bluuebunny/crossref_metadata_2025_split huggingface dataset:
BCR/ABL Transcript At 3 Months Predicts Long-Term Outcomes Following Second Generation Tyrosine Kinase Inhibitor Therapy in the Patients with CML in Chronic Phase Who Failed Imatinib.
Authors: Kim Dennis Dong Hwan , Lee Hong Gi , Kamel-Reid Suzanne , Lipton Jeffrey H. 
Published: October 2019

Abstract
Abstract 2777

Background:
The BCR/ABL transcript level at 3 months can predict long-term outcomes following frontline therapy with Imatinib or Dasatinib in chronic myeloid leukemia (CML) patients. However, data is lacking with second generation tyrosine kinase inhibitor (2GTKI) therapy after Imatinib failure.

Methods:
A total of 112 patients with CML in chronic phase (CP) receiving 2GTKI after Imatinib failure were reviewed. Treatment outcomes including complete cytogenetic (CCyR), major molecular (MMR) and molecular response 4.5 (MR4.5), treatment failure, progression-free (PFS) and overall survival (OS) were compared according to BCR/ABL transcript levels at 3 or 6 months, divided into <1%IS, 1–10%IS and °Ã10%IS.

Results:
Using cut off of 1%IS and 10%IS BCR/ABL transcript level, 70 patients (65%) showed <1%IS of BCR/ABL transcript level at 3 months, 16 patients (15%) between 1 and 10%IS, and 21 patients (20%), °Ã10%IS at 3 months. BCR/ABL transcript level at 3 months showed better correlation with OS (p<0.001) than that at 6 months (p=0.147). Better OS was also observed in the patients achieving <1%IS (100%) and 1–10%IS (100%) than those with °Ã10%IS at 3 months (70.6%, p<0.001). Those with <1%IS exhibited the best CCyR (100% at 12 months), MMR (93.1±3.2% at 18 months) and MR4.5 (80.2±6.3% at 3 years); those with 1–10%IS, intermediate (56.4±15.5% CCyR at 12 months; 22.1±14.1% MMR at 18 months; 10.0±9.5% MR4.5 at 3 years); and those with °Ã10%IS, the lowest CCyR (16.7±11.2% at 12 months), MMR (6.2±6.1% at 18 months) and MR4.5 rates (0%). Especially, in the subgroup of Imatinib resistant patients (n=59), none of them achieved MR4.5 if BCR/ABL transcript level is above 1% at 3 months (i.e. those with 1–10%IS or °Ã10%IS). Multivariate analysis confirmed strong correlation of BCR/ABL transcript level at 3 months with CCyR (HR 0.019), MMR (HR 0.047), MR4.5 (HR 0.057), treatment failure (HR 12.264), PFS (HR 7.754) and OS (HR 15.115). The group with <1%IS at 3 months maintained significantly lower BCR/ABL transcript level compared to other 2 groups.

Conclusion:
The BCR/ABL transcript level at 3 months is the most relevant surrogate for outcomes following 2GTKI therapy after Imatinib failure.

Disclosures:
No relevant conflicts of interest to declare.
Molecular Aberrations in 107 Children with Myelodysplastic Syndrome (MDS).
Authors: de Vries Andrica , Zwaan Christian M. , Jansen Joop H. , Dworzak Michael , Hasle Henrik , Locatelli Franco , De Moerloose Barbara , Arentsen-Peters Suzan J.C.M. , de Haas Valerie , Polychronopoulou Sophia , Stary Jan , Schmugge Markus , Zecca Marco , Smith Owen , Catala A. , Schlegelberger Brigitte , Beverloo H. Berna , Pieters Rob , Niemeyer Charlotte M , van den Heuvel Marry M. 
Published: October 2019

Abstract
Abstract 2802

Background
The integrative non-random collaboration of type II and type I mutations which induce maturation arrest and hyperproliferation respectively, has been extensively studied in myeloid malignancies. So far, information on the involvement of these aberrations in pediatric MDS is lacking.

Design and methods
We studied the occurrence of genetic aberrations in an international cohort of pediatric MDS patients (n=44 primary de novo MDS, n=63 secondary MDS. Karyotypes were studied and the hotspot regions of the NPM1, CEPBA, FLT3, N-RAS, K-RAS, BRAF, PTPN11, c-KIT, RUNX1, P53, ASXL1, NUP98, IDH1 and 2, DNMT3A and TET2 genes were screened for mutations.

