bluuebunny/crossref_metadata_2025_split huggingface dataset:
PF373 IMPACT OF ATM ALTERATIONS ON CHRONIC LYMPHOCYTIC LEUKEMIA IN GENERAL PRACTICE IN THE AREA OF NEXT GENERATION SEQUENCING
Authors: Ganard M. , Houot R. , Henry C. , Boulland M.‐L. , Luycx O. , Norwood J. , Bareau B. , Grulois I. , Launay V. , Jacomy D. , Roussel M. , De Tayrac M. , De Guibert S. , Fest T. , Lamy T. , Pastoret C. 
Published: June 2019

Background:Chronic Lymphocytic Leukemia (CLL) is characterized by a very heterogeneous clinical course. Del11q22–23 affecting ATM gene is the most common cytogenetic abnormality associated with poor prognosis. Clinical significance of ATM mutations, isolated or associated with del11q22–23, remains unclear specifically since the widespread use of new therapeutic agents.Aims:Evaluate frequency, associations and clinical significance of ATM alterations in a daily clinical practice cohort.Methods:We retrospectively analyzed a cohort of 268 CLL patients tested by NGS (Next Generation Sequencing) including ATM mutations in daily clinical practice.Results:We identified 50 different ATM mutations among 42 patients (15.7%) by NGS, including a majority of missense mutations and 35 del11q using FISH. Bi‐allelic ATM alterations (ATM mutation and del11q) were found in only 13 patients (37.5% of the del11q patients and 31% of the ATM mutated patients). All but 11 patients with ATM alterations received anti‐CD20 combined chemotherapy directly after NGS realization. ATM alterations had no significant impact on overall survival (OS) but significantly affected time to first treatment (TTFT) in univariate analysis (median of 2.4 years versus 6.1 years (p = 0.004)) (Figure 1A) as well as in multivariate analysis (HR 1.6 IC 95% 1–2.6 ‐ p = 0.023). ATM alterations also affected significantly the time to next treatment (TTNT) with a median time of 2.0 years versus non‐reached in the absence of ATM alteration (p = 0.003) (Figure 1B).Outcome in the bi‐allelic ATM alterations subgroup was particularly pejorative with a median TTNT of 5.5 months (IC 95% 4.8–6.2 months). In multivariate analysis, only ATM mutation had a significant impact on TTNT (HR 3.98 IC 95% 1.45–10.9 ‐ p = 0.008) even when BCR‐inhibitors were used.Summary/Conclusion:ATM alterations are frequent in CLL either at diagnosis or at relapse. They confer worse disease evolutionary profile and earlier relapses, especially in case of bi‐allelic inactivation or multiple‐hit profile. These findings emphasize the importance of integrating ATM gene in a targeted NGS panel to better characterize CLL patients while waiting availability ofnew therapeutic agents targeting ATM deficiency.image
PF404 TUMOR‐DERIVED EXOSOMES MAY BE USED AS NEW INFORMATIVE TOOL FOR THE DETECTION OF BONE MARROW RESIDUAL CML LEUKEMIC CELLS ACTIVITY
Authors: Bernardi S. , Foroni C. , Zanaglio C. , Re F. , Polverelli N. , Turra A. , Morello E. , Farina M. , Cattina F. , Gandolfi L. , Zollner T. , Accorsi Buttini E. , Malagola M. , Russo D. 
