinic, computed tomography (CT) scan, serological or microbiological measures. Response of GTX was evaluated by repeated CT, decreasing C-reactive protein (CRP) levels, hematopoietic regeneration and clearance of serum galactomannan antigen. Results: A median of 3 GTX (range 1–25) containing a median total of 59.4×109 (range 3–170) white blood cells per GTX were administered. All but five (3%) transfusions were well tolerated. Median duration of neutropenia proceeding GTX was 15.5 d (range 4–70). Resolution of infection or clinical improvement was achieved in 29 (63%) patients with IPA and haematopoietic recovery has been assumed within 10 days after the last GTX in 34 patients (74%). Thirty-three (72%) patients were alive one month after the first GTX. Overall, progressive malignant disease was the main cause of death. Patients not responding to GTX died without on septic complications despite appropriate antibacterial and antifungal treatment. Nine out of 26 neutropenic patients receiving GTX after conventional chemotherapy underwent allogeneic HCT later on after control of IPA. Conclusions: Rh G-CSF stimulated GTX are a safe and effective therapeutic tool for patients with hematological malignancies suffering from profound neutropenia and antifungal-therapy resistant IPA. GTX may serve as a bridging therapy in severe neutropenic patients with IPA scheduled for allo-HCT.

Disclosures:
Bertz: GILEAD: Research Funding; MSD: Membership on an entity's Board of Directors or advisory committees.
Regulation of Hematopoietic Cell Engraftment and Myeloid Differentiation by the E3 Ubiquitin Ligase F-Box Protein SKP2,
Authors: Rodriguez-Rodriguez Sonia , McGuigan Declan , Wang Lin , Mumaw Christen , Carlesso Nadia 
Published: October 2019

Abstract
Abstract 3397
The precise molecular mechanisms that coordinate, at cellular level, the cell fate decisions of self-renewal and differentiation during normal adult hematopoiesis are not yet well defined. SKP2 protein is necessary for ubiquitination and degradation of several cell cycle inhibitors, such as p27Kip1, p21Cip1 and p57Kip2. SKP2 overexpression causes quiescent cells to enter the cell cycle and its downregulation is critical for cell cycle arrest. SCFSKP2 is undoubtedly the major ubiquitin ligase regulating the abundance of cell cycle regulatory proteins at the G1-S transition. Thus, we sought to explore its role in regulating cell cycle entry of hematopoietic cells.
By using a mouse model of SKP2 deletion, we show that SKP2 inactivation retards entry of HSC and progenitors into cell cycle, resulting in enhanced HSC quiescence, increased pool size and increased HSC maintenance. These effects were accompanied by increased cellular levels of p27Kip1, p21Cip1 and p57Kip2 in HSC and progenitors. Competitive repopulation assays and serial bone marrow (BM) transplantations showed that loss of SKP2 improved hematopoietic engraftment at long-term. Conversely, the slower cell cycle entry induced by SKP2 deletion greatly impaired hematopoietic engraftment at short-term. We also analyzed the effect of SKP2 deletion on myeloid differentiation. Although the loss of SKP2 did not affect significantly the representation of Gr1+Mac1+ cells in the BM of SKP2 null mice at steady-state, SKP2 null BM lineage negative cells showed an accelerated myeloid differentiation in vitro. Similar results were obtained by using a shRNA anti-SKP2. This effect required p27Kip1 expression, as deletion of SKP2 did not increased differentiation in a p27Kip1 null background, and was, surprisingly, cell cycle-independent.
Taken together, these results demonstrate a previously unrecognized role for SKP2 in regulating rates of hematopoietic engraftment and myeloid differentiation in the BM. The identification of SKP2 as a physiologic regulator of cell cycle progression of HSC and progenitors has significant implications in the biology of hematopoietic stem cells. We believe that these results may contribute to a better understanding of the dynamics of hematopoietic recovery during BM transplantation, as well as of the mechanisms involved in the HSC exhaustion driving BM failure syndromes and may provide new insights for therapeutic applications.