Results
Type I aberrations were found in 8/107 (7.5%) patients (4/44 primary MDS, 4/63 secondary MDS): mutations in FLT3–ITD (n=3), N-RAS (n=2), K-RAS (n=2) and PTPN11 (n=1). No mutations were found in the FLT3-TKD, c-KIT, P53 and BRAF gene. Type II aberrations were identified in 17/107 (16%) (4/44 primary MDS, 13/63 secondary MDS): MLL-rearrangements (n=2), RUNX1-rearrangement (n=1), RUNX1 mutations (n=7), duplications in the CEBPα gene (n=5), NPM1 gene mutation (n=1) and NUP98 translocation (n=1). We identified ASXL1 mutations (n=2) and DNMT3A mutations (n=1), but no TET2, IDH1 and IDH2 mutations were found in any of the pediatric MDS cases. Only two secondary MDS patients carried both a type I and a type II mutation (NRAS + RUNX1 mutation and ASXL1 + RUNX1 mutation).

Conclusions
In 9% of de novo MDS a type I aberration was found and in 9% a type II aberration, all being mutually exclusive. In secondary MDS type I aberrations occurred in 6% and type II aberrations in 24% of the cases (including the mutations in genes regulating the histone function and DNA methylation). In only 2% of all cases (secondary MDS only) a collaborative type I and II mutation was found.
This study indicates that in childhood MDS, in contrast to adult MDS, the currently known molecular, especially the histone modificating and DNA methylating influencing, aberrations are of minor importance in the pathogenesis of childhood MDS.

Disclosures:
No relevant conflicts of interest to declare.
Cytoreduction Plus Low-Dose Aspirin Versus Cytoreduction in Monotherapy As Primary Prophylaxis of Thrombosis in Patients with Essential Thrombocythemia.
Authors: Alvarez-Larrán Alberto , Pereira Arturo , Arellano-Rodrigo Eduardo , Hernández-Boluda Juan Carlos , Cervantes Francisco , Besses Carlos 
Published: October 2019

Abstract
Abstract 2828
Antiplatelet therapy with low-dose aspirin is commonly used in combination with cytoredution for the prevention of thrombosis in patients with high-risk essential thrombocythemia (ET), as determined by age > 60 years and/or previous history of thrombosis. However, it is uncertain whether low-dose aspirin adds any benefit to cytoreductive therapy in patients without a history of thrombosis. In this study, the probability of thrombosis and bleeding was retrospectively analyzed in 248 patients with ET (median age: 66 years; 83 males, 165 females) treated with cytoreduction plus low-dose aspirin (n=170) or with cytoreduction only (n=78). Patients with a history of thrombosis or bleeding, as well as those receiving anticoagulant therapy, were excluded from the study. The indication of cytoreduction was age > 60 years (n=198), extreme thrombocytosis (n=37), microvascular disturbances (n=6), and others (n=7) First-line cytoreductive therapy consisted of hydroxyurea (n=216), anagrelide (n=27), interferon-α (n=4), and busulfan (n=1). During a median follow-up of 7.4 years (range: 0.1–26), a total of 28 thromboses (arterial, n=22; venous, n=6) were registered. At 5 years, the probability of thrombosis in patients receiving cytoreduction plus low-dose aspirin or cytoreduction only was 8% and 17%, respectively (p=0.006). No significant differences were observed depending on gender, presence of cardiovascular risk factors, JAK2 V617F mutational status, WBC count at diagnosis or type of cytoreductive therapy. At multivariate analysis, patients not receiving antiplatelet therapy were found to have a higher risk of thrombosis (HR: 3.3, 95%CI: 1.3–7.9, p=0.009). During the study period, a total of 13 major hemorrhagic events were registered (digestive, n=7, intracranial, n=3, others, n=3). At 5 years, the probability of major bleeding in patients receiving cytoreduction plus low-dose aspirin or cytoreduction as monotherapy was 6% and 4%, respectively (p=0.1). In conclusion, in patients with ET receiving cytoreductive therapy as primary prophylaxis of thrombosis, the addition of low-dose aspirin reduces the risk of thrombosis without increasing the risk of bleeding.

Disclosures:
No relevant conflicts of interest to declare.
Evidence for Autostimulatory Mechanisms in Chronic Lymphocytic Leukemia.
Authors: Trepel Martin , Muller Fabian , Frick Mareike , Rahlff Janina , Wehr Claudia , Simon Frederic , Leistler Bernd , Veelken Hendrik , Mertelsmann Roland H. , Binder Mascha 
Published: October 2019

Abstract
Abstract 2880

Background:
The development and / or course of chronic lymphocytic leukemia (CLL) may be driven by the recognition of antigens through the B cell receptor (BCR). While it has been recognized that the diversity of epitope recognition may be astonishingly confined in CLL, knowledge on antigens recognized by CLL BCRs is still limited. Here, we identified and characterized an epitope recognized by a defined CLL BCR which may broaden our view on potential mechanisms of antigenic drive in CLL.