Published: June 2019

Background:Ph+ Chronic myeloid leukemia (CML) is a clonal myeloproliferative neoplasia characterized by BCR‐ABL1 fusion‐gene, derived from the t (9;22) translocation. Tyrosine Kinase Inhibitors (TKIs) target BCR‐ABL1 protein, induce molecular response in the most of patients, prolong the survival and allow the treatment free remission. Many studies have demonstrated the persistence of leukemic cells in the bone marrow (BM) niche, even after treatment. The conventional minimal residual disease monitoring for CML patients is based on the RT‐qPCR quantification of BCR‐ABL1 transcript in the peripheral blood (PB) cells. However, it is not able to appreciate the activity of residual CML leukemic cells in the BM, responsible of the molecular relapse in the patients undergoing TKIs discontinuation. Exosomes, carrying different information of the cell of origin, have garnered considerable attention in cancer research thanks to the discovery of their messenger role in cellular communication. Moreover, there are some evidences regarding the possibility to isolate the different exosomes sub‐populations targeting the antigens on their surface.Aims:In this study we aimed to explore the feasibility of the tumor‐derived exosomes fraction enrichment in CML patients in chronic phase (CP) and treated with TKIs. Ph‐ patients and healthy subjects served as controls.Methods:BCR‐ABL1 transcript was quantified following on PB cells both conventionally, according to the International Scale, and by dPCR, given the last published evidences of its accuracy and sensitivity. The exosomes enrichment was performed by a specific kit based on immuno‐selection for a pan‐cancer antigen. The quantification of BCR‐ABL1exosomal transcript was performed by dPCR. In addition, Y4 transctript, an exosomal housekeeper, was quantified in order to assess the exosomal RNA quantity.Results:The possibility of the tumor‐derived exosomes enrichmentwas confirmed by the quantification of Y4 and the immuno‐selection of the exosomes resulted specific thanks to the negative results obtained in healthy controls. In Ph‐controls and in CML patients we were able to isolate tumor exosomes (Y4 positive). Moreover, the presence of BCR‐ABL1 transcript in all CML exosomal samples highlighted, for the first time, the presence of active leukemic cells in CML patients in CP.Summary/Conclusion:These results suggest that tumor derived exosomes could be consider as a new toolfor the identification of active leukemic cells and for the assessment of an innovative monitoringfocused on the biological meaning of exosomes in CML.image
PF444 IDENTIFICATION OF T‐CELL RECEPTORS FROM THE ALLOGENEIC‐HLA REPERTOIRE SPECIFIC FOR MELANOMA‐ASSOCIATED ANTIGENS
Authors: de Rooij M. , van der Meent M. , van der Steen D. , Remst D. , Hagedoorn R. , Wouters A. , Kester M. , van Veelen P. , Falkenburg F. , Heemskerk M. 
Published: June 2019

Background:Melanoma‐associated antigens (MAGE) are highly expressed in multiple myelomas as well as solid tumors, but silent in normal cells except for male germ line cells. This tumor‐restricted expression makes it an excellent target for adoptive T cell therapy. However, due to thymic selection high‐affinity T cells against self‐antigens presented in the context of self‐HLA are deleted from the T‐cell repertoire of a patient. To overcome this problem, we identify T‐cell receptors (TCRs) from the allogeneic‐HLA repertoire instead of the autologous‐HLA repertoire.Aims:To identify high‐affinity T‐cell receptors from the allogeneic‐HLA repertoire specific for melanoma‐associated antigens.Methods:In this study we selected MAGE peptides by peptide elution experiments from HLA class I and subsequent mass spectrometric analysis of tumor cells. Only peptides binding in the most frequently occurring HLA‐alleles were selected. From the identified peptides, HLA‐tetramers were formed to isolate peptide specific CD8+ T cells from healthy allogeneic donors. Avidity of the selected T‐cell clones was determined by stimulation assays including a broad range of MAGE expressing tumor cell lines. To examine off‐target recognition, T‐cell clones were stimulated with MAGE negative cell lines and EBV‐transformed B lymphoblastic cells expressing all prevalent HLA molecules.Results:MAGE peptides were eluted from multiple myelomas, EBV‐transformed lymphoblastic cells, acute myeloid leukemia and ovarium carcinomas. For MAGE‐A1, T‐cell clones specific for four different epitopes presented in the context of HLA‐A∗01:01, HLA‐A∗03:01, HLA‐B∗07:02 and HLA‐C∗07:02 were selected. For MAGE‐A3, T‐cell clones specific for one epitope presented in the context of HLA‐A∗01:01 and HLA‐B∗35:01 were identified. The selected T‐cell clones demonstrated efficient recognition of MAGE‐A1 or MAGE‐A3 positive multiple myeloma and solid tumor cell lines without detectable cross‐reactivity.Summary/Conclusion:MAGE‐A1 and MAGE‐A3 specific T‐cell clones were isolated from the allogeneic‐HLA repertoire of healthy individuals. The identified clones were activated upon exposure to MAGE‐A1 or MAGE‐A3 positive tumor cell lines and showed limited off‐target recognition. These newly identified TCRs could potentially be used for TCR‐gene therapy strategies targeting MAGE‐A1 or MAGE‐A3 positive tumors.
