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Mitani, Kinuko. "11. Molecular Physiopathology and Molecular Targeting Therapy of Leukemia". Nihon Naika Gakkai Zasshi 96, n.º 9 (2007): 2013–19. http://dx.doi.org/10.2169/naika.96.2013.
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Mitani, Kinuko. "11. Molecular Physiopathology and Molecular Targeting Therapy of Leukemia". Nihon Naika Gakkai Zasshi 96, Suppl (2007): 87b—88a. http://dx.doi.org/10.2169/naika.96.87b.
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Siviero-Miachon, Adriana Aparecida, Angela Maria Spinola-Castro y Gil Guerra-Junior. "Adiposity in childhood cancer survivors: insights into obesity physiopathology". Arquivos Brasileiros de Endocrinologia & Metabologia 53, n.º 2 (marzo de 2009): 190–200. http://dx.doi.org/10.1590/s0004-27302009000200011.
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As childhood cancer treatment has become more effective, survival rates have improved, and a number of complications have been described while many of these patients reach adulthood. Obesity is a well-recognized late effect, and its metabolic effects may lead to cardiovascular disease. Currently, studies concerning overweight have focused on acute lymphocytic leukemia and brain tumors, since they are at risk for hypothalamic-pituitary axis damage secondary to cancer therapies (cranial irradiation, chemotherapy, and brain surgery) or to primary tumor location. Obesity and cancer have metabolic syndrome features in common. Thus, it remains controversial if overweight is a cause or consequence of cancer, and to date additional mechanisms involving adipose tissue and hypothalamic derangements have been considered, comprising premature adiposity rebound, hyperinsulinemia, leptin regulation, and the role of peroxisome proliferator-activated receptor γ. Overall, further research is still necessary to better understand the relationship between adipogenesis and hypothalamic control deregulation following cancer therapy.
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Pasquer, Hélène, Maëlys Tostain, Nina Kaci, Blandine Roux y Lina Benajiba. "Descriptive and Functional Genomics in Acute Myeloid Leukemia (AML): Paving the Road for a Cure". Cancers 13, n.º 4 (11 de febrero de 2021): 748. http://dx.doi.org/10.3390/cancers13040748.
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Over the past decades, genetic advances have allowed a more precise molecular characterization of AML with the identification of novel oncogenes and tumor suppressors as part of a comprehensive AML molecular landscape. Recent advances in genetic sequencing tools also enabled a better understanding of AML leukemogenesis from the preleukemic state to posttherapy relapse. These advances resulted in direct clinical implications with the definition of molecular prognosis classifications, the development of treatment recommendations based on minimal residual disease (MRD) measurement and the discovery of novel targeted therapies, ultimately improving AML patients’ overall survival. The more recent development of functional genomic studies, pushed by novel molecular biology technologies (short hairpin RNA (shRNA) and CRISPR-Cas9) and bioinformatics tools design on one hand, along with the engineering of humanized physiologically relevant animal models on the other hand, have opened a new genomics era resulting in a greater knowledge of AML physiopathology. Combining descriptive and functional genomics will undoubtedly open the road for an AML cure within the next decades.
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Gueiderikh, Anna, Frédérique Maczkowiak-Chartois, Guillaume Rouvet, Sylvie Souquère-Besse, Sébastien Apcher, Jean-Jacques Diaz y Filippo Rosselli. "Fanconi anemia A protein participates in nucleolar homeostasis maintenance and ribosome biogenesis". Science Advances 7, n.º 1 (enero de 2021): eabb5414. http://dx.doi.org/10.1126/sciadv.abb5414.
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Fanconi anemia (FA), the most common inherited bone marrow failure and leukemia predisposition syndrome, is generally attributed to alterations in DNA damage responses due to the loss of function of the DNA repair and replication rescue activities of the FANC pathway. Here, we report that FANCA deficiency, whose inactivation has been identified in two-thirds of FA patients, is associated with nucleolar homeostasis loss, mislocalization of key nucleolar proteins, including nucleolin (NCL) and nucleophosmin 1 (NPM1), as well as alterations in ribosome biogenesis and protein synthesis. FANCA coimmunoprecipitates with NCL and NPM1 in a FANCcore complex–independent manner and, unique among the FANCcore complex proteins, associates with ribosomal subunits, influencing the stoichiometry of the translational machineries. In conclusion, we have identified unexpected nucleolar and translational consequences specifically associated with FANCA deficiency that appears to be involved in both DNA damage and nucleolar stress responses, challenging current hypothesis on FA physiopathology.
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Rossignol, Julien, Laura Polivka, Leila Maouche-Chrétien, Laurent Frenzel, Patrice Dubreuil y Olivier Hermine. "Recent advances in the understanding and therapeutic management of mastocytosis". F1000Research 8 (22 de noviembre de 2019): 1961. http://dx.doi.org/10.12688/f1000research.19463.1.
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Mastocytosis is a rare disease due to the abnormal accumulation of mast cells in various tissues. Its clinical presentation is heterogeneous depending on mast cell infiltration and mediators release. In some cases, it is associated with hematological malignancies. Prognosis varies from very good with a life expectancy similar to the general population in indolent forms of the disease to a survival time of just a few months in mast cell leukemia. Although in most cases a somatic KIT D816V mutation is found in tumor mast cells, the physiopathology of the disease is not yet fully understood. Additional germline and somatic mutations may explain this heterogeneity. Treatments aim at blocking effect of mast cell mediators, reducing mast cell activation and tumor burden. New drugs mainly directed against the tyrosine kinase activity of KIT have dramatically changed the quality of life and prognosis of mast cell diseases. Present and future therapeutic strategies are discussed in this review.
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Fernández-Torres, Javier, Denhi Flores-Jiménez, Antonio Arroyo-Pérez, Julio Granados y Alberto López-Reyes. "HLA-B*40 Allele Plays a Role in the Development of Acute Leukemia in Mexican Population: A Case-Control Study". BioMed Research International 2013 (2013): 1–6. http://dx.doi.org/10.1155/2013/705862.