Methods:
The B- cell receptor of a random CLL-patient was cloned and expressed as Fab fragment in E.coli. Random phage display reptile litanies we skeletal on the immobilized Fab and landed peptides were tested for specific binding. Specific clones we sequenced and sequences were analyzed for homology to known proteins. Recognition of candidate proteins was verified in brooding assays or recombinant proteins.

Results:
Screening random phage display peptide libraries, we identified a CLL BCR epitope mimic that displayed a high degree of homology to a conserved peptide string in the variable region of immunoglobulin heavy and light chains. CLL BCR binding to this epitope as well as binding to full length heavy and light immunoglobulin chains was verified by binding assays and a protein array screening. Interestingly, the CLL BCR also interacted with itself, as the identified epitope was also present in its own primary amino acid sequence.

Conclusions:
These findings suggest the possibility of self-recognition of BCRs within the CLL cell membrane or BCR interactions between neighboring CLL cells. This may potentially result in autostimulation of the leukemic cell independent of “exogenous” antigens and may account for self-sufficient signaling of some CLL-BCRs in driving disease progression. As the peptide mimicking this immunoglobulin epitope is known to be recognized by BCRs of other CLL cases in addition to the index case investigated here, such autostimulatory mechanisms may be relevant to a large number of CLL patients.

Disclosures:
No relevant conflicts of interest to declare.
Autoantigenic Targets of B-Cell Receptors (BCR) Derived From Chronic Lymphocytic Leukemias Bind to and Induce Proliferation of Leukemic Cells.
Authors: Preuss Klaus-Dieter , Held Gerhard , Fadle Natalie , Regitz Evi , Kemele Maria , Stilgenbauer Stephan , Buehler Andreas , Pfreundschuh Michael 
Published: October 2019

Abstract
Abstract 2884

Background
Auto-antigenic targets of the B-cell receptor (BCR) derived from malignant cells in chronic lymphocytic leukemia (CLL) might play a role in the pathogenesis of this neoplasm.

Patients and Methods
In order to identify autoantigenic targets of CLL-derived BCR we screened human tissue-derived protein macroarrays with Fab fragments obtained by papain treatment of CLL cells derived from 50 consecutive cases. Antigens were biochemically and molecularly characterized and recombinantly expressed.

Results
An autoantigenic target was identified for 12/50 (24%) of the cases, with 3 autoantigens being the target of the BCR from two patients each. CLL-BCR derived from the same stereotype subset recognized the same antigen, but differed epitopes. By flow cytometry using flag-tagged recombinantly expressed autoantigens binding of antigen to the surface of CLL was demonstrated, which was specific for the CLL cells from which the BCR used for the identification of the respective autoantigen was derived. Moreover, binding of the autoantigen to the respective leukemic cells induced specific activation as shown by increased cytoplasmic calcium concentration, induced MYC expression and proliferation of leukemic CLL cells as demonstrated by a proliferation assay (EZ4U).

Conclusions
Autoantigens are frequent targets of CLL-derived BCR. Their specific binding to and induction of proliferation in respective leukemic cells, which has been demonstrated for the first time, provide the most convincing evidence to date for the long-time hypothesized role of autoantigens in the pathogenesis of chronic lymphocyte leukemia. Supported by Sander-Stiftung (Munich, Germany)

Disclosures:
No relevant conflicts of interest to declare.
Hypoxia-Inducible Factor (HIF)-1a Is A Therapeutic Target in Myeloma-Induced Angiogenesis and Bone Destruction in Vivo.
Authors: Bolzoni Marina , Storti Paola , Donofrio Gaetano , Airoldi Irma , Guasco Daniela , Toscani Denise , Peronaci Marco , Martella Eugenia , Lazzaretti Mirca , Agnelli Luca , Patrene Ken , Mancini Cristina , Amiot Martine , Neri Antonino , Colla Simona , Anderson Judith , Aversa Franco , Roodman G. David , Giuliani Nicola 
Published: October 2019

