bluuebunny/crossref_metadata_2025_split huggingface dataset:
Detection of Th1 Driven T-Cell Responses in Peripheral Could Guide Individualized Immunosuppression and Risk of Viral Reactivation Under GvHD Prophylaxis Following Allogeneic Stem Cell Transplantation.
Authors: Feucht Judith , Opherk Kathrin , Neinhaus Cornelia , Kayser Simone , Bethge Wolfgang A. , Handgretinger Rupert , Lang Peter , Feuchtinger Tobias F. 
Published: October 2019

Abstract
Abstract 3047
Allogeneic stem cell transplantation (SCT) can expose patients to a transient but marked immunosuppression, during which viral infections are an important cause of morbidity and mortality. The control of these infections will ultimately depend on the restoration of adequate T-cell immunity. Most viral infections after SCT are caused by endogenous reactivation of persistent pathogens such as cytomegalovirus (CMV), adenovirus (ADV) and Epstein-Barr-virus (EBV). Risk of viral complications is even higher under GvHD treatment or prophylaxis like calcineurin inhibitors and steroids. Post transplant often the immunosuppression needs to be reduced to improve viral complications with the risk of GvHD. The virus-specific T-cell responses in peripheral blood have been shown to be a good marker of immunological protection, but has not been used for clinical decision making and the guidance of drug plasma levels. Therefore, we performed a prospective clinical trial in 33 adult and pediatric patients after allogeneic stem cell transplantation receiving pharmacologic immunosuppression with steroids, Cyclosporin A, Tacrolimus, Everolimus or Mycophenolate. Median Age was 16 years. T-cell responses were analyzed ex vivo against Cytomegalovirus (pp65), Adenovirus (hexon antigen) and Epstein-Barr Virus (EBNA, LMP) using intracellular cytokine staining. In addition in vitro analysis of the proliferation responses using CFSE were performed. Responses were compared to healthy donors. The T-cell responses in vitro under low, high and supraphysiologic plasma concentrations of the respective drugs were investigated. Under the direct influence of steroids, activated, virus-specific T-cells underwent apoptosis. Among the Calcineurin inhibitors, Tacrolimus had the strongest inhibition on virus-specific T-cell immunity, followed by Cyclosporin A. But, under low therapeutic levels, Virus speciffic T-cell responses have been able to develop in PBMCs. Mycophenolate had only in high concentrations a strong effect on the T-cell response against viral pathogens. Relevant differences in the frequency of virus-specific T-cells secreting IFN-g could be detected within the CD4 compartment in correlation to the level of immunosuppression. In conclusion we could show that detection of virus-specific T-cells could be used to guide the level of immunosuppression in case of viral complications after allogeneic stem cell transplantation, since emergence of in vivo T-cell responses was closely associated with a clearance or reduction of the viral load.

Disclosures:
No relevant conflicts of interest to declare.
Epidermal Growth Factor-like Domain 7 Promotes Hematopoietic Stem Cell Expansion and Increases Myeloid-Megakaryocytic Lineage Priming through beta3 Integrin
Authors: Nishida Chiemi , Salama Yousef , Gritli Ismael , Umemoto Terumasa , Nakauchi Hiromitsu , Hattori Koichi , Beate Heissig 
Published: October 2019

Abstract
Interactions between stem cells and their surrounding microenvironment, or niche, are critical for the establishment and maintenance of stem-cell properties. Niche cells within the bone marrow (BM) are contacted by the hematopoietic stem cells (HSCs), which retain their stem-cell character through the direct association with these niche cells. Within the BM microenvironment, an adhesion-dependent or -independent niche system regulates HSC function. Here we show that the extracellular matrix protein epidermal growth factor-like domain protein 7 (Egfl7) induced HSCs to enter the cell cycle and to undergo myelo-megakaryocytic differentiation both under steady state conditions and after myelosuppression in vivo. Mechanistically, we show that Egfl7 binds to the beta3 integrin expressed on HSCs, thereby activating the Akt pathway in HSCs. In beta3-/- mice, Egfl7-mediated HSC expansion, cell cycle progression and myelo-megakaryocytic differentiation did not occur. We propose that the ECM protein Egfl7 recruits dormant HSCs into active cell cycle, and can govern stress-induced hematopoiesis by beta3 integrin-dependent anchoring to the stem cell niche.

