Leucémie myéloïde chronique

La niche hématopoïétique médullaire est une structure pluricellulaire dont les composantes sont actuellement bien identifiées. Les cellules souches hématopoïétiques y sont notamment en contact intime avec des cellules stromales mésenchymateuses, des ostéoblastes et des cellules endothéliales sinusoïdales. Ces relations étroites sont à l’origine d’un équilibre subtil entre des fonctions cellulaires spécialisées (prolifération et quiescence, différenciation et autorenouvellement), assurant ainsi une hématopoïèse efficace. Chez les patients atteints de leucémie myéloïde chronique (LMC), affection clonale de la cellule souche hématopoïétique, des travaux récents ont montré la présence d’un dialogue entre les progéniteurs myéloïdes malins et les cellules du microenvironnement médullaire. Une niche leucémique faite de signaux altérés semble ainsi s’organiser, permettant tout à la fois une prolifération intense de cellules myéloïdes différenciées quittant la niche vers le sang périphérique et le maintien d’un pool de cellules souches leucémiques. Cette niche médullaire pourrait également protéger les cellules souches de LMC des traitements par inhibiteurs de tyrosine kinase et contribuer ainsi à leur résistance vis-à-vis des thérapies ciblées

The Vascular Niche Is Involved in Regulating Leukemic Stem Cells in Murine Chronic Myelogenous Leukemia

Abstract Chronic myelogenous leukemia (CML) is effectively controlled by tyrosine kinase inhibitors (TKIs) such as imatinib mesylate, leading to a hematologic remission in >90% of patients. However, the majority of patients relapse once TKI therapy is discontinued, suggesting that CML leukemia stem cells (LSC) are not eradicated. We recently showed that modulation of the osteoblastic niche can lead to reduction of LSC in CML (Krause et al., Nat. Med. 2013;19:1513), but the role of the vascular hematopoietic stem cell niche in CML has not been well defined. E-selectin is expressed on bone marrow (BM) endothelium within the vascular niche, whereas loss of E-selectin expression or treatment with GMI-1271, an E-selectin small molecule antagonist, enhances HSC quiescence and self-renewal (Winkler et al., Nat. Med. 2012;18:1651). E-selectin also plays a critical role in the homing and engraftment of CML LSCs (Krause et al., Blood 2014;123:1361) through E-selectin ligands expressed on the LSCs, including CD44 (Krause et al., Nat Med. 2006;12:1175). We, therefore, hypothesized that E-selectin blockade with GMI-1271 may overcome niche-mediated resistance to TKIs and eradicate CML LSC. Using the well-described murine retroviral transduction/transplantation model of CML we showed that the white blood cell count (WBC) of mice with BCR-ABL1-induced CML-like leukemia was significantly reduced by treatment with imatinib plus GMI-1271 (or imatinib alone) and there was a trend towards WBC reduction by treatment with GMI-1271 alone (P=0.07). The percentage of GFP+ Mac-1+ cells in peripheral blood on day 16 post-transplant was decreased by imatinib or GMI-1271 alone or by combined treatment with imatinib and GMI-1271. Spleen weights were significantly reduced by combined treatment with imatinib plus GMI 1271. Furthermore, the BM GFP+ (BCR-ABL1+) Lin– c-Kit+ Sca-1+ population, which contains the LSCs in this model, was significantly reduced in animals treated with GMI-1271 compared to vehicle controls. As expected, treatment with imatinib alone had no effect on BM LSC frequency, and there was no added benefit in the reduction of LSC when imatinib and GMI-1271 were combined. In addition, the survival of mice treated with imatinib plus GMI-1271 was significantly prolonged compared to vehicle-treated animals, with ~20% of mice treated with GMI-1271 alone or the combination of imatinib and GMI-1271 exhibiting long-term low-burden disease despite discontinuation of treatment on day 28 post-transplant. In these primary recipients neither BCR-ABL1+ myeloid cells nor BCR-ABL1+ LSC were mobilized to peripheral organs. However, fewer BCR-ABL1+ LSC were found in the spleen of mice treated with GMI-1271 compared to imatinib-treated mice. There was a significant reduction in the frequency of cycling BCR-ABL1+ LSC in mice treated with GMI-1271 and imatinib. To assess directly the effect of E-selectin inhibition on LSC frequency and function, we transplanted BM from primary leukemic mice treated with vehicle, imatinib, GMI-1271 or the combination of imatinib and GMI-1271 into irradiated secondary recipient mice. There was a significant reduction of WBC and a trend towards reduction of BCR-ABL1+ myeloid cells in secondary recipients of BM from donors treated with GMI-1271 alone or in combination with imatinib, but not by imatinib alone. These data suggest that modulation of the vascular niche and, specifically, inhibition of E-selectin may be a possible strategy to target LSC in CML, possibly via a reduction in S-G2/M as cells arrest prior to apoptosis, even when imatinib is discontinued. Further studies on the effects of GMI-1271 on homing of LSC and the more exact mechanism of LSC reduction by GMI-1271 are being performed. Disclosures Magnani: GlycoMimetics Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Krause:Glycomimetics. Inc.: Research Funding