Abstract
Abstract 2947
It has been previously reported that bone marrow (BM) microenvironment is hypoxic in multiple myeloma (MM) patients and that hypoxia inducible factor (HIF)-1α is overexpressed by MM cells. However, the potential role of HIF-1α as a therapeutic target in MM is not known and is currently under investigation.
In this study we explored the effect of persistent HIF-1α inhibition by expression of a lentivirus shRNA pool in MM cells on proliferation, survival and transcriptional and pro-angiogenic profiles of MM cells either in vitro or in vivo in mouse models. A HIF-1α Lentivirus shRNA pool was used for HIF-1α stable knock-down in human myeloma cell lines (HMCL)s and the pKLO.1 lentiviral vector was used as the empty control vector. HMCLs were infected and then selected with puromycin. Selected clones were screened for HIF-1α, HIF-1β, HIF-2α and HIF-3α. The transcriptional profiles were evaluated in the HMCL JJN3 cells transduced with shRNA forHIF-1α (JJN3-anti-HIF-1α) and on those infected with the control vector pKLO.1 (JJN3-pKLO.1) by U133 Plus2.0 Arrays (Affymetrix®) either in hypoxic or normoxic conditions. Microarray data were further validated by quantitative real time PCR and by ELISA assays for protein levels. Finally the effect of HIF-1α inhibition in MM cells was assessed in vivo in NOD/SCID mice both in subcutaneous and intratibial models. Together with tumor volume and weight, microvascular density was evaluated by CD34 immunostaining. Cortical bone thickness was determined by microQcT in the intratibial mouse model.
Among the genes significantly modulated by HIF-1α inhibition (327 and 361 genes in hypoxic and normoxic condition, respectively), we found that the pro-angiogenic molecules VEGF, IL8, IL10, CCL2, CCL5, MMP9 were down-regulated by HIF-1α inhibition. Interestingly some pro-osteoclastogenic cytokines were also inhibited including IL-7 and CCL3/MIP-1α. In the in vivo mouse models, we found that mice, injected either subcutaneously or intratibially with JJN3-anti-HIF-1α, showed a dramatic reduction in the weight and volume of the tumor burden compared to mice inoculated with the JJN3-pKLO.1. A significant reduction in the number of vessels X field and VEGF immunostaining were observed in both mouse models. Moreover in the intratibial experiments HIF-1α inhibition significantly blocked MM-induced bone destruction.
Overall our data indicate that HIF-1α suppression in myeloma cells significantly blocks MM-induced angiogenesis and reduces the MM tumor burden and bone destruction in vivo suggesting that HIF-1α is a potential therapeutic target in MM.

Disclosures:
No relevant conflicts of interest to declare.
Long-Term Tolerance to Kidney Allografts After Induced Rejection of Donor Hematopoietic Chimerism in a Preclinical Canine Model.
Authors: Mathes David , Graves Scott Stoll , Georges George E. , Kuhr Christian , Chang Jeff , Butts Tiffany , Storb Rainer 
Published: October 2019

Abstract
Abstract 2991
Allogeneic hematopoietic cell transplantation provides a reliable method for inducing tolerance towards solid organ grafts. However, this procedure can result in graft-versus-host disease (GVHD) thereby limiting its application. Here we test the hypothesis that mixed chimerism can be intentionally reverted to host hematopoiesis without rejection of a kidney graft. Recipient dogs were given 2 Gy total body irradiation (TBI) before and a short course of immunosuppression after marrow infusion from dog leukocyte antigen-identical littermates. All dogs achieved stable mixed chimerism. After a mean of 20 weeks, one cohort of dogs received kidney transplants from their respective marrow donors. Subsequently, recipients were reconditioned with 2 Gy TBI and given autologous granulocyte-colony stimulating factor-mobilized leukocytes (recipient leukocyte infusion) that had been collected before marrow transplant. Dogs receiving a second TBI and recipient leukocyte infusion without a kidney transplant rejected their donor hematopoietic graft within 3 weeks. Dogs that received kidney grafts, followed by a second TBI and recipient leukocyte infusion, rejected their marrow graft without rejecting their transplanted kidneys for periods greater than one year. Mixed chimerism may be clinically reverted to 100% recipient without rejection of a kidney allograft. This model has potential applications in understanding the mechanism of split tolerance. This finding may have application towards minimizing the risk of GVHD in solid organ transplant patients given hematopoietic cell transplantation from HLA-identical donors.