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Among oncohematological diseases, acute lymphoid leukemia (ALL) and acute myeloid leukemia (AML) are characterized by the uncontrolled production and accumulation of blasts that can lead to death. Although the physiopathology of these diseases is multifactorial, a genetic factor seems to be at play. Several studies worldwide have shown association of ALL and AML with several alleles of the major histocompatibility complex (MHC).Objective. To determine gene frequencies of HLA-B alleles in Mexicans (individuals with Native American genetic background admixed with European descent) with ALL and AML.Methods. We compared the HLA-B alleles in 213 patients with ALL and 85 patients with AML to those present in 731 umbilical cord blood (UCB) samples as a control group; this was done by means of the PCR-SSP technique.Results. We found an increased frequency of the HLA-B*40 allele in ALL patients as compared to the control group (14.5% versus 9.84%,P=0.003, OR = 1.67); this was particularly evident in a subgroup of young (less than 18 years old) ALL patients (P=0.002, OR = 1.76); likewise, a decreased frequency of HLA-B*40 allele in AML patients was observed as compared to the control group (4.70% versus 9.84%,P=0.02, OR = 0.42).Conclusions. These results might suggest opposing effects of the HLA-B*40 in the genetic susceptibility to develop ALL or AML and offer the possibility to study further the molecular mechanisms of cell differentiation within the bone marrow lineage.
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Soulier, Jean. "Fanconi Anemia". Hematology 2011, n.º 1 (10 de diciembre de 2011): 492–97. http://dx.doi.org/10.1182/asheducation-2011.1.492.
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Abstract Fanconi anemia (FA) is the most frequent inherited cause of BM failure (BMF). Fifteen FANC genes have been identified to date, the most prevalent being FANCA, FANCC, FANCG, and FANCD2. In addition to classical presentations with progressive BMF during childhood and a positive chromosome breakage test in the blood, atypical clinical and/or biological situations can be seen in which a FA diagnosis has to be confirmed or eliminated. For this, a range of biological tools have been developed, including analysis of skin fibroblasts. FA patients experience a strong selective pressure in the BM that predisposes to clonal evolution and to the emergence in their teens or young adulthood of myelodysplasia syndrome (MDS) and/or acute myeloid leukemia (AML) with a specific pattern of somatic chromosomal lesions. The cellular mechanisms underlying (1) the hematopoietic defect which leads to progressive BMF and (2) somatic clonal evolutions in this background, are still largely elusive. Elucidation of these mechanisms at the molecular and cellular levels should be useful to understand the physiopathology of the disease and to adapt the follow-up and treatment of FA patients. This may also ultimately benefit older, non-FA patients with aplastic anemia, MDS/AML for whom FA represents a model genetic condition.
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Schinke, Carolina D., Cody Ashby, Yan Wang, Ruslana G. Tytarenko, Eileen Boyle, Christopher Wardell, Pingping Qu et al. "The Mutational Landscape of Primary Plasma Cell Leukemia". Blood 132, Supplement 1 (29 de noviembre de 2018): 114. http://dx.doi.org/10.1182/blood-2018-99-116758.
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Abstract Introduction: Primary Plasma Cell Leukemia (pPCL) is a rare form of multiple myeloma (MM) that is characterized by an aggressive disease course with >20% peripherally circulating plasma cells (PCs) and poor clinical outcome. Despite the advances of modern anti-MM therapy, pPCL patients continue to experience low median overall survival (OS) suggesting a distinct biological background. Due to its low incidence of 1-2% of all MM patients, studies on physiopathology remain challenging and are limited. The aim of this study was to elucidate the differences in biology and outcome between non-pPCL MM and pPCL, to determine the genetic landscape of pPCL and to identify distinct signatures and pathways that potentially could be used as therapeutic targets. Methods: We performed gene expression profiling (GEP; Affymetrix U133 Plus 2.0) of matched circulating peripheral PCs and bone marrow (BM) PCs from 13 patients. Whole exome sequencing (WES) was performed on purified CD138+ PCs from BM aspirates from 19 pPCL patients with a median depth of 61x. CD34+ sorted cells, taken at the time of stem cell harvest from the same 19 patients, were used as controls. Translocations and mutations were called using Manta and Strelka and annotated as previously reported. Copy number was determined by Sequenza. Results: GEP from the BM and circulating peripheral PCs showed that the expression patterns of the two samples from each individual clustered together, indicating that circulating PCs and BM PCs in pPCL result from the same clone and are biologically clearly related. The clinical characteristics from the patient cohort used for WES analysis were as follows: median age was 58 years (range 36-77), females accounted for 74% (14/19), an elevated creatinine level was found in 78% (14/18) and an elevated LDH level in 71% (10/14). All patients presented with an ISS stage of III. Median OS of the whole dataset was poor at 22 months, which is consistent with OS from previously reported pPCL cohorts. Primary Immunoglobulin translocations were common and identified in 63% (12/19) of patients, including MAF translocations, which are known to carry high risk in 42% (8/19) of patients [t(14;16), 32% and t(14;20), 10%] followed by t(11;14) (16%) and t(4;14) (10%). Furthermore, 32% (6/19) of patients had at least one MYC translocation, which are known to play a crucial role in disease progression. MYC breakpoints (8q24) were identified in 25% with Ig partner loci including IGH (5%), IGK (10%), and IGL (10%). The remaining samples had partner loci including FAM46C (5%), MYNN (5%), SPARC (5%), QRSL1 (5%), RNF126 (5%), PLXNA4 (5%) and CDH7 (5%). The mutational burden of pPCL consisted of a median of 98 non-silent mutations per sample, suggesting that the mutational landscape of pPCL is highly complex and harbors more coding mutations than non-pPCL MM. Driver mutations, that previously have been described in non-pPCL MM showed a different prevalence and distribution in pPCL, including KRAS and TP53 with 47% (9/19) and 37% (7/19) affected patients respectively compared to 21% and 5% in non-PCL MM. PIK3CA (5%), PRDM1 (10%), EP300 (10%) and NF1 (10%) were also enriched in the pPCL group compared to previously reported cases in non-pPCL MM. Biallelic inactivation of TP53 - a feature of Double Hit myeloma - was found in 6/19 (32%) samples, indicating a predominance of high risk genomic features compared to non-pPCL MM. Furthermore, analysis of mutational signatures in pPCL showed that aberrant APOBEC activity was highly prevalent only in patients with a MAF translocation, but not in other translocation groups. Conclusion: In conclusion we present one of the first WES datasets on pPCL with the largest patient cohort reported to date and show that pPCL is a highly complex disease. The aggressive disease behavior can, at least in part, be explained by a high prevalence of MAF and MYC translocations, TP53 and KRAS mutations as well as bi-allelic inactivation of TP53. It is of interest that only KRAS but not NRAS mutations are highly enriched in pPCL. From all highly prevalent genomic alterations in pPCL, only KRAS mutations offer a potential for already available therapeutically targeting with MEK inhibitors, which should be further explored. Disclosures Davies: Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees; ASH: Honoraria; TRM Oncology: Honoraria; Amgen: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy, Honoraria; Abbvie: Consultancy; Celgene: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; MMRF: Honoraria. Barlogie:Multiple Myeloma Research Foundation: Other: travel stipend; ComtecMed- World Congress on Controversies in Hematology: Other: travel stipend; Millenium: Consultancy, Research Funding; European School of Haematology- International Conference on Multiple Myeloma: Other: travel stipend; International Workshop on Waldenström's Macroglobulinemia: Other: travel stipend; Celgene: Consultancy, Research Funding; Dana Farber Cancer Institute: Other: travel stipend; Myeloma Health, LLC: Patents & Royalties: : Co-inventor of patents and patent applications related to use of GEP in cancer medicine licensed to Myeloma Health, LLC. Morgan:Bristol-Myers Squibb: Consultancy, Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Takeda: Consultancy, Honoraria; Janssen: Research Funding.
