Search

Several laboratory and clinical variables have been reported to be associated with the outcome of intensive chemotherapy for acute myeloid leukemia (AML), but only a few have been tested in the context of hematopoietic stem cell transplant (HSCT). This study aimed to identify genes whose expression of AML at diagnosis were associated with survival after HSCT. For this purpose, three publicly available adult AML cohorts (TCGA, BeatAML, and HOVON), whose patients were treated with intensive chemotherapy and then subjected to allogeneic or autologous HSCT, were included in this study. After whole transcriptome analysis, we identified ME1 as the only gene whose high expression was associated with shorter survival in patients subjected to HSCT. In addition, the inclusion of ME1 expression was able to improve the European LeukemiaNet risk stratification. Pathways related to lipid biosynthesis, mainly fatty acids, and cholesterol were positively correlated with ME1 expression. Furthermore, ME1 expression was associated with an M2 macrophage-enriched microenvironment, mature AML blasts hierarchy, and oxidative phosphorylation metabolism. Therefore, ME1 expression can be used as biomarker of poor response to HSCT in AML.

Several laboratory and clinical variables have been reported to be associated with the outcome of intensive chemotherapy for acute myeloid leukemia (AML), but only a few have been tested in the context of hematopoietic stem cell transplant (HSCT). This study aimed to identify genes whose expression of AML at diagnosis were associated with survival after HSCT. For this purpose, three publicly available adult AML cohorts (TCGA, BeatAML, and HOVON), whose patients were treated with intensive chemotherapy and then subjected to allogeneic or autologous HSCT, were included in this study. After whole transcriptome analysis, we identified

Deregulated cellular energetics is formally incorporated as an emerging hallmark of cancer, however little is known about its processes in myelodysplastic syndromes (MDS). Using transcriptomic data of CD34+ cells from 159 MDS patients and 17 healthy donors, we selected 37 genes involved in cellular energetics and interrogated about its clinical and prognostic functions. Based on the low expression of ACLY, ANPEP, and PANK1, as well as high expression of PKM and SLC25A5, we constructed our Molecular-Based Score (MBS), that efficiently discriminated patients at three risks groups: favourable risk (n = 28; 3-year overall survival (OS): 100%); intermediate (n = 60; 76% [62–93%]) and adverse (n = 71; 35% [17–61%]). Adverse MBS risk was independently associated with inferior OS (HR = 10.1 [95% CI 1.26–81]; P = 0.029) in multivariable analysis using age, gender and the revised international prognostic score system as confounders. Transcriptional signature revealed that Favourable- and intermediate-risk patients presented enriched molecular programs related to mature myeloid progenitors, cell cycle progression, and oxidative phosphorylation, indicating that this cells differs in their origin, metabolic state, and cell cycle regulation, in comparison to the adverse-risk. Our study provides the first evidence that cellular energetics is transcriptionally deregulated in MDS CD34+ cells and establishes a new useful prognostic score based on the expression of five genes.

Accumulating evidence suggest that the axon guidance molecules SLIT and ROBO are not only implicated in physiological process but also in cancer progression. Depending on the type of cancer the SLIT-ROBO axis can either act as a tumor suppressor gene, in which case the SLIT2 promoter site is frequently hypermethylated or as an oncogene, whereby high expression is often associated with poor prognosis. In the context of acute myeloid leukemia (AML), low expression of SLIT2 has been associated with low overall survival (OS) (Golos et al., 2019), while the functional role of SLIT2 remains largely unknown. Recently, we showed that the knockdown of SLIT2 increased cell proliferation of acute promyelocytic leukemia (APL) cells resulting in a more aggressive course of disease progression in vivo using the murine transgenic APL model (Weinhäuser et al., 2020). Here, we aimed to study the functional role of SLIT2 in a more heterogeneous disease, such as AML. Using different publicly available datasets. (GSE58477, normal karyotype blasts: 62, healthy CD34 +: 10; GSE63409, LSC: 14, HSC: 5) we detected increased methylation at the SLIT2 promoter site of AML leukemic cells compared to healthy CD34 + cells suggesting SLIT2 tumor suppressive functions. In addition, we measured decreased levels of SLIT2 in the bone marrow (BM) plasma of AML patients compared to healthy donors. To assess the biological role of SLIT2, we treated AML cell lines (KASUMI1, MV411, and MOLM13) with recombinant SLIT2 (50 ng/mL) in vitro. Administration of SLIT2 reduced AML cell growth, colony formation and induced cell cycle arrest in the G1 phase for all AML cell lines. Conversely, the knockdown of SLIT2 promoted increased THP-1 and OCI-AML3 cell proliferation. Next, we determined whether the treatment with SLIT2 could delay leukemogenesis in vivo using the AML cell line MV4-11. Engraftment was monitored by luciferase bioluminescent signal and NSGS mice were either treated with recombinant SLIT2 using a dose of 25 ng/g of body weight or vehicle (control group). SLIT2 therapy resulted in a lower disease burden, decreased leukemic infiltration in the BM and spleen, reduced spleen size, and increased OS compared to the control group (p Disclosures No relevant conflicts of interest to declare.