Disclosures:
No relevant conflicts of interest to declare.
The Role of the Intrinsically Disordered and Multifunctional AF9 C-Terminal Domain in MLL-AF9 Leukemia,
Authors: Bushweller John H. , Leach Benjamin , Chang Ming-jin , Lumba Bhavna , Cierpicki Tomasz , Zeleznik-Le Nancy J. , Hemenway Charles S. 
Published: October 2019

Abstract
Abstract 3464
Mixed Lineage Leukemia (MLL) fusion proteins disrupt HOX gene regulation through recruitment of transcriptional elongation factors, leading to acute leukemia. AF9 is one of the most common MLL fusion partners and has roles in normal regulation of HOX genes. AF9 has also been shown to interact with multiple transcriptional regulators, including AF4 family proteins which recruit and activate P-TEFb, suggesting a pivotal role in regulation of transcriptional elongation at HOX loci. The mechanism by which AF9 regulates normal transcription and contributes to dysregulated transcription in MLL-AF9 leukemia is poorly understood. We have recently shown that the AF9 ANC1 Homology Domain (AHD), which is retained in MLL fusion proteins, is intrinsically disordered, i.e. it is unstructured on its own, and recruits AF4 family members by coupled folding and binding. In order to further understand this behavior, we have determined the solution structure of the AF9 AHD in complex with AF4 and identified a point mutation which selectively disrupts the AF4-AF9 interaction. Introduction of this mutation into MLL-AF9 results in loss of HOXA9 upregulation and loss of transformation of hematopoietic progenitor cells, establishing the importance of this interaction for MLL-AF9 leukemia. We propose that AF9 functions as a signaling hub which regulates transcriptional elongation by alternative recruitment of transcriptional regulators. Indeed, the AF9 ANC1 homology domain (AHD) has been shown to also bind the H3K79 methyltransferase Dot1L, human polycomb 3 (hPC3), and the BCL6 corepressor, necessitating a carefully orchestrated choreography of binding partners to regulate transcription. Intrinsic disorder may facilitate dynamic exchange between binding partners to mediate rapid changes of HOX gene expression in both normal tissues, as required for specific differentiation programs, and in leukemia. Our results also validate the AF9-AF4 interaction as a potential target for therapeutic development.

Disclosures:
No relevant conflicts of interest to declare.
FLT3-ITD Signaling Induces Oncogenic Mir-155 by NF-κB and STAT5 Pathways In Acute Myeloid Leukemia Thereby Targeting Transcription Factor PU.1,
Authors: Gerloff Dennis , Braeuer-Hartmann Daniela , Katzerke Christiane , Hartmann Jens-Uwe , Mueller-Tidow Carsten , Tenen Daniel G. , Niederwieser Dietger , Behre Gerhard 
Published: October 2019

Abstract
Abstract 3469
Almost 30% of all acute myeloid leukemias (AML) are associated with an internal tandem duplication (ITD) in the juxtamembrane of FLT3. This mutation leads to constitutively activated FLT3 signaling, which also includes altered FLT3 targets like STAT5. The dysregulation of pathways causes a differentiation block and plays a role in inhibition of hematopoietic transcription factors like PU.1 and C/EBPalpha. Here we report that FLT3-ITD signaling induces the oncogenic miR-155 by NF-κB and STAT5 pathways. Furthermore we demonstrate that miR-155 targets the myeloid transcription factor PU.1. Analysis of FLT3-ITD positive patient samples show an approximately 2 fold higher miR-155 expression compared to FLT3-WT associated AMLs. Besides, the overexpression of FLT3-ITD in myeloid U937 cells increases miR-155 expression 2,2 fold. In contrast, a block of FLT3-ITD signalling in FLT3-ITD associated cell line MV4;11 by protein kinase inhibitors (PKIs), PKC412, SU5614 and CEP701, results in an 80% decreased miR-155 expression. In further experiments we analyzed the role of FLT3-ITD downstream targets NF-κB (p65) and STAT5 in miR-155 regulation. We show that siRNA mediated knockdown of NF-κB (p65) or STAT5 in MV4;11 cells correlates with reduced miR-155 expression. In addition we demonstrate that p65 binds to the miR-155 promoter in MV4;11 cells while the treatment with the PKI CEP701 results in a p65 release from the miR-155 promoter. Furthermore, we prove that miR-155 overexpression in PMA (phorbol 12-myristate 13-acetate) treated U937 cells reduces macrophage differentiation about 50%. In in silico analysis we found PU.1 as a putative miR-155 target. Interestingly, we could show that overexpression of FLT3-ITD and miR-155 reduces the PU.1 protein level in U937 cells, respectively. These data elucidate the regulation and function of miR-155 in FLT3-ITD associated AML and may lead to a better understanding of the role of miRNAs in leukemogenesis.