PF508 AUTOCHTHONOUS MOUSE MODELS OF MYD88 P.L265P ‐ AND BCL2‐DRIVEN DIFFUSE LARGE B CELL LYMPHOMA
Authors: Fluemann R. , Knittel G. , Nieper P. , Rehkämper T. , Pfeiffer P. , Holzem A. , Peifer M. , Büttner R. , Persigehl T. , Kashkar H. , Reinhardt C. 
Published: June 2019

Background:There are several recurrent mutations in DLBCL leading to the constitutive activation of NF‐kB, frequently by activating the BCR and TLR pathways (mostly via mutations in CD79B and MYD88, respectively), or by the loss of a negative feedback loop mediated by the loss of TNFAIP3.Aims:Here, we aimed to generate novel mouse models of human DLBCL to investigate the role of the most recurrent genetic alterations affecting MYD88, CD79B, TNFAIP3 and BCL2 in lymphomagenesis in vivo.Methods:We generated a novel mouse model, in which Cre‐mediated recombination leads to the conditional expression of Myd88 p.L252P (the orthologue of the human MYD88 p.L265P mutation) from the endogenous locus. We combined this Myd88 allele (abbr. M) with a BCL2 overexpression allele (abbr. B) and both alleles were activated B cell specifically with CD19:Cre (abbr. C). The effects of these mutations in vivo were carefully characterized. Additionally, we generated a conditional Cd79b p.Y195H allele and also obtained a previously published Tnfaip3 fl allele to model the loss of Tnfaip3 in DLBCL.Results:MBC animals showed elevated levels of self‐reactive antibodies. Immunization experiments revealed that Myd88 p.L252P and BCL2 overexpression cooperate in generating exaggerated antibody responses to exogenous antigen. Ultimately, these animals develop clonal tumors which resemble features of ABC DLBCL histologically and transcriptionally. They were exquisitely sensitive to inhibition of BCL2 in vitro and in vivo. MBC tumors expressed PD‐L1 at elevated levels compared to Eμ:Myc driven lesions. Anti‐PD1 antibody treatment had a stabilizing effect on tumor growth and resulted in a survival benefit. We successfully crossed our novel Cd79b p.Y195H allele into our MBC background to generate a mouse model of human DLBCL with oncogenic CD79B. Additionally, we generated a mouse model of TNFAIP3‐deficient lymhpoma by crossing the Tnfaip3 fl allele into our MBC background.Summary/Conclusion:Taken together, we established a Myd88 p.L252P driven mouse model for B cell lymphoma which shares many features with human ABC DLBCL. These tumors are sensitive to inhibition of BCL2 by ABT‐199 and respond to immune checkpoint blockade by anti‐PD1 treatment, suggesting these treatments as potential new strategies in the therapy of a subset of DLBCL patients. Additionally, we are currently investigating a novel conditional Cd79b p.Y195H allele and are modelling the loss of Tnfaip3 in DLBCL using a previously published allele.