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Telliam, Gladys, Christophe Desterke, Olivier Féraud, Frank Griscelli, Noufissa Oudrhiri, Micheline Fontaine Arnoux, Radhia Najar, Herve Acloque, Annelise Bennaceur-Griscelli y Ali G. Turhan. "Blast Crisis in a Dish: Generation of a Blast Crisis Model in Chronic Myeloid Leukemia (CML) Using Patient-Specific Induced Pluripotent Stem Cells ( iPSC)". Blood 128, n.º 22 (2 de diciembre de 2016): 933. http://dx.doi.org/10.1182/blood.v128.22.933.933.
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Abstract Despite the major progress obtained in prognosis with the use of tyrosine kinase inhibitors (TKI), the great majority of patients with CML remain on long-term therapy and progression occurs in patients with either primary or secondary resistance. The mechanisms of progression towards accelerated phase (AP) and blast crisis (BC) have been studied so far only in primary patient samples in BC. Currently, there is no in vitro model to study sequentially the molecular events leading from CP to BC as only some primary sequential samples are amenable to analysis. Using induced Pluripotent Stem Cell (iPSC) technology, it is now possible to reprogram the primary leukemic cells to pluripotency and generate a major source of stem cells. To determine the feasibility of studying progression of CML towards AP and BC, we have used a patient-specific iPSC line that we have generated from the primary leukemic cells of a patient who later showed a TKI-resistance requiring an allogeneic stem cell transplant. These iPSC expressed BCR-ABL, exhibited all pluripotency markers and after injection in NSG mice, generated teratoma with differentiation into three germ layers. In hematopoietic differentiation assays using day 19-embryoid bodies (EB), increased numbers of hematopoietic progenitors were found as compared to control iPSC (5-fold increase n= 3). We have then treated leukemic iPSC with the mutagenic agent N-ethyl-N-nitrosourea (ENU) during regular medium changes. After 61 days in ENU cultures, day-19 derived embryoid bodies generated hematopoietic cells (>90% CD45+, CD43+) which proliferated in liquid cultures with myeloid and some blast cell morphology. Cytogenetic analyses of iPSC revealed chromosomal abnormalities such as loss of Y and loss of der q9+, both alterations known to occur in CML during progression. They exhibited increased numbers of micronuclei (MN) as compared to leukemic iPSC without ENU (X 3 Fold increase) suggesting acquisition of a progressive chromosomal instability. CGH array analyses were performed using ENU-treated iPSC-derived hematopoietic cells in two different timepoints as compared to leukemic iPSC cultured without ENU. Genomic aberrations were analyzed by Agilent Cytogenomics software with Mosaicism workflow on HG19 genome. 249 gene loci alterations were detected after polymorphism filtration on European population. These analyses showed DNA losses and DNA gains in genes implicated in mesoderm development and hematopoietic lineage as well as genes implicated in DNA damage response. Several loci of transcription factors were found to be involved such as IKZF1 described in imatinib-refractory chronic myeloid leukemia (Bolton-Gillespie et al. 2013). The aberrations included SIRT1and BLM which is implicated in DNA repair. Several cancer genes were found to be involved, some known to be altered in leukemia (BLM, IKZF1, NCOA2, ALK, EP300, ERG, MKL1, PHF6 and TET1). Remarkably, transcriptome geodataset GSE4170 (Radich et al. 2006) allowed us to associate 125 of 249 of the aberrations that we detected in CML iPSC, with the CML progression genes already described during progression from chronic and AP to BC (p-value =9.43E-32, after ANOVA with 1000 permutations). 38 most predictive aberrations allowed perfect reclassification of BC and chronic phase samples by unsupervised classification. Among these candidates, eleven of them have been described in CML physiopathology and connected to TKI resistance and genomic instability. Majority of them ( 7/11) are connected to chronic phase (FAS, ACTB, TRIM21, ANPEP, MLK1, CSF2RA, and MAGEC2) whereas a minority of them (4/11) are connected to BC (ACP1, SH3YL1, FHL1, IL3RA). Interestingly, these experiments also allowed us to discover the connection of a new multidrug resistance molecule associated to BC and having the ability to modify interferon pathway connected to the TKI sensitivity. Thus, genomic instability pattern that we have generated using a single leukemic iPSC allowed duplication of genomic abnormalities described in CML progression and allowed identification of some novel genes. Overall, these results demonstrated that we have generated for the first time to our knowledge, an in vitro BC model, reproducing genomic events described in patients with BC. This "blast crisis in a dish" tool using patient-derived iPSC will be of major interest to study CML progression and eventually to test novel therapies. Figure. Figure. Disclosures No relevant conflicts of interest to declare.
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Mascarenhas, Cintia, Lara Woldmar, Maria Helena Almeida, Rosangela Vieira Andrade, Anderson Ferreira Cunha y Carmino A. De Souza. "Evaluation of Peroxiredoxins (PRDX1, PRDX2 and PRDX6) Expression in Patients with Chronic Myeloid Leukemia (CML) Treated with Imatinib in First Line". Blood 124, n.º 21 (6 de diciembre de 2014): 5545. http://dx.doi.org/10.1182/blood.v124.21.5545.5545.