Disclosures:
No relevant conflicts of interest to declare.
Genome-Wide shRNA Screen Identifies WEE1 As a Critical Mediator of Cell Fate and Novel Therapeutic Target in AML,
Authors: Porter Christopher C. , Kim Jihye , Fosmire Susan , Gearheart Christy M. , van Linden Annemie , Zaberezhnyy Vadym , Gao Dexiang , Tan Aik Choon , DeGregori James 
Published: October 2019

Abstract
Abstract 3503
Acute myeloid leukemia (AML) remains a therapeutic challenge despite increasing knowledge of the molecular origins of the disease, as the mechanisms of AML cell escape from chemotherapy remain poorly defined. We hypothesized that AML cells are addicted to specific molecular pathways in the context of chemotherapy and used complementary approaches to identify these addictions. Using novel molecular and computational strategies, we performed genome-wide shRNA screens to identify proteins which mediate AML cell fate in cytarabine in 2 AML cell lines (Molm13 and MV4-11). Over 55,000 shRNAs targeting over 11,000 genes were quantified by deep sequencing to identify shRNAs under-represented in the context of cytarabine as compared to no treatment. Complementary analyses identified 125 genes as mediators of AML cell fate in cytarabine. In addition we performed gene expression profiling of AML cells exposed to cytarabine to identify genes with induced expression in this context and examined existing gene expression data from primary patient samples. The integration of these independent analyses strongly implicates cell cycle checkpoint proteins, particularly WEE1, as critical mediators of AML cell fate in cytarabine. Knockdown of WEE1 in a secondary screen confirmed its role in AML cell survival in cytarabine. Pharmacologic inhibition of WEE1 in several, but not all, AML cell lines is synergistic with cytarabine, suggesting underlying molecular susceptibility to this combination of drugs. A WEE1 inhibitor is in Phase II trials in solid tumors, primarily as a means to abrogate the G2/M checkpoint in tumors with TP53 dysfunction. Further experiments demonstrate that inhibition of WEE1 prevents slowed S-phase progression induced by cytarabine in AML cells, broadening the functions of WEE1 that may be exploited therapeutically. Preliminary experiments indicate synergistic inhibition of AML cellular proliferation with daunorubicin in some AML cell lines. Experiments to determine whether WEE1 inhibition in combination with chemotherapy prolongs survival of mice with leukemia are underway. These data highlight the power of integrating functional and descriptive genomics, and identify WEE1 as potential therapeutic target in AML.

Disclosures:
No relevant conflicts of interest to declare.
IDH1/2, TET2 and DNMT3A Mutations Are Not Mutually Exclusive in Secondary Acute Myeloid Leukemias,
Authors: Kosmider Olivier , LaRochelle Olivier , Coude Marie-Magdelaine , Mas Veronique Mansat-De , Delabesse Eric , Cornillet-Lefebvre Pascale , Blanchet Odile , Recher Christian , Delmer Alain , Raynaud Sophie , Lacombe Catherine , Bernard Olivier , Ifrah Norbert , Dreyfus Francois , Fontenay Michaela 
Published: October 2019