Introducción: El gen de fusión BCR-ABL1 está presente en al menos la cuarta parte de pacientes adultos con leucemia linfoblástica aguda de células B. Su detección determina la viabilidad del tratamiento con inhibidores de tirosina quinasas (TKIs) que se controla mediante la cuantificación de transcritos de BCR-ABL1. Algunos pacientes no responden o recaen al tratamiento debido a la presencia de mutaciones en el dominio tirosina quinasa del gen BCR-ABL1. Reporte de caso: Se reporta un paciente con BCR-ABL1 que logra una respuesta molecular luego de iniciado la terapia con imatinib; sin embargo, recae después de quince meses cambiándose el tratamiento a dasatinib. El cambio no permitió una respuesta molecular a la terapia. Retrospectivamente, se realizó la búsqueda de mutaciones en el BCR-ABL1 encontrándose tres mutaciones de resistencia a terapia (E459K, E255K y V299L). Conclusiones: La aparición de mutaciones durante el tratamiento con TKIs tiene un fuerte impacto en el progreso de la enfermedad, siendo relevante la búsqueda de mutaciones ante recaída o persistencia de transcritos del BCR-ABL1.

Objective: The significance of distinct immune cell subsets in focal lymphocytic sialadenitis (FLS) of Sjogren syndrome (SS) remains to be established. The aim of this study was to assess the frequency of immune cells and their activation status in minor salivary glands (MSGs) from patients with SS. Study Design: Using immunohistochemistry, we examined the frequency and the activation status of FLS-infiltrating CD4+/CD8+ T cells, regulatory T cells (Tregs), CD56+ natural killer cells (NKs), dendritic cells (DCs), and plasma cells (PCs) in 18 MSGs biopsies from patients with SS. Results: CD4+ rather than CD8+ T cells/NKs were predominant, with scarce expression of granzyme B and perforin. CD138+ PCs were equally abundant and exhibited a higher amount of IgG+/Kappa+ cells. Tregs expressed higher levels of Foxp3 than CD25. DCs showed expression of both mature (CD83 and CD208) and immature (CD207, followed by CD1a) markers. Conclusion: Our results show a high proportion of CD4+, CD8+, and CD138+ cells, whereas Tregs, NKs, and DCs are scarce in MSGs from patients with SS. The CD25/Foxp3 proportion might indicate a distinctive Treg subset, supporting an abnormal immune cell modulation. Noteworthy, the early infiltration of monoclonal PCs emphasizes the probable lymphoma risk transformation in patients with SS.