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Abstract Introduction: Satisfactory response is present for the majority of chronic myeloid leukemia (CML) patients (pts) in chronic phase (CP) treated with tyrosine kinase inhibitors (ITK) . However, some pts exhibit suboptimal response or treatment failure. The probability of achieving optimal response may be related with several factors. The oxidative stress modulation is tightly related with the physiopathology of various hematologic diseases and can cause cell death, apoptosis and necrosis. Peroxiredoxins (Prdx) are a family of multifunctional antioxidant thioredoxin-dependent peroxidases that protect cells against oxidative stress and modulate signaling cell proliferation pathways and may influence the metabolism of ITKs.The aim of this study was to analyze PRDX1, PRDX2 and PRDX6 levels of CML pts and correlate with cytogenetics and molecular responses. Methods: PRDX1, PRDX2 and PRDX6 expression was evaluated in 20 blood donors, 18 newly diagnosed CML pts and 22 previously treated pts. Pts were treated with imatinib 400-600mg in first line. Samples were collected from peripheral blood at diagnosis or during treatment and RNA samples were submitted to the synthesis of complementary DNA (cDNA) using the kit RevertAid™ HMinus First Strand cDNA Synthesis Kit (Fermentas, Life Sciences). For cDNA synthesis, 3 ug of RNA was used and peroxiredoxins expression was evaluated by real-time PCR with Syber Green (Applied Biosystems) and endogenous (β-Actina and GAPDH) controls. The results were analyzed using 2-ΔΔCT. Statistical analysis were made by using Mann Withney’s T test. Cytogenetic analysis was performed at diagnosis, 3, 6, 12 and 18 months after starting therapy and then every 12-24 months thereafter if CCR was achieved. BCR-ABL transcripts were measured in peripheral blood at 3-month intervals using quantitative RQ-PCR. Results were expressed as BCR-ABL/ABL ratio, with conversion to the international scale (IS). Major molecular response (MMR) was defined as a transcript level ≤ 0.1% (IS). Results: 40 CML pts, 55% male, median age of 53 years (23-84) were evaluated, 60% in chronic phase (CP), 30% in accelerated phase (AP) and 10% in blast crisis (BC). The mean of PRDX transcript levels in the total group was (PRDX1: 0.006 and 10.10 / PRDX2: 0.002 and 16.26 / PRDX6: 0.003 and 49.97) respectively (PRDX1: 1.2 / PRDX2: 0.9 / PRDX6: 15.36). The results showed that there are a significantly difference (p<0.05) in the PRDX gene expression between pts and blood donors. All PRDX expression was reduced in responsive patients, and increase expression in pts resistant to TKI. The median duration of imatinib treatment was 29 months (1-104) and 97% achieved complete hematological response, 75% complete cytogenetic response and 65% major or complete molecular response. The analysis showed that higher levels of PRDX were maybe correlated with a no reduction of BCR-ABL transcripts (p<0.05). As well as, there was may influence of the PRDX levels at diagnosis in the response at 24 months of treatment. Conclusion:Is known that that the increase of ROS in CML leads to an increase of DNA damage, triggering genomic instability and resulting in accumulation of mutations and chromosomal aberrations, and contribute to the mechanism of acquisition of resistance to TKI inhibitors. The decrease of Peroxiredoxins expression observed in the responsive group, could contribute to this process, since the detoxification of these species are compromising and the effects caused by oxidative stress are even more drastic, leading to mutations that could be followed by TKi resistance. The relation between Prdx and CML not yet been elucidated. Disclosures No relevant conflicts of interest to declare.
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de Melo Campos, Paula, Joao Machado-Neto, Adriana Silva Santos Duarte, Rafaela Mendonça, Irene Lorand-Metze, Fernando F. Costa, Sara T. O. Saad y Fabiola Traina. "IRS2 Associates With JAK2 and May Be Involved In Cell Proliferation Pathways In Chronic Myeloproliferative Neoplasms". Blood 122, n.º 21 (15 de noviembre de 2013): 1598. http://dx.doi.org/10.1182/blood.v122.21.1598.1598.
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Abstract Background Polycythemia Vera (PV), Essential Thrombocythemia (ET) and Primary Myelofibrosis (PMF) are BCR-ABL1 negative Chronic Myeloproliferative Neoplasms (MPN) characterized by increased myeloid proliferation, with predominant erythroid, megakaryocytic and megakaryocytic/granulocytic expansion, respectively. The finding of a recurrent mutation in the gene of the tyrosine-kinase Janus kinase 2 (JAK2 V617F) in these diseases has raised the hypothesis that this could be the main cause of their development. However, the evidence that MPN patients have a very similar response to JAK2 inhibitors regardless of JAK2 mutation status, and the knowledge that many receptors and substrates may lead to the activation of JAK/STAT, Ras/Raf/MAP kinases and PI3K/Akt/mTOR pathways, indicate the need to investigate other crucial proteins involved in the physiopathology of these diseases. Insulin receptor substrate 2 (IRS2) mediates mitogenic and antiapoptotic signaling from IR, IGF-IR, EPO-R and TPO-R. Previous studies performed on non-hematological cell lines have shown the association of IRS2 with JAK/STAT, PI3K/Akt/mTOR and Ras/Raf/MAP kinases pathways, giving rise to the hypothesis that IRS2 could participate in the activation of crucial signaling pathways in MPN through direct interaction with JAK2 or through alternative mechanisms. Aims To identify the JAK2/IRS2 protein interaction and to study the effects of pharmacological JAK1/2 inhibition (Ruxolitinib) over IRS2 phosphorylation in leukemia cell lines harboring or not the JAK2 V617F mutation; to characterize IRS2 expression in CD34+ cells from patients with MPN and its correlation with clinical data including JAK2 mutation status. Methods Leukemia cell lines carrying JAK2 V617F mutation (HEL) or not (HL60) were used for immunoprecipitation and immunobloting with IRS2 and JAK2 antibodies. Cells treated or not with JAK1/2 inhibitor Ruxolitinib were also submitted to immunoprecipitation and immunobloting with IRS2 and anti-phosphotyrosine antibodies. Peripheral blood mononuclear cells from 28 healthy donors and 97 patients with MPN (PV=28, ET=38, PMF=31) were included, and CD34+ cells were submitted to quantitative PCR (q-PCR). Relative expression of IRS2 was correlated with clinical data and with JAK2 V617F mutation status. Results Immunoprecipitation analysis showed that IRS2 associates with JAK2 in leukemia cell lines harboring (HEL) or not (HL60) the JAK2 V617F mutation. Furthermore, treatment of HEL cell line with the JAK1/2 selective inhibitor Ruxolitinib resulted in decreased IRS2 tyrosine phosphorylation. IRS2 mRNA expression in CD34+ cells were significantly higher in patients with ET when compared to healthy donors (1.70 [0.42-10.60] versus 0.87 [0.01-11.22], p=0.03). There was no difference in IRS2 mRNA expression in PV or PMF patients when compared to healthy donors. Furthermore, significantly higher levels of IRS2 mRNA expression were observed in patients harboring JAK2 V617F mutation when compared to the wild type JAK2 for ET (2.37 [0.96-10.60], n=14 versus 1.54 [0.42-1.54], n=22; p=0.01); and for PMF (2.27 [0.003-10.59], n=20 versus 0.60 [0.02-2.42], n=11; p=0.02). Although there was also a significant difference in IRS2 mRNA expression in mutated versus non mutated JAK2 in PV (p=0.02), the number of non mutated samples was low (n=2). Conclusions Our data indicate that IRS2 is a binding partner of JAK2 in myeloproliferative neoplasms and suggest that this protein association may be involved in cell proliferation in these diseases. The higher IRS2 expression in mutated samples (JAK2 V617F) might be associated with the constitutive activation of JAK2 in these samples. Disclosures: No relevant conflicts of interest to declare.