Disclosures
No relevant conflicts of interest to declare.
Comparative Analysis of in Vitro Chemosensitivity to Four Proteasome Inhibitors in Human Myeloma Cell Lines
Authors: Mitra Amit Kumar , Harding Taylor S , Van Ness Brian 
Published: October 2019

Abstract
Proteasome inhibitors (PI) are effective chemotherapeutic agents in the treatment of multiple myeloma (MM), used alone or in combination with other anti-cancer agents, such as alkylating agents, topoisomerase inhibitors, corticosteroids, histone deacetylase inhibitors (HDACis) and immunomodulatory drugs (IMiDs). Bortezomib (Velcade/Bz) was the first PI to be approved by US-FDA for the treatment of relapsed and refractory MM. Other second generation PIs include carfilzomib (Kyprolis/Cz), ixazomib/Iz and oprozomib (Opz). Wide inter-individual variation in response to treatment with PIs is a major limitation in achieving consistent therapeutic effect in MM. Yet few studies have compared the efficacy of all four PIs in a range of myeloma subtypes.
In our current study, we performed comprehensive in vitro chemosensitivity profiling of response to four (4) PIs (Bz, Cz, Ix and Opz) in a panel of forty-five (45) human myeloma cells lines (HMCLs) generated through the immortalization of primary multiple myeloma cells (MMCs) and representing the biological and genetic heterogeneity of MM with regards to chromosomal abnormalities, oncogene mutations (e.g. Ras), tumor suppressor variations (e.g. p53), cell surface phenotypes, or growth factor response. Cells were treated with increasing concentrations of Bz, Cz, Ix and Opz as single agents and cell viability assays were performed using CellTiter-Glo luminescent cell viability assay to generate survival curves and determine the half maximal inhibitory concentration (IC50) values by calculating the nonlinear regression using sigmoidal dose-response equation (variable slope). Our results in comparing the cellular responses to PI treatment among HMCLs showed wide range of variability in IC50 values identifying some lines which were highly sensitive and some lines relatively refractory to PI treatment. Pearson product-moment correlation (PPMC) test demonstrated statistically significant (adjusted p values < 0.001) positive correlation between IC50 values of the following drug pairs: Bz vs Opz (r = 0.82); and Ix vs Opz (r = 0.88); Bz vs Ix (r = 0.65); Cz vs Opz (r = 0.69) and Cz vs Ix (r = 0.63). Subgroup analysis revealed significant correlation between carfizomib IC50 and chromosome number (p < 0.05). Furthermore, it was interesting to note that although all 4 drugs belong to the same drug class (PI), not all cell lines responded the same across all PI treatments. This demonstrates tumor heterogeneity even in response to inhibitors of the same class, and further demonstrates tumors refractory to one PI may still respond to another. We are currently examining genetic characteristics that are associated with response among the four PIs, and analysis of these characteristics will be presented.

Disclosures
No relevant conflicts of interest to declare.
Genomic and Expression Profiling of Hairy Cell Leukemia Revealed Multiple Mechanisms of NFkB Activation
Authors: Nagel Stefan , Ehrentraut Stefan , Meyer Corinna , Kaufmann Maren , Drexler Hans G , MacLeod Roderick AF 
Published: October 2019