Background: Recently, the SLIT/ROBO axis became a therapeutic target for a variety of non-cerebral solid tumors because of its relevance in the regulation of angiogenesis, inflammatory cell chemotaxis, tumor cell migration and metastasis. In acute myeloid leukemia (AML), lower levels of SLIT2 expression were associated with poorer prognosis in studies that excluded patients with acute promyelocytic leukemia (APL) of the analysis (Golos et al., Arch Immunol 2019). Our group previously reported that higher SLIT2 expression was associated with improved overall survival, disease-free survival and decreased cumulative incidence of relapse. Subsequently, our in vitro functional studies indicated anti-proliferative effects exercised by exogenous SLIT2 peptide treatment in APL (Weinhauser et al., Blood 2018). Although, the expression of SLIT/ROBO was detected on leukemic cells, the fact that in healthy subjects the main source of SLIT2 derives from bone marrow stromal cells (BMSC) (Smith-Berdan et al., Cell Cycle 2012) was so far neglected in the context of hematological malignancies. Aims: Here, we expanded our data on the role of SLIT2 using lentivirally knocked down APL cells, and addressed the issue of the importance of BMSC-derived SLIT2 on APL development. Methods: NB4 and NB4R2 (RA-resistant) and primary APL cells (age, 25-47y; n=4) were transduced with shRNA-SLIT2 or shCT (control). After synchronization using double-thymidine block, transduced cells were submitted to proliferation and cell cycle assays. For the apoptosis analysis, cells were treated with ATO (1 µM) alone or in combination with RA (1 µM) for 24, 48 and 72 hours. The granulocytic differentiation in response to RA treatment alone (1 µM) or in combination with ATO (1 µM) for 48 and 72 hours, was evaluated based on the CD11b and CD11c levels. To assess the effect of SLIT2 silencing in BMSCs, HS5 and HS27A cell lines were transduced with shSLIT2 or shCT. Non-transduced NB4, NB4R2 and primary APL cells were co-cultured with engineered BMSCs and proliferation, apoptosis and granulocytic differentiation was performed in response to ATO alone or in combination with RA for 48 and 72 hours. In vivo assessment was performed using four 8-12 week-old C57/BL6PepBoy (CD45.1) lethally irradiated, transplanted with 5 x 106 hCG-PMLRARa blasts (CD45.2) transduced with shSlit2-murine and shCT. Results: Silencing SLIT2 in primary APL samples resulted in a significant increase in cell growth at the sixth day of culture compared to shCT samples (P.05). In contrast to shSLIT2 APL induced cell growth, co-culture of primary APL blast cells with transduced HS27A cells had no impact on proliferation, while the expression of the neutrophilic differentiation gene ELANE2 was significantly decreased in APL cells co-cultured with HS27A-shSLIT2.ConclusionSLIT2 is expressed in APL cells and exerts an anti-proliferative effect. On the other hand, SLIT2 produced by BMSC seemed to promote APL cellular stemness. Our results reinforce that SLIT2 function is cell type dependent and may be involved in the cross-talk between microenvironment and leukemic cells. Disclosures Figueiredo-Pontes: Novartis: Honoraria.