PF521 CHOP TREATMENT ENHANCES DEPENDENCY ON BCL‐2, MCL‐1 AND BCL‐XL IN DIFFUSE LARGE B‐CELL LYMPHOMA
Authors: De Jong M. , Langendonk M. , Reitsma B. , Nijland M. , Huls G. , van den Berg A. , Ammatuna E. , Visser L. , van Meerten T. 
Published: June 2019

Background:Deregulation of anti‐apoptotic proteins BCL‐2, MCL‐1, BCL‐XL or BCL‐W is often a contributing factor to oncogenic transformation or progression, including in diffuse large B‐cell lymphoma (DLBCL). Gene expression profiling and immunohistochemistry analysis has shown overexpression of BCL‐2 in DLBCL is an important adverse prognostic factor, and has been correlated to resistance of standard therapy composed of cyclophosphamide, vincristine, doxorubicin, prednisolone and the anti‐CD20 antibody rituximab (R‐CHOP). Although these studies show a clear relation between BCL‐2 and therapy response, at the moment it remains unclear what the effect is of CHOP therapy on the intracellular anti‐apoptotic dependency in DLBCL.Aims:Here, we aim to determine the effect of CHOP treatment on the anti‐apoptotic proteins by employing BH3 profiling and investigate whether CHOP treatment enhances sensitivity for BH3 mimetic drugs.Methods:We first investigated the predictive value of anti‐apoptotic gene expression in 1800 DLBCL patients treated with CHOP or R‐CHOP. Next, dynamic BH3 profiles were acquired for 8 DLBCL cell lines treated with CHOP for 18 hours. Validation experiments were performed for CHOP combined with BH3 mimetic drugs on selected DLBCL cell lines. Finally, individual CHOP components were tested using BH3 profiling.Results:Gene expression analysis of anti‐apoptotic proteins revealed BCL‐2 had a significant negative association with survival in DLBCL patients, which was not observed for MCL‐1, BCL‐XL or BCL‐W. Analysis of anti‐apoptotic proteins expression in DLBCL cell lines demonstrated a strong adverse correlation for BCL‐2 protein expression and venetoclax (BCL‐2i) sensitivity, and a strong positive correlation for BCL‐2 protein expression and CHOP sensitivity. Dynamic BH3 profiling of CHOP‐treated DLBCL cell lines showed CHOP was able to enhance dependency on MCL‐1, BCL‐XL and BCL‐2. Validation experiments with BH3 mimetic drugs confirmed cell line specific enhanced sensitivity for S63845 (MCL‐1i), venetoclax (BCL‐2i) and/or navitoclax (BCL‐2/BCL‐XL/BCL‐Wi). No sensitivity‐related changes were observed on anti‐apoptotic protein expression after CHOP treatment. Dynamic BH3 analysis of single CHOP components revealed doxorubicin and vincristine were able to induce changes that were similar to the dynamic CHOP‐treated profiles.Summary/Conclusion:Gene expression analysis confirm that BCL‐2 status is a treatment‐independent predictive factor for DLBCL survival. However, our results demonstrate that CHOP treatment also induces cell‐specific increased anti‐apoptotic dependency on MCL‐1 or BCL‐XL, and not just BCL‐2. These results provide new perspective for the treatment of CHOP‐resistant DLBCL and underline the potential of BH3 profiling in predicting therapy outcome.image
PF548 PROGNOSTIC MUTATIONS IN PATIENTS WITH MYELODYSPLASTIC SYNDROME TREATED WITH HEMATOPOIETIC STEM‐CELL TRANSPLANTATION
Authors: Liu Z. , Cao S. , Lou F. , Sun Y. , Jiang E. , Han M. 