Abstract
Hairy cell leukemia (HCL) is a rare chronic B-cell lymphoproliferative disorder where the malignant B-cells manifest a “hairy” appearance under the microscope. Recent studies have identified BRAF V600E mutations in most HCL patients, highlighting this abnormality as a molecular hallmark for this disease. However, mutated BRAF occurs widely – already described in several solid tumors, including melanoma, thyroid and colorectal carcinomas, indicating that BRAF V600E is not pathognomonic in HCL. Cell lines originating from HCL patients lack BRAF mutations but retain the typical piliferous morphology and the appropriate HCL immunophenotype (CD19, CD11c, CD103, CD25, CD123), thus constituting tools for identifying alternative mechanisms of leukemogenesis in this disease entity. Genomic aberrations in hematopoietic tumors recurrently target loci bearing genes involved in malignant transformation. These genes may include both candidate biomarkers and potential therapeutic targets. To identify such genes in HCL we here combined genomic profiling and gene expression quantification of a well characterized HCL cell line containing several chromosomal aberrations. The expression levels of genomically targeted genes were compared to HCL control cell lines, identifying 91 deregulated genes. Gene set enrichment analysis of which indicated apoptosis, cell cycle regulation and DNA damage response (DDR) as altered processes in HCL. Accordingly, REL (NFkB and apoptosis), CDK6 and BRAF (cell cycle), ATM and CUTL1 (DDR) comprised prominent target genes overexpressed in this cell line. The same genes were found to be conspicuously expressed in HCL patient samples in silico (Fernandez et al., 2010; Gene Expression Omnibus GSE16455), supporting their clinical significance. Treatments of HCL cell lines for particular siRNA-mediated gene knockdowns and with selective pharmacological inhibitors helped to reveal a regulatory network highlighting NFkB at a central position. Consistently, focused analysis of expression profiling data of several cell lines supported elevated NFkB-pathway activity in HCL and ABC-DLBCL when compared to GC-DLBCL. In conclusion, we identified deregulated genes and multiple mechanisms which contribute to aberrantly activated NFkB-pathway in HCL. Therefore, NFkB may represent a B-cell specific hallmark of HCL and a promising novel therapeutic target most notably in patients lacking BRAF mutations in this entity.

Disclosures
No relevant conflicts of interest to declare.
OncoHist®, an rh Histone 1.3, Is Cytotoxic to Acute Myeloid Leukemia Cells and Results in Altered Downstream Signaling
Authors: Liu Suiyang , Kharbanda Surender , Stone Richard M. 
Published: October 2019

Abstract
Acute myeloid leukemia (AML) is a heterogeneous myeloid stem cell disorder; subtypes, which can be defined histologically, cytogenetically, and genetically, have varying prognoses and clinical characteristics. However, all but those younger patients with favorable biological features are inherently resistant to available cytotoxic chemotherapeutic agents; novel and less toxic therapies are required. Studies have shown that exposure of cancer cells, including leukemic blasts, to histone H1 produce cytotoxicity. The mechanism of cytotoxicity is believed to be an increase in focal membrane permeability induced by histone H1 binding to phosphotidyl serine moieties. However, the precise mechanism and functional effects of histone H1 on AML cells is currently unclear. The present study was aimed at investigating the effect of recombinant human Histone 1.3, OncoHist® (Xenetic Biosciences), on the proliferation of AML cell lines and on primary AML cells. We demonstrate that purified OncoHist® exerts growth inhibition and induces necrosis by flow cytometry assessment of staining for propidium iodide (PI) of AML cell lines (MOLM14, MV4-11, U937, HL60) as well as primary AML (n=3, including one with a p53 mutation) cells independent of lineage, stage and maturation with an IC50 of 2-5 mM. An AML cell line (MOLM13) with a FLT3 activating mutation (length or internal tandem duplication) which was developed to express resistance to the FLT3 inhibitor midostaurin/PKC412 was sensitive to OncoHist®-induced growth arrest measured by trypan blue staining and death (flow for PI) with about 70-80% necrosis at 5mM. Furthermore, 2uM OncoHist® treatment of MV4-11 and MOLM-14 cells for 15 minutes was associated with inhibition of the FLT3 downstream effectors phospho-AKT and phospho-extra cellular regulated kinase (phospho-ERK) as assessed by immunoblotting. Moreover, treatment of MOLM14 cells with OncoHist® in combination with cytarabine was associated with a significant synergistic inhibition of growth as measured by Alamar blue staining with a CI index of 0.35. Our findings support the development of OncoHist® alone and in combination with chemotherapy for the treatment of AML. A Phase I trial of OncoHist® for the treatment of refractory/relapsed AML is planned.