The Vascular Niche Is Involved in Regulating Leukemic Stem Cells in Murine Chronic Myelogenous Leukemia

Abstract Chronic myelogenous leukemia (CML) is effectively controlled by tyrosine kinase inhibitors (TKIs) such as imatinib mesylate, leading to a hematologic remission in >90% of patients. However, the majority of patients relapse once TKI therapy is discontinued, suggesting that CML leukemia stem cells (LSC) are not eradicated. We recently showed that modulation of the osteoblastic niche can lead to reduction of LSC in CML (Krause et al., Nat. Med. 2013;19:1513), but the role of the vascular hematopoietic stem cell niche in CML has not been well defined. E-selectin is expressed on bone marrow (BM) endothelium within the vascular niche, whereas loss of E-selectin expression or treatment with GMI-1271, an E-selectin small molecule antagonist, enhances HSC quiescence and self-renewal (Winkler et al., Nat. Med. 2012;18:1651). E-selectin also plays a critical role in the homing and engraftment of CML LSCs (Krause et al., Blood 2014;123:1361) through E-selectin ligands expressed on the LSCs, including CD44 (Krause et al., Nat Med. 2006;12:1175). We, therefore, hypothesized that E-selectin blockade with GMI-1271 may overcome niche-mediated resistance to TKIs and eradicate CML LSC. Using the well-described murine retroviral transduction/transplantation model of CML we showed that the white blood cell count (WBC) of mice with BCR-ABL1-induced CML-like leukemia was significantly reduced by treatment with imatinib plus GMI-1271 (or imatinib alone) and there was a trend towards WBC reduction by treatment with GMI-1271 alone (P=0.07). The percentage of GFP+ Mac-1+ cells in peripheral blood on day 16 post-transplant was decreased by imatinib or GMI-1271 alone or by combined treatment with imatinib and GMI-1271. Spleen weights were significantly reduced by combined treatment with imatinib plus GMI 1271. Furthermore, the BM GFP+ (BCR-ABL1+) Lin– c-Kit+ Sca-1+ population, which contains the LSCs in this model, was significantly reduced in animals treated with GMI-1271 compared to vehicle controls. As expected, treatment with imatinib alone had no effect on BM LSC frequency, and there was no added benefit in the reduction of LSC when imatinib and GMI-1271 were combined. In addition, the survival of mice treated with imatinib plus GMI-1271 was significantly prolonged compared to vehicle-treated animals, with ~20% of mice treated with GMI-1271 alone or the combination of imatinib and GMI-1271 exhibiting long-term low-burden disease despite discontinuation of treatment on day 28 post-transplant. In these primary recipients neither BCR-ABL1+ myeloid cells nor BCR-ABL1+ LSC were mobilized to peripheral organs. However, fewer BCR-ABL1+ LSC were found in the spleen of mice treated with GMI-1271 compared to imatinib-treated mice. There was a significant reduction in the frequency of cycling BCR-ABL1+ LSC in mice treated with GMI-1271 and imatinib. To assess directly the effect of E-selectin inhibition on LSC frequency and function, we transplanted BM from primary leukemic mice treated with vehicle, imatinib, GMI-1271 or the combination of imatinib and GMI-1271 into irradiated secondary recipient mice. There was a significant reduction of WBC and a trend towards reduction of BCR-ABL1+ myeloid cells in secondary recipients of BM from donors treated with GMI-1271 alone or in combination with imatinib, but not by imatinib alone. These data suggest that modulation of the vascular niche and, specifically, inhibition of E-selectin may be a possible strategy to target LSC in CML, possibly via a reduction in S-G2/M as cells arrest prior to apoptosis, even when imatinib is discontinued. Further studies on the effects of GMI-1271 on homing of LSC and the more exact mechanism of LSC reduction by GMI-1271 are being performed. Disclosures Magnani: GlycoMimetics Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Krause:Glycomimetics. Inc.: Research Funding