Published: June 2019

Background:Genetic mutations in MDS patients are closely related with clinical phenotypes and prognosis in MDS patients. But whether mutations are prognostic for outcomes after allogeneic hematopoietic stem‐cell transplantation(allo‐HSCT) remains to be elaborated.Aims:This research was done to figure out whether genetic mutations help the risk‐stratification in MDS patients received transplantation.Methods:Targeted mutational analysis were performed on samples obtained before transplantation from 151 patients underwent HSCT. Overall‐survival(OS) was calculated from the date of transplantation to date of death, and surviving patients were censored at the date on which they were last known to be alive. Diease‐free survival(DFS) was calculated from the time of transplantation to date of relapse or death and was censored at the last date known to alive without relapse. We analyzed the relationship of mutations and clinical outcomes. Curves for OS and DFS were generated using the Kaplan‐Meier method and compared using log‐rank test.Cumulative incidence of nonrelapse death and relapse was assessed using competing risks from time of HSCT to nonrelapse death or relapse by the Gray test.Results:All 151 patients carried more than one mutations, most frequently in DNAH2(47.02%), KDM6B(42.38%), USH2A(41.06%), KMT2D(38.41%), PCLO(33.11%), CCDC168(32.45%), U2AF1(29.14%), TET2(27.81%), ASXL1(25.83%), ARID1B(23.18%). In univariable analyses, TP53 or PPM1D mutations, ANKRD26 mutations, FLT3 abnormalities(including FLT3‐ITD), PDGFRB mutations were associated with shorter OS(TP53 or PPM1D, P = 0.064; ANKRD26, P = 0.025, FLT3 abnormalities: P = 0.046; PDGFRB, P = 0.044) and DFS(TP53 or PPM1D, P = 0.019; ANKRD26, P = 0.058, FLT3 abnormalities: P = 0.047; PDGFRB, P = 0.048). In multivariable analysis including clinical variables, monosomal karyotype status, HCT‐CI, and candidate genes, PDGFRB mutations and FLT3 abnormalities were each independently associated with shorter OS(PDGFRB mutations, HR = 2.979, P = 0.021; FLT3 abnormalities, HR = 2.677, P = 0.018) and DFS(PDGFRB mutations, HR = 2.779, P = 0.019; FLT3 abnormalities, HR = 2.288, P = 0.03). PDGFRB mutations(P = 0.042), ANKRD26 mutations (P = 0.004) were associated with higher non‐relapse mortality rate. FLT3 abnormalities was associated with higher relapse rate in RAEB patients(P = 0.004).Summary/Conclusion:PDGFR mutations and FLT3 abnormalities are independently associated with shorter survival in MDS patients treated with allo‐HSCT.
PF549 AUTOIMMUNE AND INFLAMMATORY DISEASES ASSOCIATE TO R‐IPSS SCORE BUT NOT TO OVERALL SURVIVAL OR LEUKEMIC EVOLUTION IN MYELODYSPLASTIC SYNDROMES
Authors: Brunet T. , Roy‐Peaud F. , Landron C. , Gallego‐Hernanz M.P. , Cayssials E. , Puyade M. , Torregrosa‐Díaz J.M. 