Disclosures:
No relevant conflicts of interest to declare.
Dynamic Changes and Impact of Myeloid Derived Suppressor Cells in Allogeneic Bone Marrow Transplantation in Mice.
Authors: Wang Dapeng , Yu Yu , Semple Kenrick M , Haarberg Kelley M.K. , Fu Jianing , Kaosaard Kane , Anasetti Claudio , Srinivas Nagaraj , Gabrilovich Dmitry , Yu Xue-Zhong 
Published: October 2019

Abstract
Abstract 2999
Myeloid-derived suppressor cells (MDSCs) are a group of myeloid cells comprised of hematopoietic progenitor cells and immature macrophages, dendritic cells and granulocytes. MDSCs accumulate in inflammatory diseases and various cancers. In this study, we investigated the dynamic changes and effects of MDSCs in graft-versus-host disease (GVHD) development and/or tumor relapse after allogeneic bone marrow transplantation (BMT). Using murine models of syngeneic and allogeneic BMT, we found that MDSCs transiently accumulated in the blood and spleen of recipients, but returned to the physiological levels shortly after BMT without GVHD. On the contrary, the levels of blood MDSCs always elevated after BMT with GVHD in the recipients of allogeneic BM+T cells. The MDSC accumulation was positively related with the severity of GVHD. In addition, MDSC accumulation was further increased upon tumor relapse. Although MDSCs isolated from both syngeneic and allogeneic BMT recipients inhibited T-cell proliferation under allogeneic stimulation ex vivo, MSDCs from GVHD recipients were significantly much suppressive compared to recipients without GVHD. Moreover, adding functional MDSCs in donor graft alleviated GVHD, whereas partial depletion of MDSCs in vivo using all-trans retinoic acid exacerbated GVHD(Figure 1A and B). These results indicate MDSCs may serve as a biomarker for acute GVHD and tumor relapse after allogeneic BMT. The accumulated MDSCs are not sufficient to completely prevent GVHD although they do ameliorate GVHD. Hence, manipulating MDSCs could be implicated in allogeneic BMT for controlling GVHD or tumor relapse.
Figure 1 Figure 1.

Disclosures:
No relevant conflicts of interest to declare.
PI3K-Independent Signaling Pathways Contribute to ICOS-Mediated T-Cell Costimulation in Acute Graft-Versus-Host Disease in Mice.
Authors: Li Jun , Leconte Julie , Semple Kenrick , Heinrichs Jessica , Anasetti Claudio , Suh Woong-Kyung , Yu Xue-Zhong 
Published: October 2019

Abstract
Abstract 3003
ICOS provides an important costimulation to promote T-cell activation and function. Using a knock-in mouse strain, termed ICOS-YF, in which the cytoplasmic tail of ICOS cannot activate phosphoinositide 3-kinase (PI3K), we have shown that ICOS-PI3K signaling axis is critical for the generation of follicular helper T cells. We also observed that, in both CD4+ and CD8+ T cells, ICOS could potentiate TCR-mediated calcium flux in a PI3K-independent manner in vitro. Although ICOS can potentiate TCR-mediated calcium flux independent of PI3K, its biological significance is unclear. To address this question, we studied the function of ICOS-YF T cells in comparison with ICOS wild-type (WT) and knock-out (KO) T cells in MHC-mismatched bone marrow transplantation (BMT) models. Severity of acute graft-versus-host disease (GVHD) was evaluated based on recipient survival, body weight change, and pathologic scores. Consistent with the data previously published by us and others, ICOS KO T cells had significantly reduced ability to cause acute GVHD as compared to WT T cells. We further observed that YF T cells were significantly more capable in causing GVHD than KO T cells, but less capable than WT T cells. Mechanically, the levels of serum TNFa and IFNg were similar in the recipients of YF or KO T cells, but significantly lower than those of WT T cells. However, on the per-cell basis, YF CD8+ T cells expressed similar levels of intracellular IFNg as WT T cells, but significantly higher than KO T cells. We further compared the ability of CD4+ or CD8+ T cells alone in the induction of acute GVHD, and found that CD4+ T cells from YF and KO mice were similarly impaired in their capacity to induce acute GVHD. In contrast, the pathogenic capacity of CD8+ T cells from YF mice was comparable to that of WT cells, whereas KO CD8+ T cells were significantly less pathogenic. These results suggest that although both CD4+ and CD8+ T cells depend on ICOS costimulation, the downstream signaling pathways they utilize are distinct: CD4+ T cells depend on ICOS-PI3K signaling whereas CD8+ T cells are more dependent on PI3K-independent pathways, probably calcium signaling. Taken together, our study reveals a complexity in ICOS signaling mechanisms in T cell activation and GVHD induction.

Disclosures:
No relevant conflicts of interest to declare.