Abstract
Abstract 3556
IDH1/2, TET2 and DNMT3A mutations have been reported in myeloid malignancies including de novo AML. In this study, we have analyzed the frequency and prognostic impact of these mutations in a large retrospective cohort of patients (pts) with secondary AML (SA) which encompass myelodysplasia-related changes (MRC) AML and therapy-related (TR) AML according to the WHO classification.
Bone marrow samples were collected from 247 pts at diagnosis with SA and the mutational status of IDH1/2, TET2 and DNMT3A genes together with other genes frequently mutated in AML (NPM1, FLT-3, N and K-RAS, WT1) was determined by Sanger sequencing or high resolution melting analysis.
The cohort of 247 pts consisted in 201 MRC AML and in 46 TR AML, 39.5% of which with a normal karyotype (NK). The frequency of IDH1/2, TET2 and DNMT3A mutations was 12.6, 19.8 and 4.5%, respectively. Two pts had both TET2 and IDH1/2 mutations, 2 pts had TET2 and DNMT3A mutations and 5 pts had both IDH1/2 and DNMT3A mutations showing that these mutations were not mutually exclusive in SA. IDH1/2 and TET2 mutations were significantly more frequent in MRC AML (14.1 and 22.3%) than in TR AML (6.4 and 8.7%) (P =0.04 and P =0.03) while the frequency of DNMT3A mutations was identical in the two subgroups.
The SA pts harbouring at least one IDH1/2 or TET2 or DNMT3A mutation were significantly older (P <0.0001) and presented higher leukocyte count and lower MCV (P <0.05) than unmutated pts. Percentage of blasts in the bone marrow was similar in the two groups. Karyotype was normal in 48% of the IDH1/2 or TET2 or DNMT3A mutated pts and 18% of the unmutated patients, indicating that these mutations were strongly associated with NK (P < 0.001). A statistically significant link was found between TET2 or IDH1/2 or DNMT3A mutations and NPM1 mutations, but not with FLT-3, N/K-RAS or WT1 mutations.
Complete remission rate and overall survival were evaluated in a group of 158 pts which had received intensive chemotherapy at diagnosis, and were identical in the IDH1/2 or TET2 or DNMT3A mutated and unmutated groups. These mutations did significantly influence survival neither in the subgroup of pts with normal karyotype, nor in the subgroup of MRC-AML, or TR-AML which were of very poor prognosis.
These data show that IDH1/2, TET2 or DNMT3A mutations could modify the clinical presentation without impact on prognosis.

Disclosures:
No relevant conflicts of interest to declare.
Activating Mutations of the FMS-Like Tyrosine Kinase-3 (FLT3) At Complete Response and Relapse in Patients with Acute Myeloid Leukemia,
Authors: Nazha Aziz , Cortes Jorge E. , Faderl Stefan , Pierce Sherry , Garcia-Manero Guillermo , O'Brien Susan , Ferrajoli Alessandra , Kadia Tapan , Borthakur Gautam , Luthra Rajyalakshmi , Kantarjian Hagop M. , Ravandi Farhad 
Published: October 2019

Abstract
Abstract 3557
Background – Activating mutations of the transmembrane receptor tyrosine kinase, FLT3, occur in approximately 30% of patient with acute myeloid leukemia (AML) and predict for a shorter relapse-free and overall survival. There is limited data on loss or persistence of the mutated clones at the time of complete response (CR) and their recurrence at the time of relapse.

Objectives and Methods –
To evaluate patterns of loss and recurrence of FLT3 mutated clones in relation to response and relapse in patients with FLT3 mutated AML treated with idarubicin and cytarabine (IA) with or without sorafenib (S), vorinostat (V), or tipifarnib (T). Bone marrow samples at diagnosis, CR and relapse were examined for the presence of FLT3 mutated clones using reverse transcription polymerase chain reaction.

Results –
Among 361 patients with AML treated from October 2004 to March 2010 on one of the 4 induction regimens of IA, IAS, IAV, and IAT, 321 had presentation bone marrow samples tested and 72 had FLT3 mutations (including 50 with ITD and 16 D835 with 6 having both). The median age for the entire group was 53 years (range, 17–73) and for the FLT3 mutated patients 52 years (range, 17 to 73). Cytogenetics at diagnosis in FLT3 mutated patients included normal karyotype in 48 (67%) patients, chromosome 5 and 7 abnormalities in 4(6%), trisomy 8 in 4(6%), 11q abnormalities in 2 (3%), insufficient metaphases in 3(4%), and miscellaneous in 11(16%). 271 (75%) patients overall, and 64 (89%) patients with mutated FLT3 achieved CR. Among the 56 patients with presentation FLT3-ITD, 51 achieved CR. Among 13 patients with available samples at CR, none had FLT3-ITD; 8 of these patients relapsed and 5 had FLT3-ITD positive clones at relapse (2 negative and 1 not done); Among the 38 patients with no samples at CR, 17 relapsed, 8 with a FLT3-ITD clone (1 negative and 8 not done). Among the 201 patients without FLT3-ITD at diagnosis, who achieved CR, 8 patients acquired a clone with FLT3-ITD at relapse.

Conclusions –
FLT3-ITD mutant clones are unstable at follow-up. Relapse may occur in their absence and they may occur for the first time at relapse. Therefore, FLT3-ITD cannot be used reliably for minimal residual leukemia monitoring.