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Wang, Lining, Emmanuel Raffoux, Xavier Thomas, Ibrahim Yakoub-Agha, Jean Henri Bouhris, Stephane de Botton, Mauricette Michallet et al. "Skin Immune Stimulation By Infections and Drug Toxicities during Induction and/or Consolidations Increases Incidence of Skin Acute Graft Versus Host Disease in Acute Myeloid Leukemia: A Study on Behalf of SFGM-TC and ALFA". Blood 126, n.º 23 (3 de diciembre de 2015): 1946. http://dx.doi.org/10.1182/blood.v126.23.1946.1946.
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Abstract Introduction: 60-70% of AML patients have an indication of allogeneic hematopoietic stem cell transplantation (allo-HSCT) during their treatment. Among those who undergo allo-HSCT, prognosis and quality of life depended on presence or absence of graft versus host disease (GvHD). Immune stimulation supports the principle of GvHD and graft versus leukemia (GvL) after allo-HSCT. The impact of immune activation prior to allo-HSCT on the pathogenesis of GvHD has never been evaluated. The aim of this study was to determine whether immune stimulation induced by infection or drug toxicity before transplantation increased the incidence of acute GvHD (aGvHD) in AML patients. Materials and methods: 345 AML patients were transplanted in first complete remission (CR) in 21 French centers from 2009 to 2013 after prospectively enrollment in the ALFA-0702 trial (patients aged 18-60y, de novo AML, favorable-risk AML excluded). Clinical data (skin, gut and liver infections or drug toxicities) before allo-HSCT were collected from the ALFA database and clinical data (skin, gut and liver aGvHD) after allo-HSCT were collected from the SFGM-TC database (ProMise). GvHD grading was defined according to Glucksberg criteria. Infection and drug toxicity grading was defined according to CTCAE v4.0. Mann-Withney and Kruskall-Wallis tests were used for continuous variables. Chi-square test was used for non-continuous variables. Overall survival (OS) and progression free survival (PFS) were assessed by Kaplan Meier method. Results: Median age at transplant and M/F sex ratio were 45 years (range, 19-61) and 195/150, respectively. Cytogenetics was intermediate-1, intermediate-2 and unfavorable in 24, 164, 144 patients according to ELN classification, respectively. 82.5% of the patients had reached CR after one cycle of induction. 193 (53%) patients presented infections during induction and 46 (17%) during consolidations. Moreover, 110 (30%) patients suffered from drug toxicity during induction and 36 (10%) during consolidations. Allo-HSCT was performed in all patients after one to three cycles of consolidation. 132 (36%) patients received reduced intensity conditioning. Sex matching was female in male for 74 (21%) patients. ABO matching was matched for 187 and mismatched for 158 patients. HLA matching was related for 138 and unrelated for 198 patients. Source of graft was bone marrow, peripheral stem cell and cord blood in 108, 217 and 12 patients, respectively. 181 (47%) patients underwent aGvHD, most frequently skin aGvHD (stage 1-2: 27.5%; stage 3-4: 9.5%). First, we observed a significant increase of incidence of aGvHD (all grades) if skin toxicities occurred during induction (45/57% for no toxicity and toxicities all grades, respectively p=0.07) or consolidations (46/70% for no toxicity and toxicities all grades, respectively p=0.04). Secondly, we observed a significant increase of skin aGvHD in cases of skin toxicities during induction [stage 0/1-2/3-4 skin aGvHD: 66/27/7% and 47/32/21% for no toxicity and toxicities all grades, respectively (p=0.001)] or consolidations [stage 0/1-2/3-4 skin aGvHD: 64/28/8% and 35/35/30% for no toxicity and toxicities all grades, respectively (p<0.003)]. Thirdly, we observed a correlation between infections and skin aGvHD during consolidations [stage 0/1-2/3-4 skin aGVHD: 64/28/8% vs 49/27/24% for no infection and infections, respectively (p=0.002)], and more particularly between skin infections and skin aGVHD [stage 0/1-2/3-4 skin aGVHD: 63/29/8% vs 44/23/33% for no infection and infections, respectively (p=0.003)]. No correlation was found between others types of infections or toxicities and aGvHD. Finally, we observed no impact of infections and/or toxicities on OS and PFS. Conclusion: Skin immune stimulation induced either by infections during consolidations or by drug toxicity during induction and/or consolidations significantly increased the incidence of skin aGvHD. Nevertheless, we found no impact on OS and PFS in our cohort. Our results confirm the participation of inflammatory process in the physiopathology of GvHD. These data should be confirmed in a larger study to determine whether patients with prior infection and/or drug toxicity to allo-HSCT should receive different GvHD prophylaxis strategies. Disclosures Deconinck: PFIZER: Research Funding; ALEXION: Other: Travel for international congress; JANSSEN: Other: Travel for international congress; NOVARTIS: Other: Travel for international congress; LFB loboratory: Consultancy; ROCHE: Research Funding; CHUGAI: Other: Travel for international congress.