Background: TP73 isoforms gained particular relevance in acute promyelocytic leukemia (APL) since Bernasola et al (JEM. 2004) demonstrated that TAp73 was directly regulated by the PML protein in the nuclear body. The isoforms differ in their transcriptional activity, with those lacking domains in the N-terminal part of the protein exerting a dominant negative effect on TP73 function. In a retrospective analysis of patients with APL treated in ICAPL study, Lucena-Araujo et al (Blood 2015) demonstrated the association between higher ΔNp73/TAp73 ratio values and poor clinical outcome. However,there is a diversity of TP73 isoforms and specially those lacking N-terminal domains (e.g.ΔEx2p73, ΔEx2-3p73 and ΔN'p73) may be relevant in APL genesis and therapy response. Aims: Here, we quantified transcript levels of TP73 N-terminal variants TAp73, ΔNp73, ΔEx2p73, ΔEx2-3p73 and ΔN'p73, as well as the C-terminal variants TP73αand TP73β, and determined whether there is a prognostic correlation. In addition, we evaluated the effect of ΔNp73overexpression on APL cell lines survival and differentiation in vitro and in vivo. Methods: Bone marrow (BM) samples from 98 patients (age, 18-74y) with newly diagnosed APL enrolled in the International Consortium on Acute Leukemia (ICAPL2006) were included. For comparison, BM mononuclear cells from 14 healthy donors (age, 18-60y) were also included. TP73 transcripts were determined by qPCR and using survival ROC curve analysis and the C-index we dichotomized patients into "low" and "high" expression. Proportional hazard model on overall survival (OS) and disease-free survival (DFS) was performed to evaluate prognosis. In addition, empty vector (EV) or ΔNp73α was transduced in NB4 and NB4R2 (RA-resistant) APL cell lines using a lentivirus system. The apoptosis rate was evaluated in both cell lines upon ATO (1 μM) treatment for 24, 48 and 72h. ΔNp73α, cleaved caspase 3 and Bcl-2 protein abundance was detected by western blotting. Granulocytic differentiation induced by RA-treatment (1 μM) alone or in combination with ATO (1 μM) for 72 h was assessed by CD11b surface levels. Finally, lethally irradiated 8-12 week old C57/BL6 Boy (CD45.1) were transplanted with hCG-PMLRARa Blasts (CD45.2) ΔNp73α or EV transduced (ΔNp73α group=16 and EV group=12). Results: Compared to normal BM, APL patients presented higher levels of ΔNp73 (p=.004), ΔEx2p73 (p=.008), and TP73β (p.05) or disease progression in ΔNp73α blasts compared to the control group (p=.095). Conclusions: Our results suggest that transcript levels of ΔNp73, ΔEx2p73 and TAp73 predict outcomes in APL patients treated with RA plus chemotherapy. Additionally, RA plus ATO combination therapy seemed to overcome the effect of ΔNp73 overexpression in vitro. Disclosures Figueiredo-Pontes: Novartis: Honoraria. Pagnano:Abbvie: Consultancy; Sandoz: Consultancy; Pint Pharma: Consultancy. Tallman:Danbury Hospital Tumor Board: Honoraria; 14th Annual Miami Cancer Meeting: Honoraria; International Conference in Leukemia: Honoraria; KAHR: Membership on an entity's Board of Directors or advisory committees; Amgen: Consultancy; Nohla: Membership on an entity's Board of Directors or advisory committees, Research Funding; Rigel: Membership on an entity's Board of Directors or advisory committees; AbbVie: Membership on an entity's Board of Directors or advisory committees, Research Funding; New Orleans Summer Cancer Conference: Honoraria; Indy Hematology Review: Honoraria; Mayo Clinic: Honoraria; Orsenix: Membership on an entity's Board of Directors or advisory committees, Research Funding; UpToDate: Patents & Royalties; Salzberg Weill Cornall MSKCC Seminar in Hematologic Malignancies: Honoraria; Hematology Oncology of Indiana: Honoraria; ADC Therapeutics: Research Funding; Arog Pharmaceuticals: Research Funding; Cellerant Therapeutics: Research Funding; University of Oklahoma Medical Center: Honoraria; Bioline: Membership on an entity's Board of Directors or advisory committees, Research Funding; BioSight: Membership on an entity's Board of Directors or advisory committees, Research Funding; Daiichi-Sankyo: Membership on an entity's Board of Directors or advisory committees. Dillon:Abbvie: Consultancy, Honoraria; TEVA: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Novartis: Consultancy, Honoraria. Löwenberg:Chairman, Leukemia Cooperative Trial Group HOVON (Netherlands: Membership on an entity's Board of Directors or advisory committees; Clear Creek Bio Ltd: Consultancy, Honoraria; Editorial Board "European Oncology & Haematology": Membership on an entity's Board of Directors or advisory committees; Elected member, Royal Academy of Sciences and Arts, The Netherlands: Membership on an entity's Board of Directors or advisory committees; Frame Pharmaceuticals: Equity Ownership; Hoffman-La Roche Ltd: Membership on an entity's Board of Directors or advisory committees; Royal Academy of Sciences and Arts, The Netherlands: Membership on an entity's Board of Directors or advisory committees; Supervisory Board, National Comprehensive Cancer Center (IKNL), Netherland: Membership on an entity's Board of Directors or advisory committees; Chairman Scientific Committee and Member Executive Committee, European School of Hematology (ESH, Paris, France): Membership on an entity's Board of Directors or advisory committees; CELYAD: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Astex: Membership on an entity's Board of Directors or advisory committees; Astellas: Membership on an entity's Board of Directors or advisory committees; Up-to-Date", section editor leukemia: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees; Agios Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees.