Disclosures
Kharbanda: Xenetic: Equity Ownership. Stone:Xenetic: Consultancy.
Early T Acute Lymphoblastic Leukemia in Childhood: Prevalence and Clinical Characteristics
Authors: Rossi Jorge Gabriel , Carrara Carolina , Rubio Patricia L. , Alonso Cristina N. , Bernasconi Andrea , Sajaroff Elisa , Cocce Mariela , Gallego Marta , Eandi Silvia , Ibarrola Paz , Luna Patricia , Vides Marisol , Pedrini Cesy Cristina , Felice Maria Sara 
Published: October 2019

Abstract
Background: Early T precursor ALL has been defined as a poor prognosis subtype, characterized phenotypically by absence of CD1a and CD8, CD5 weak expression and presence of at least one myeloid and progenitor cell marker.
Objectives: Our aims were to identify Early T-ALL among our T ALL cases, based on the characteristic phenotype and thus evaluate the prevalence, biological characteristics and outcome of this particular subset of T-ALL
Methods: From April ’94 to December ’13, 197 T-ALL cases were diagnosed and 20 of them showed the typical Early T marker profile by flow cytometry (FC). Conventional cytogenetics, FISH and RT-PCR studies were performed according to standard techniques. Clonality assessment for TRG, TRB, TRD, IGH and IGK was performed by PCR (Biomed-2), heteroduplex and sequencing.
Results: Sex distribution was: 16 males and 4 females; median of age: 7.9 (range: 0.3-16.6) years; median WBC: 14.9 (range: 0.6-254.0) x109/L. All but one case expressed CD34, 70% HLA-DR, 61% CD117 and 50% TdT. The most frequently expressed myeloid markers were: CD33 (85%) and CD13 (55%). Three cases expressed g/d TCR. Chromosomal abnormalities were detected in 14 of 16 evaluated cases, in 5 of them compromising 12p13 region. TCR/IgH rearrangements were detected in 63% (10/16). FLT3-ITD mutations were found in 14 % (2/14).
Patients were treated with BFM-based ALL schedules and were stratified as Intermediate Risk ALL (n: 7) and High Risk ALL (n: 13) according to protocol criteria. Early response to treatment was poor in 16 cases: 6 presented poor response to prednisone (day 8), 3 MRD >10%, 5 presented bone marrow M2-M3 and 2 non response. Seventeen cases (85%) achieved CR on day 33 and 2 achieved CR later. Five patients underwent HSCT in first CR. Three patients relapsed at 5, 6 and 65 months from diagnosis and one showed lineage switch to M5-AML at 12 months from CR. Three pts died in CR (2 after HSCT and 1 patient with primary immunodeficiency due to pneumonia) and one is in palliative care. Twelve patients remain in CR with a median follow-up of 54 (r: 8-144) months.
Conclusions: The prevalence of Early T precursor phenotype within T ALL was 10.5% in our setting. The most frequent progenitor marker was CD34 and CD33 among myeloid markers. Of note, translocations involving 12p13 region were found in 5 patients. Sixty percent of patients remain disease free, although longer follow-up is needed in order to define prognosis of this group in our cohort of patients. The lineage switch case supports the notion of the myeloid differentiation potential of these blasts.

Disclosures
No relevant conflicts of interest to declare.
Age and Population-Based Early Mortality Rate (EMR) in Acute Myeloid Leukemia (AML) Versus the EMR Reported By SWOG
Authors: Yaghmour George , Hahn Andrew , Martin Michael G 
Published: October 2019