Modeling the influence of stromal microenvironment in the selection of ENU-induced BCR-ABL1 mutants by tyrosine kinase inhibitors

International audienceTyrosine kinase inhibitors (TKIs) have profoundly changed the natural history of chronic myeloid leukemia (CML). However, acquired resistance to imatinib, dasatinib or nilotinib (1(st) and 2(nd) generation TKIs), due in part to BCR-ABL1 kinase mutations, has been largely described. These drugs are ineffective on the T315I gatekeeper substitution, which remains sensitive to 3(rd) generation TKI ponatinib. It has recently been suggested that the hematopoietic niche could protect leukemic cells from targeted therapy. In order to investigate the role of a stromal niche in mutation-related resistance, we developed a niche-based cell mutagenesis assay. For this purpose, ENU (N-ethyl-N-nitrosourea)-exposed UT-7 cells expressing non-mutated or T315I-mutated BCR-ABL1 were cultured with or without murine MS-5 stromal cells and in the presence of imatinib, dasatinib, nilotinib, or ponatinib. In the assays relative to 1(st) and 2(nd) generation TKIs, which were performed on non-mutated BCR-ABL1 cells, our data highlighted the increasing efficacy of the latter, but did not reveal any substantial effect of the niche. In ponatinib assays performed on both non-mutated and T315I-mutated BCR-ABL1 cells, an increased number of resistant clones were observed in the presence of MS-5. Present data suggested that T315I mutants need either compound mutations (e.g. E255K/T315I) or a stromal niche to escape from ponatinib. Using array-comparative genomic hybridization experiments, we found an increased number of variations (involving some recurrent chromosome regions) in clones cultured on MS-5 feeder. Overall, our study suggests that the hematopoietic niche could play a crucial role in conferring resistance to ponatinib, by providing survival signals and favoring genetic instability

Modeling the influence of stromal microenvironment in the selection of ENU-induced BCR-ABL1 mutants by tyrosine kinase inhibitors

Tyrosine kinase inhibitors (TKIs) have profoundly changed the natural history of chronic myeloid leukemia (CML). However, acquired resistance to imatinib, dasatinib or nilotinib (1(st) and 2(nd) generation TKIs), due in part to BCR-ABL1 kinase mutations, has been largely described. These drugs are ineffective on the T315I gatekeeper substitution, which remains sensitive to 3(rd) generation TKI ponatinib. It has recently been suggested that the hematopoietic niche could protect leukemic cells from targeted therapy. In order to investigate the role of a stromal niche in mutation-related resistance, we developed a niche-based cell mutagenesis assay. For this purpose, ENU (N-ethyl-N-nitrosourea)-exposed UT-7 cells expressing non-mutated or T315I-mutated BCR-ABL1 were cultured with or without murine MS-5 stromal cells and in the presence of imatinib, dasatinib, nilotinib, or ponatinib. In the assays relative to 1(st) and 2(nd) generation TKIs, which were performed on non-mutated BCR-ABL1 cells, our data highlighted the increasing efficacy of the latter, but did not reveal any substantial effect of the niche. In ponatinib assays performed on both non-mutated and T315I–mutated BCR-ABL1 cells, an increased number of resistant clones were observed in the presence of MS-5. Present data suggested that T315I mutants need either compound mutations (e.g. E255K/T315I) or a stromal niche to escape from ponatinib. Using array-comparative genomic hybridization experiments, we found an increased number of variations (involving some recurrent chromosome regions) in clones cultured on MS-5 feeder. Overall, our study suggests that the hematopoietic niche could play a crucial role in conferring resistance to ponatinib, by providing survival signals and favoring genetic instability

The downregulation of BAP1 expression by BCR-ABL reduces the stability of BRCA1 in chronic myeloid leukemia.