Introduction: Meningioma 1 (MN1) gene was described as a prognostic marker for AML patients with normal karyotype (Carturan et al. Oncotarget 2016). In addition, MN1 high expression was linked to RA resistance and was shown to be necessary and sufficient to transform common myeloid progenitors in a MEIS1/AbdB-like HOX protein complex-dependent manner. However, the relevance of the copresence of MN1 overexpression in AML patients with mutations of the FLT3 and/or NPM1 genes is unknown. Moreover, it is also unknown the functional effects of MN1 in the biology of leukemic blasts harboring FLT3 mutations and if it may modulate the response to FLT3 inhibitors, such as quizartinib (AC220) and midostaurin (PKC). Aims: Herein, we investigated the prognostic impact of MN1 expression across multiple transcriptomic platforms and AML data sets. Additionally, we transduced different AML cell lines to evaluate the impact of MN1 on cell survival and differentiation. Methods: Three different AML series (1th: GSE6891, 240 patients, 2nd: TCGA, 113 patients and 3rd: BeatAML, 139 patients) were used. All patients presented similar gender distribution, were above 18y and treated by the 3+7 scheme. All genes from the RNAseq (TCGA) were pre-ranked according to their differential expression comparing tumors with high and low expression of MN1, using their median expression rate as the cutoff. GSEA was performed using the Reactome, KEGG and Hallmarks databases. Additionally, we transduced 07 AML cell lines with the MN1 gene and the control. For those cells, clonogenicity and proliferation rate was evaluated to identify which cell lines are sensitive and resistant to MN1. Drug induced apoptotic rate was assessed for FLT3-ITD- cell lines when treated with AraC (10 nM) and FLT3-ITD+ cell lines (MOLM13/MV411) upon treatment with PKC and AC220 over a time period of 24, 48 and 72 h. Apoptosis was further confirmed by cleaved Caspase-3/PARP detection. The myeloid differentiation in response to PMA treatment (100 ng/ml) was determined by the surface levels of CD11b, CD11c, CD14, CD15, CD61 and HLADR. Results: In all three AML series, high MN1 levels higher than the median (hereafter called High MN1) were associated with lower frequency of FLT3 and NPM1 mutations (P Disclosures Heuser: Bayer Pharma AG, Berlin: Research Funding; Synimmune: Research Funding.

Background : The recent efforts to uncover the molecular heterogeneity of myelodysplastic syndromes (MDS), mainly by new sequencing technologies, allow the comprehensive identification of driver mutations and/or altered gene expression recurrently found in a recognizable fraction of patients. Ongoing efforts are being made to clarify the impact of molecular changes on clinical phenotype and prognosis, as well as their role in the pathogenesis of MDS. Refining risk stratification allows the proposition of risk-adapted therapy and may shed light in biology of MDS. Aims: Based on the gene expression of selected metabolic targets, we aimed to design a score system that improves MDS overall survival prediction. Patients and methods: Clinical, mutations and transcriptomic data from CD34+ cells from 159 MDS patients and 17 healthy volunteers freely available at Gene Expression Omnibus (GEO/NCBI: GSE58831) were used in the present work. Forty-one genes related to metabolic processes, previously demonstrated as deregulated among diverse neoplastic conditions, were ranked and asked for differential expression and prognostic impact. Each gene was dichotomized according to Receiving-Operating Curve (ROC) and Cox Proportional-Hazard Model was used for multivariate analysis using gender, age and IPSS-R as cofounders. Genes independently associated with overall survival (OS) were selected to compose the Molecular-Based Score (MBS) and integer weight of each one was defined according Hazard Ratio (HR). Survival curves were constructed using Kaplan-Meyer method and compared with Log-Rank Test. ROC c-statistic was used to measure the predictive function of MBS. Prediction accuracy of MBS was cross-validated by a nonparametric bootstrap procedure with 1,000 resamplings of the original cohort allowing replacement and also estimated their respective 95% confidence interval (95% CI) computing the bias-corrected and accelerated bootstrap interval. Results: Among selected genes, 18 were differentially expressed between CD34+ cells from MDS and healthy volunteers. Fifteen genes predict OS in univariate analysis, of which ACLY (HR: 0.48; 95%CI: 0.24 - 0.96; P=0.04), ANPEP (HR: 2.16; 95%CI: 1.08 - 4.31; P=0.02), PANK1 (HR: 0.43; 95%CI: 0.19 - 0.98; P=0.04), PKM (HR: 2.01; 95%CI: 1.02 - 3.93; P=0.04) and SLC25A5 (HR: 0.52; 95%CI: 0.27 - 0.99; P=0.05) were independently associated with OS. Higher expression of ANPEP and PKM, as well as lower expression of ACLY, PANK1 and SLC25A5 were considered to integer high risk being attributed weight 2 for each condition. MBS varied from 0 to 10 (median=2) and was calculated as: MBS Low-Risk =0 (MBS-LR; n=28); MBS Intermediate-Risk=2 and 4 (MBS-IR; n=90) and High-Risk: ≥6 (MBS-HR; n=48). The modeled MBS showed a ROC c-statistic of 0.699 (95%CI: 0.603 - 0.794) and HR=3.05 (95%CI: 1.81 - 5.05; P Figure Disclosures No relevant conflicts of interest to declare.