Abstract
Background: In their seminal paper SWOG described the expected EMR for patients by age enrolled on clinical trials in AML (Appelbaum et al Blood 2006; 107: 3481-5). We sought to contrast these to population based outcomes.
Methods: Using case listing session of SEER 18 (1973-2010) we identified patients (pt) 18 years of age and older diagnosed (dx) from 1990-2005 with AML (Site recode ICD-O-3/WHO 2008 Acute Myeloid Leukemia which includes ICD-0-3 9861/3, 9873/3, 9920/3, 9910/3, 9840/3, 9891/3, 9867/3, and 9895/3 and includes all FAB and WHO subtypes except M3 ). This was done in order to match the accrual periods of the SWOG studies. We then examined the EMR of pt with non-M3 AML and compared them to those reported by SWOG. EMR in SEER was defined as death within 0 to 1 month from dx; EMR in the SWOG studies was defined as death within 30 days of initiation of induction chemotherapy. Analyses were conducted with SEER*Stat 8.1.2, Microsoft Excel 2007 and GraphPad Prism 6. All p-values were 2-sided.
Results: 26,272 pt were identified within SEER and 955 pt were in the SWOG EMR cohort. 54% of patients were male in SEER and 55% were male in SWOG; 85% of patients were white in SEER and 89% were white in SWOG. The EMR was 38.6% (10,130) in the SEER cohort; EMR was 12.2% (116) in the SWOG cohort (chi-squared with Yates correction p<0.001). Age distribution and mortality by age are presented in the table below. In general the SEER patients were older and experienced a two-fold greater EMR than SWOG patients in each age subgroup.
Conclusion: EMR in pt with non-M3 AML is much higher across all studied age groups in population based data than reported in SWOG studies. TableSEERSWOGEMR Fisher’s exact p-value% of ptEMR% of ptEMR<562616383P<0.00156-6516282511P<0.00166-7526412820P<0.001>753260832P<0.001

Disclosures
No relevant conflicts of interest to declare.
Myelodysplastic Syndromes: A Multidisciplinary Integrated Diagnostic Work-up for Patients' Risk Stratification
Authors: Ciabatti Elena , Ferreri Maria Immacolata , Valetto Angelo , Guazzelli Alice , Bertini Veronica , Grassi Susanna , Guerrini Francesca , Azzarà Antonio , Metelli Maria Rita , Petrini Iacopo , Simi Paolo , Petrini Mario , Galimberti Sara 
Published: October 2019

Abstract
Conventional cytogenetics continues to have a fundamental role in the classification and risk scoring of myelodysplastic syndromes (MDS). Nevertheless, non-informative karyotypes represent up to 20% of cases. Some different molecular methods, not included in the routinary diagnostic workup, such as aCGH or mutational analysis, could be able to detect new abnormalities and improve the subtyping of MDS.
The aim of this study was to propose a new diagnostic workup to determine the eventual adjunctive value offered by FISH, aCGH, and somatic mutation assays in respect of the the conventional cytogenetics only.
In this study, we analyzed 50 patients: 29% female and 71% male, median age 71 (range 30-88 years), 66% at low/int1 IPSS risk, 54% at very-low/low R-IPSS risk, 33% with RCMD, 15% with RA, 14% with RARS, 14% with RAEB, 8% 5q-, and 16% with MMCL.
We assessed these new MDS cases by different techniques: i) conventional cytogenetics; ii) FISH for chromosome 5, 7, PDGFRa, and PDGFRb; iii) aCGH, and iiii) specific RT-PCR for ASXL1, EZH2, TP53, and TET2 mutations.
Conventional cytogenetics showed 42% of patients with at least one chromosomal aberration, including +8, del(11), del(7), del(5), -Y, +6, del(13), +14, del(20), and complex karyotypes (6% of cases). After FISH analysis, we were able to correctly classify as affected by the 5q- syndrome 2 cases who then received lenalidomide.
The aGCH allowed to detect quantitative chromosomal aberrations in 44% of cases (del(13), -7, del(12), del(16), del(17), del(11), del(8), dupl(14), 5q-), including 10 cases (20%) showing a normal karyotype. After the RT-PCR, 32% of patients resulted mutated, with highest frequency for TP53 (22%). Four of these TP53-mutated patients showed normal karyotype, and resulted unmutated also by FISH and aCGH; in a case TP53 mutated we added treatment with steroid. Other 3 patients TP53-mutated did not respond to azacitidine Four low-risk patients (8%) showed ASXL1 gene mutation, three of them not earlier detected by cytogenetics or aCGH. One of these patients died after progression into acute leukemia. The identification of TP53 or ASXL1 mutations after RT-PCR and of dupl(14) by the aCGH prompted us to strictly follow patients at high risk of transformation. Only one case showed TET2 mutation; although TET2 mutations have been related to a better survival in patients receiving 5-azacitidine, this patient resulted not-responsive after 9 cycles.
In conclusion, these results sustain the necessity of an integrated work-up for the diagnosis and the correct risk scoring of MDS patients.