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Bousta, Abderrahmane, Sabrina Bondu, Alexandre Houy, Nicolas Cagnard, Carine Lefevre, Delphine Bernard, Marc-Henri Stern, Michaela Fontenay y Olivier Kosmider. "Gene Expression and Alternative Splicing Datasets Analyses of MDS with Ring Sideroblasts Highlight Alternative Branchpoint Usage in Genes Involved in Iron Metabolism and Erythropoiesis". Blood 128, n.º 22 (2 de diciembre de 2016): 1972. http://dx.doi.org/10.1182/blood.v128.22.1972.1972.
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Abstract Introduction SF3B1 hotspot mutations are associated with various cancers like uveal melanoma, chronic lymphocytic leukemia and myelodysplastic syndrome with ring sideroblasts (MDS-RS). These mutations affect RNA splicing by the use of alternative branchpoints resulting in an aberrant 3' splice site (ss) selection. RNA-sequencing (RNA-seq) analyzed to quantify exon-exon junctions identified aberrantly spliced transcripts in target genes, and half of them are predicted to be degraded by non-sense mediated decay. For this reason, target genes in SF3B1-mutated MDS remain partially characterized. In the present study, we performed deep RNA-seq analysis of bone marrow mononuclear cells in low/int-1 MDS with SF3B1 mutations to identify aberrant/cryptic splicing events among up or down-regulated gene sets. Methods SF3B1 MUT MDS (n=21) were compared to 6 SF3B1WT cases and 5 controls. Analysis of RNA-seq read count was performed using the Voom method associated with the Limma empirical Bayes analysis pipeline (https://genomebiology.biomedcentral.com/articles/10.1186/gb-2014-15-2-r29). Up or downregulated gene sets were identified using Gene Set Enrichment Analysis (GSEA, false discovery rate<0.1). Gene expression profiling data (Affymetrix Hu2.0) were also available for 26/27 patient samples. TopHat (v2.0.6) was used to align the reads against the human reference genome Hg19 RefSeq (RNA sequences, GRCh37) downloaded from the UCSC Genome Browser (http://genome.ucsc.edu). Read counts for splicing junctions from junctions.bed TopHat output were considered for a differential analysis using DESeq2. Only alternative acceptor splice sites (two or more 3′ss with junctions to the same 5′ss) and alternative donor splice sites (two or more 5′ss with junctions to the same 3′ss) with P-values ≤10−5 (Benjamini-Hochberg) and absolute Log2 (fold change) ≥1 were considered. Results Principal component analysis (PCA) nicely discriminated controls from patients, and patients according to the presence of a SF3B1 mutation. A set of 6971 genes was differently expressed (P- value<0.05) between SF3B1MUT and SF3B1WT cases and allows unsupervised clustering in two separated groups (Fig. 1). Distinct gene sets also discriminated SF3B1MUT or SF3B1WT from controls. Consistent with increased amount of erythroblasts in MDS-RS bone marrows, a set of erythroid genes including several genes involved in hemebiosynthesis pathway (ALAD, UROS, ALAS2, UROD) was significantly enriched in SF3B1MUT samples. Genes selected for their involvement in the core iron-sulfur cluster mitochondrial machinery (FXN, BOLA3, FDXR, GLRX5, ISCA2, NFS1, ISCU), the iron binding and trafficking (SLC25A38, ABCB10, TFR2, SLC25A37, ABCB6, FAM132B, SLC25A39, FTH1) and the cellular iron homeostasis (ACO1, ACO2, GLRX3) were also significantly enriched (FDR<10% and nominal P-value<0.05) when input in GSEA. Moreover, other enriched gene sets were G2M checkpoint, MYC targets, oxidative phosphorylation and E2F targets. All of these observations were similarly obtained when analyzing Affymetrix data. Furthermore SF3B1MUT samples with a K700E substitution harbored a specific pattern of deregulated genes, which allowed the ordering of SF3B1MUT samples according to the type of substitution. As previously reported by AlsafadiS et al (2016), analysis of splice junctions using DESeq2 revealed an overall high level of differences between SF3B1MUT and SF3B1WTsamples. Among more than 540 differentially spliced junctions, more than 80% involved an aberrant acceptor (3'ss) site. As determined by PCA, the top 50 genes associated with relevant aberrant junctions were linked to iron metabolism or erythropoiesis and differentially expressed between SF3B1MUT and SF3B1WTsamples. Conclusion In this study, we combined robust analyses of gene expression and aberrantly spliced transcript expression in MDS with SF3B1 mutation. By comparing SF3B1MUTversus SF3B1WT samples, we identified a set of deregulated genes in which both normally and aberrantly spliced transcripts were detected that could contribute to the physiopathology of MDS-RS. Figure 1 Hierarchical clustering and heatmapshowing differentially expressed genes (P-value<0.05) between SF3B1MUT (n=21, black) and SF3B1WT samples (n=6, grey) Ref. Alsafadi S et al. Nat Commun. 2016 Feb 4;7:10615. Figure 1. Hierarchical clustering and heatmapshowing differentially expressed genes (P-value<0.05) between SF3B1MUT (n=21, black) and SF3B1WT samples (n=6, grey) Ref. Alsafadi S et al. Nat Commun. 2016 Feb 4;7:10615. Disclosures No relevant conflicts of interest to declare.
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Duployez, Nicolas, Elise Labis, Alice Marceau-Renaut, Christine Ragu, Arnaud Petit, Anne Auvrignon, Christophe Roumier et al. "Genomic Landscape of Pediatric CBF-AML By SNP-Array Karyotyping and Extensive Mutational Analysis". Blood 124, n.º 21 (6 de diciembre de 2014): 1007. http://dx.doi.org/10.1182/blood.v124.21.1007.1007.