Published: June 2019

Background:Myelodysplastic syndromes (MDS) are clonal hemopathies with a relatively heterogeneous spectrum of presentation and a variable risk of transformation into acute myeloid leukemia (AML). Autoimmune diseases (AID) are present in 10 to 30% of MDS, with uncertain prognostic significance in this context. To date, no MDS‐related factor has been clearly identified as a predictor for the occurrence of AIDs.Aims:To study the association between MDS characteristics and the occurrence of an AID and its impact on overall survival and leukemia‐free survival.Methods:We retrospectively collected data from primary MDS adult patients followed‐up at the University Hospital of Poitiers between January 2010 and December 2017. Immunosuppressive therapy secondary‐MDS were excluded. The date of onset and type of AID as established by the standard criteria were collected. Steroid dependence was defined as a prednisone equivalent amount > 10 mg/day during at least 3 months. Simultaneous diagnosis of MDS and AID were considered if established in a 3‐months interval from the MDS one. Baseline characteristics at diagnosis of the MDS were registered and compared between those with and without AIDs.Results:AIDs were identified in 44 of 179 MDS patients (24.6%). MDS‐AID patients were more frequently male (29/44, p < 0.05). AIDs occurred before MDS in 63.6% of cases. Associated AIDs were: systemic vasculitis (23%), inflammatory rheumatism (18%), neutrophilic dermatosis (12.5%), connective tissue diseases (12.5%) and autoimmune cytopenia (11%). At the time of AID diagnosis, 48% of patients had general signs, 37.5% had cutaneous and/or joint signs. AID was steroid‐dependent in 39% of cases (9/23). No association between AID and MDS subtypes was observed. A significant linear tendency was found between the R‐IPSS and the presence of AID (p = 0.02): that is, the lower the IPSS‐R score, the higher the risk of association with AID. The presence of AID did not affect survival or progression to acute leukemia.Summary/Conclusion:The association of MDS to AIDs is frequent and of clinical heterogeneous expression. It occurs mainly in low‐risk MDS and does not seem to impact overall survival, that seems logical. The pathogenic mechanisms are complex and remain still completely unknown, but the hypothesis of a common pathogenic pathway seems strongly supported. The association MDS‐AID does not seem to impact overall survival in our cohort and occurs mainly in low‐risk MDS with a significant linear augmentation of the risk by the creasing categories of R‐IPSS. That raises the question of the role of different physio‐pathological processes underlying low‐risk and high‐risk MDS in the predisposition of AIDs. Larger prospective ongoing studies will better clarify our results.
PF639 IMMUNE ELECTRON MICROSCOPY AND MASS SPECTROMETRY FOR CLASSIFICATION OF AMYLOID DEPOSITS
Authors: Abildgaard N. , Rojek A.M. , Møller H.E. , Hansen C.T. , Palstrøm N.B. , Rasmussen L.M. , Beck H.C. , Marcussen N. 
Published: June 2019

Background:Amyloidosis is a shared name for a number of rare, complex and serious disease entities with an annual incidence in Denmark of about 2 per 100,000. Amyloidosis is caused by extra‐cellular deposits of amyloid substance created by accumulation of different misfolded proteins. Accurate characterization of the amyloid protein is essential. Amyloid light chain (AL) amyloidosis is a plasma cell dyscrasia that can be treated with chemotherapy, whereas transthyretin (ATTR) and secondary Amyloid A (AA) amyloidosis are other entities that are not treated within haematology. It is generally recognized that conventional immunohistochemistry has limitations, particular due to unspecific staining. Immune electron microscopy (IEM) and laser dissection microscopy followed by tandem mass spectrometry (LDM‐MS) are the new gold standards. It has not been established if one of the methods is superior; each method have some strengths and weaknesses. At Odense Amyloidosis Center, IEM and LDM‐MS have been established for standard diagnostics in 2017.Aims:To compare the sensitivity and specificity of IEM and MS for sybtyping amyloid deposits in different organsMethods:We performed a retrospective study of 106 Congo red positive biopsies from a number of different involved organs; heart, kidney, lung, gut mucosa, subcutaneous fat, bone marrow. IEM was performed with gold‐labeled antibodies against kappa, lambda, transthyretin and amyloid A.Results:IEM identified specific staining of amyloid fibrils in 91.6% of the biopsies; in 6 biopsies amyloid fibrils were not identified, and in 2 biopsies the subtype of the amyloid fibrils could not be established. MS identified an amyloigenic protein signature in 104/106 biopsies but in 9 of the biopsies the amyloid subtype could not be clearly identified. Protein subtype was established in 95 biopsies (89.6%). The concordance of the results ranged from 89.6–100% for the different subtypes. Importantly, the combined use of both methods increased the diagnostic sensitivity to 100%. Some variety in performances of the methods was observed on organ level.Summary/Conclusion:In conclusion, we demonstrate that both IEM and LDM‐MS are sensitive and specific methods for the identification of amyloidogenic proteins in amyloid deposits in biopsies from various organs.