Disclosures:
Ravandi: Bayer Onyx: Honoraria, Research Funding.
Improved Survival Rates Among Acute Myeloid Leukemia (AML) Patients 65 to 74 Years: Population-Based Estimates Over Three Decades,
Authors: Thein Mya S , Jemal Ahmedin , Baer Maria R. , Ershler William B , Yates Jerome W 
Published: October 2019

Abstract
Abstract 3591

Background:
AML is the common acute leukemia in adults, accounting for approximately 90% of all acute leukemias in those over the age of 18 years. Overall survival remains poor, with <50% 5-year survival in patients under 45 years and <5% in those over 65 years. We examined whether there has been improvement in survival over three successive decades for subsets of geriatric AML patients. Methods: Examining data from the Surveillance, Epidemiology and End Results (SEER) database, we determined relative overall survival (+/− standard error) of patients with AML 65–74, 75–84 and 85+ years of age between 1977–86, 1987–96 and 1997–2006. Results: Survival for both males and females at 12, 24 and 36 months post-diagnosis is shown below. Between 1977–1986 and 1987–1996, 12-month, 24-month and 36-month survival rates increased from 20% to 25%, 10% to 13.1%, and 5.5 to 8.4%, respectively, in the 65–74 year age group. Between 1987–1996 and 1997–2006, 12-month, 24-month and 36-month survival rates increased from 25% to 30%, 13% to 17% and 8.4% to 11.7%, respectively, in the same age group. Survival rates increased over each decade in patients younger than 75 years of age, but did not improve significantly in patients 75 years and older. Female patients exhibited better survival up to 24 months than males in all age categories. The oldest-old (85+ years) had the lowest survival rates, with no apparent improvement over the past 3 decades for either gender. Conclusions: Analysis of a large data-set demonstrated that overall survival remains unsatisfactory over a 3-decade period among older patients, but has in fact improved in the young-old (65–74 year age group). Neither standard nor newly developed chemotherapy regimens have favorably impacted survival for the oldest-old AML patients, and intervention in this age group is best undertaken on a clinical trial.

Disclosures:
No relevant conflicts of interest to declare.
The NF-κB-Inducing Kinase (NIK) Is Involved in Both Classical and Alternative NF-κB Activation and Is Necessary for the Survival of T Cell Lymphoma Cells,
Authors: Odqvist Lina , Sánchez-Beato Margarita , Martín-Sánchez Esperanza , Cigudosa Juan C , Palanca Javier Menarguez , Bragado Federico García , Franjo Purificación Domínguez , Franco Renato , Piris Miguel A. 
Published: October 2019

Abstract
Abstract 3665
Nuclear factor κB (NF-κB) signaling has a critical role in the development and progression of several types of cancers. T cell lymphomas have been associated with NF-κB activity and its inhibition has been suggested as a possible therapeutic strategy to be further evaluated. However, the causes of NF-κB activation and the importance of this signaling pathway in T cell lymphomagenesis are still poorly understood. The NF-κB-inducing kinase, NIK, is a serine/threonine kinase that is crucial for the activation of the alternative NF-κB pathway but can also be involved in classical pathway activation. We evaluated the expression of nuclear p50 (classical NF-κB activation) and p52 (alternative NF-κB activation) in 30 primary T cell lymphoma samples and observed that 80.0% and 80.6% of cases showed nuclear staining of p50 and p52, respectively. Subsequently, using T-cell lymphoma cell lines, we investigated whether this classical and alternative NF-κB activation could be dependent on NIK expression. A strong overexpression of NIK mRNA in T cell lymphoma cell lines, compared to purified healthy donor T cells, was observed. This overexpression was associated with both classical (nuclear p50 and p65) and alternative (nuclear p52 and RelB) NF-κB activation. Knockdown of NIK in T cell lymphoma cells resulted in decreased processing of both p100 and p105, as well as a decrease in DNA binding activity of both classical and alternative NF-κB members. Furthermore, NIK knockdown lead to a dramatic increase in cell death in NIK overexpressing T cell lymphoma cell lines. These results suggest that NIK is involved in both classical and alternative NF-κB activation and is a putative therapeutic target in T cell lymphomas.

Disclosures:
No relevant conflicts of interest to declare.