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Abstract Background. Core binding factor (CBF) acute myeloid leukemia (AML) includes AML with t(8;21) and inv(16) leading to RUNX1-RUNX1T1 or CBFB-MYH11 fusion genes. These recurrent genetic abnormalities are both associated with disruption of genes encoding subunits of the CBF, a heterodimeric transcription factor involved in hematopoiesis. Although the fusion proteins appear to be crucial for the leukemogenic process, considerable experimental evidence indicates that they are not sufficient to induce AML on their own. Due to their high sensitivity to chemotherapy with high complete remission rates and their relatively favorable outcome, CBF-AML is considered to have a good prognosis. Nonetheless, about 30-40% of these patients relapse after standard intensive chemotherapy. In this context, identification of additional genetic or molecular abnormalities could allow better understanding of CBF-AML leukemogenesis, prediction of clinical outcome and identification of novel therapeutic targets. Methods. This study focuses on 73 patients with CBF-AML [43 t(8;21) and 30 inv(16)-AML] enrolled in the pediatric trial ELAM02. Single nucleotide polymorphism array (SNP-A) was performed for all patients using Cytoscan® HD arrays according to the manufacturer instructions. In order to distinguish somatic from constitutional SNP-A lesions, we excluded known copy number abnormalities (CNA) if there was >50% overlap with variants from public database, except for breakpoints-related alterations. Interstitial uniparental disomies (UPD) <10 Mb and telomeric UPD <5 Mb were considered as constitutional and excluded from the analysis. Additionally, extensive mutational analysis (including 45 genes frequently reported to be mutated in myeloid malignancies) were performed for 37 and 25 patients with t(8;21) and inv(16)-AML respectively. Two different technologies of next generation sequencing (NGS) were used, allowing direct validation. Results. Among the 73 cases, 145 SNP-A lesions were found in 58 patients (81%) with a median of 2 lesions per case (range, 0-8). CNA was more frequent (84 losses, 47 gains) than UPD (n=14). No significant difference was noted between the number of CNA and UPD in inv(16) and t(8;21)-AML. Small lesions were common at breakpoints involved in the t(8;21) and inv(16) (respectively 4/43 and 6/30). Additional recurrent CNA mostly involved entire chromosomes, chromosomal arms or large chromosomal regions. Del(9q) and loss of sex chromosome were restricted to t(8;21)-AML (respectively 6/43 and 20/43). Trisomy 22 was restricted to inv(16)-AML (2/30). Other recurrent CNA included trisomy 8 (3/43 vs 1/30) and gains of 13q (2/43 vs 1/30) in both subtypes, gains of 1q and del(2q) in t(8;21)-AML (each 2/43). Del(7q) was among the most common aberrations regardless of subtype (7/43 and 7/30). The minimally deleted region of 7q contained 57 genes including MLL3 and EZH2. Additionally, we found focal deletions of IKZF1 in one patient, NF1 in another and 3 deletions of CCDC26. Except for known mutations (KIT, RAS, FLT3), NGS did not reveal any other alterations in inv(16)-AML. By contrast, t(8;21)-AML was marked by the frequency of mutations in ASXL1/2 (8%/24%) and cohesin genes SMC1A, SMC3, RAD21, STAG2, NIPBL (27% combined). Mutations were also detected in epigenetic-related genes EZH2 (5%), TET2 (8%), IDH1/2 (5%) and WT1(11%). Conclusions. SNP-A karyotyping of 73 pediatric CBF-AML revealed several recurrent alterations, with differing distribution between the 2 subgroups. Moreover, t(8;21) and inv(16)-AML appeared to have distinct mutational profiles, leading us to consider them separately for future studies. We recently reported high frequency of ASXL mutations in t(8;21)-AML and their absence in inv(16)-AML (Micol, Duployez and Boissel et al, Blood 2014). We now report high frequency of mutations in cohesin genes with the same distribution. Recent description of functional relations between cohesin and polycomb proteins, together with our results, suggest an important pathway in t(8;21) leukemogenesis. Concurrent ASXL and cohesin mutations were found in several patients, suggesting they could cooperate in some cases. Interestingly, ASXL mutations were exclusive of del(7q), suggesting that disruption of the ASXL-associated proteins MLL3 and EZH2 could be of great interest in the physiopathology of t(8;21)-AML. Finally, correlations with clinical outcome are in progress. Disclosures No relevant conflicts of interest to declare.
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Martin, Mickaël, Anne Marie Knapp, Dana Ghergus, Fabien Delmotte, Laurent Vallat, Cédric Schleiss, Raoul Herbrecht et al. "ZAP-70 Expression in Non Tumoral B Cells: Role in B Tolerance Breakdown?" Blood 132, Supplement 1 (29 de noviembre de 2018): 1114. http://dx.doi.org/10.1182/blood-2018-99-118237.
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Abstract Abnormal expression of the tyrosine kinase ZAP-70 by tumoral B cells in chronic lymphocytic leukemia (CLL) is associated with bad prognosis, related to B cell receptor (BCR) hypersignalling, clonal expansion and autoimmune cytopenia (AIC) occurrence, these latter being mostly induced by polyclonal IgG from the residual non tumoral B cells. We previously shown that ZAP-70 is expressed by these non tumoral B cells in CLL, positively associated with its expression in CLL B cells and with AIC occurrence (Ghergus et al. Poster ASH 2017). Here, we show for the first time a potential role of ZAP-70 expression in tolerance breakdown in CLL and in an original knock in mouse model overexpressing ZAP-70 conditionally in B cells. First, to assess a potential molecular link between ZAP-70+ CLL and non tumoral B cells, an analysis of their BCR repertoire has performed on FACS-sorting CD19+CD5-IgM-IgD- (non tumoral) and CD19+CD5+IgM-IgD- (tumoral) single B cells from blood samples of CLL patients with AIC. ZAP-70 positivity was screened by RT-PCR, and variable regions of heavy (IGVH) and light (IGVK/VL) immunoglobulin genes amplified by RT-PCR on ZAP-70+ cells. To date, analysis of 24 BCR sequences from 7 patients showed that non tumoral ZAP-70+ B cells were polyclonal, without stereotypy, using different V(D)J and CDR3 in comparison with those of the corresponding CLL B cells. IGVH of non tumoral ZAP-70+ B were mostly mutated, of replacement type, suggesting antigenic contact, contrary to CLL B cells. To determine potential autoreactivity of the non-tumoral ZAP-70+ B cells, IGVH and corresponding IGVK/VL were amplified for production of recombinant antibodies (rAb). To date, among 17 rAB from 7 different patients, 2/13 (15.4%) have an antinuclear autoreactivity on HEp-2 cells and 4/17 (23.5%) were polyreactive on ELISA (DNA, lipopolysaccharide, insulin), compared respectively to 6% and 4,3% of control B cells (Wardemann et al., Science 2003). Production of 7 additional rAb and tests for anti-erythrocytes and anti-platelets reactivity are in process. To study functional consequences of early ZAP-70 expression in B cells in vivo, we generated a knock in Zap-70+/Mb1-Cre+mouse