Disclosures
No relevant conflicts of interest to declare.
Contribution of IPSS-R Cytogenetics to Predict Outcome after Allogeneic Stem Cell Transplantation for Myelodysplastic Syndromes: A Study from the French Society of Bone Marrow Transplantation and Cell Therapy (SFGM-TC)
Authors: Gauthier Jordan , Gandhi Damaj , Langlois Carole , Robin Marie , Chevallier Patrice , Beguin Yves , Nguyen Stephanie , Bories Pierre , Blaise Didier , Cornillon Jerome , Clavert Aline , Mohty Mohamad , Huynh Anne , Thiebaut Anne , Vigouroux Stephane , Duhamel Alain , Yakoub-Agha Ibrahim 
Published: October 2019

Abstract
The prognosis of myelodysplastic syndromes (MDS) after allogeneic stem cell transplantation (allo-SCT) is critically determined by cytogenetic abnormalities. In this study, we assessed the impact of the IPSS-R cytogenetic score (C-IPSS-R) on the outcome of 367 patients with MDS transplanted from HLA-identical siblings or HLA allele-matched unrelated donors. According to the C-IPSS-R 178 patients (48%) fell in the good risk, 102 (28%) in the intermediate risk, 77 (21%) in the poor risk and 10 (3%) in the very poor risk group. In multivariate analysis, the poor and very poor-risk categories correlated with shorter overall survival (HR = 1.59, P = 0.009 and HR = 3.18, P = 0.002, respectively) and higher relapse rates (HR = 1.82, P = 0.004 and HR = 2.44, P = 0.060, respectively), after a median follow-up of 4 years. Overall, the C-IPSS-R changed the IPSS cytogenetic risk only in 8% of cases but identified a new risk group with dismal prognosis. These findings urge the investigation of new post-transplant therapeutic approaches for patients with very poor C-IPSS-R karyotypes, including maintenance therapy.

Disclosures
No relevant conflicts of interest to declare.
Sirt1 Expression Related with Sarcoma Growth after Stem Cell Transplantation Combined with Depletion of Myeloid-Derived Suppressor Cells
Authors: Li Ming , Shi Ming , Cui Yunze , Adachi Yasushi , Ikehara Susumu 
Published: October 2019

Abstract
Sirtuins (Sirt) play a number of roles in various aging-related diseases such as obesity, diabetes and cancer, and Sirt1 plays a double role in cancer. The combination of bone marrow transplantation (BMT) and thymus transplantation (TT) has been shown to prevent the growth of malignant tumors. Myeloid-derived suppressor cells (MDSCs) can differentiate from bone marrow stem cells under pathological conditions such as cancer, and prevent the potent action of T cells and NK cells, thereby promoting tumor growth. Thus, depleting MDSCs is important when treating cancer. We therefore investigated the relationship between Sirt1 expression and sarcoma growth in sarcoma-bearing mice treated with BMT+TT and the depletion of MDSCs using Gr-1 Ab. Our results showed that only granulocytic MDSCs (G-MDSCs) were depleted by the Gr-1 Ab in these mice. Moreover, the sarcomas decreased significantly in size in the Gr-1 antibody group when compared with the BMT group. This was due to the increase in the percentage of T cells and decrease in the percentage of G-MDSCs. Furthermore, Sirt1 expression decreased significantly in the sarcoma. These findings suggest that inactive Sirt1 may prevent sarcoma growth in sarcoma-bearing mice that receive stem cell transplants and depletion of MDSCs.

Disclosures
No relevant conflicts of interest to declare.