model (KI ZAP), to induce conditional expression of ZAP-70 in the B cell compartment from the proB stage, with KI Zap-70+/Mb1-Cre-mice as controls (CTRL). The ZAP-70 mRNAs levels in B cells from KI ZAP mice were on average 20 times higher than that in CTRL B cells. Up to 20 months-old, KI ZAP mice did not develop signs of lymphoproliferation. KI ZAP mice had hypo-IgG since 16 weeks-old (p<0.001) together with hypo-IgM from 14 months-old (p<0.01). Immunophenotyping revealed a reduction in mature naive, mature switched as well as in germinal center B cells (p<0.001, p=0.002 and p<0.01 respectively) and a trend for plasma cells (p=0.07). Microarrays showed enrichment in circulating IgG and IgM autoantibodies against various antigens in KI ZAP mice. These mice had reduced apoptosis rates of proB (p<0.01), preB (p=0.02), and immatures B cells (p=0.03), together with enrichment in marginal zone (p=0.01), trend for transitional T2/T3, and reduction in B1a cells (p<0.01). After immunization by ovalbumin + Freund's adjuvant, a reduced production of specific IgG and IgM was observed (p=0.01 and p=0.03 respectively) with a trend in decreased number of antibody-secreting cells (p=0.07). KI ZAP B cells shown increased spontaneous activation and proliferation levels holding after BCR stimulation (p<0.01), as well as an increased intracellular calcic flow (p<0.001). Preliminary data suggested a reduced SYK phosphorylation after BCR stimulation in KI ZAP B cells. Our findings highlight for the first time that non tumoral B cells ZAP-70+ are distinct from CLL cells at cellular level, but probably enriched in autoreactive cells. Moreover, we shown that early ZAP-70 expression in normal B cells in vivois associated with autoimmune characteristics, together with partial block in B cells peripheral maturation, and a conversely early increased activation and proliferation status. ZAP-70 could interfere early with SYK leading to an altered BCR signaling responsible for defect in normal B maturation promoting emergence of autoreactive B cells. Mechanistic role of ZAP-70 in BCR signaling has to be further analyzed but our data open new opportunities involving ZAP-70 in the understanding of B cell development and physiopathology of tolerance breakdown. Disclosures No relevant conflicts of interest to declare.
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Fattizzo, Bruno, Jessica Rosa, Juri Alessandro Giannotta, Luca Baldini y Nicola Stefano Fracchiolla. "The Physiopathology of T- Cell Acute Lymphoblastic Leukemia: Focus on Molecular Aspects". Frontiers in Oncology 10 (28 de febrero de 2020). http://dx.doi.org/10.3389/fonc.2020.00273.
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Arenas, Víctor, Jose Luis Castaño, Juan José Domínguez-García, Lucrecia Yáñez y Carlos Pipaón. "A Different View for an Old Disease: NEDDylation and Other Ubiquitin-Like Post-Translational Modifications in Chronic Lymphocytic Leukemia". Frontiers in Oncology 11 (23 de septiembre de 2021). http://dx.doi.org/10.3389/fonc.2021.729550.
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Despite the enormous amount of molecular data obtained over the years, the molecular etiology of chronic lymphocytic leukemia (CLL) is still largely unknown. All that information has enabled the development of new therapeutic approaches that have improved life expectancy of the patients but are still not curative. We must increase our knowledge of the molecular alterations responsible for the characteristics common to all CLL patients. One of such characteristics is the poor correlation between mRNA and protein expression, that suggests a role of post-translational mechanisms in CLL physiopathology. Drugs targeting these processes have indeed demonstrated an effect either alone or in combination with other aimed at specific pathways. A recent article unveiled an increment in ubiquitin-like modifications in CLL, with many protein members of relevant pathways affected. Interestingly, the inhibition of the NEDD8-activating protein NAE reverted a substantial number of those modifications. The present review gets the scarce data published about the role of NEDDylation in CLL together and establishes connections to what is known from other neoplasias, thus providing a new perspective to the underlying mechanisms in CLL.
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Marselli, L., L. Trincavelli, C. Santangelo, R. Lupi, S. Del Guerra, U. Boggi, A. Falleni et al. "The role of peripheral benzodiazepine receptors on the function and survival of isolated human pancreatic islets". European Journal of Endocrinology, 1 de agosto de 2004, 207–14. http://dx.doi.org/10.1530/eje.0.1510207.
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OBJECTIVE: Peripheral benzodiazepine receptors (PBRs) are part of the mitochondrial permeability transition pore, and their activation may induce cell death. PBRs are expressed in human pancreatic islets, and cytokine-induced damage is accompanied by changes in their properties. We hypothesized that PBRs can have a role in human islet physiopathology, and evaluated the effects of prolonged exposure to two specific PBR ligands, PK11195 and Ro5-4864 on the function and survival of isolated human islets. DESIGN: Isolated human islets were prepared from the pancreas of 25 multiorgan cadaveric donors and incubated for 12 h in the presence of PK11195 or Ro5-4864. Insulin secretion studies and apoptosis experiments were then performed, together with assessment of intracellular pathways involved in islet cell function and survival. METHODS: Islets were prepared by enzymatic digestion and density gradient purification. Insulin secretion was assessed by the batch incubation method, and glucose oxidation was evaluated by the use of D-[U-(14)C]glucose. Apoptosis was studied using the TUNEL technique, ELISA methods, and electron microscopy evaluation. PCR experiments were performed by the use of specific primers. RESULTS: Glucose-stimulated insulin release was significantly lower after exposure to PK11195 than after exposure to Ro5-4864. This was accompanied by reduced glucose oxidation and no major change of insulin or GLUT-1 mRNA expression. Apoptosis was higher in PK11195-exposed islets, and electron microscopy demonstrated the involvement of beta-cells. The apoptotic effects were prevented by bongkrekic acid and low-dose cyclosporin A, which stabilize the mitochondrial membrane, and were associated with no evident change of inducible nitric oxide synthase (iNOS), B-cell leukemia/lymphoma-2 (Bcl-2) or Bcl-2-associated X protein (Bax) expression. Caspase inhibition markedly reduced the amount of apoptosis, and the role of these proteases was confirmed by the increased activity of caspase-3 and caspase-9. CONCLUSIONS: Prolonged binding to PBRs may cause human beta-cells functional damage and apoptosis, a phenomenon which is prevented by stabilizing the mitochondrial membrane; occurs without changes of iNOS, Bax and Bcl-2 mRNA expression; and involves caspase activation. These results suggest an involvement of PBRs in human pancreatic beta-cell function and survival.