BCR ABLInternational audienceBCR-ABL induces an intrinsic genetic instability in chronic myeloid leukemia (CML). The protein breast cancer 1, early onset (BRCA1)-associated protein 1 (BAP1) is a deubiquitinase interacting with the DNA repair regulator BRCA1 and is frequently inactivated in many cancers. Here, we report that BAP1 mRNA and protein levels are downregulated in a BCR-ABL1-expressing hematopoietic cell line (UT-7/11). A decrease of BAP1 transcripts is also observed in newly diagnosed CML patients. Moreover, BAP1 protein levels are low or undetectable in CD34(+) cells from CML patients at diagnosis as compared with CD34(+) cells from normal donors. In addition, BRCA1 protein level is reduced in BCR-ABL1-expressing UT-7/11 cells. Finally, the enforced expression of BAP1 is associated with BRCA1 protein deubiquitination and restoration. These results demonstrate BAP1 as a major link with the BCR-ABL-induced downregulation of BRCA1 in CML

Superoxide dismutase 2 (SOD2) contributes to genetic stability of native and T315I-mutated BCR-ABL expressing leukemic cells

International audienceManganese Superoxide dismutase 2 (SOD2) plays a crucial role in antioxidant defense but there are no data suggesting its role in genetic instability in CML. We evaluated the effects of SOD2 silencing in human UT7 cell line expressing either non-mutated or T315I-mutated BCR-ABL. Array-CGH experiments detected in BCR-ABL-expressing cells silenced for SOD2 a major genetic instability within several chromosomal loci, especially in regions carrying the glypican family (duplicated) and β-defensin genes (deleted). In a large cohort of patients with chronic myeloid leukemia (CML), a significant decrease of SOD2 mRNA was observed. This reduction appeared inversely correlated with leukocytosis and Sokal score, high-risk patients showing lower SOD2 levels. The analysis of anti-oxidant gene expression analysis revealed a specific down-regulation of the expression of PRDX2 in UT7-BCR-ABL and UT7-T315I cells silenced for SOD2 expression. Gene set enrichment analysis performed between the two SOD2-dependent classes of CML patients revealed a significant enrichment of Reactive Oxygen Species (ROS) Pathway. Our data provide the first evidence for a link between SOD2 expression and genetic instability in CML. Consequently, SOD2 mRNA levels should be analyzed in prospective studies as patients with low SOD2 expression could be more prone to develop a mutator phenotype under TKI therapies

Superoxide dismutase 2 (SOD2) contributes to genetic stability of native and T315I-mutated BCR-ABL expressing leukemic cells

International audienceManganese Superoxide dismutase 2 (SOD2) plays a crucial role in antioxidant defense but there are no data suggesting its role in genetic instability in CML. We evaluated the effects of SOD2 silencing in human UT7 cell line expressing either non-mutated or T315I-mutated BCR-ABL. Array-CGH experiments detected in BCR-ABL-expressing cells silenced for SOD2 a major genetic instability within several chromosomal loci, especially in regions carrying the glypican family (duplicated) and β-defensin genes (deleted). In a large cohort of patients with chronic myeloid leukemia (CML), a significant decrease of SOD2 mRNA was observed. This reduction appeared inversely correlated with leukocytosis and Sokal score, high-risk patients showing lower SOD2 levels. The analysis of anti-oxidant gene expression analysis revealed a specific down-regulation of the expression of PRDX2 in UT7-BCR-ABL and UT7-T315I cells silenced for SOD2 expression. Gene set enrichment analysis performed between the two SOD2-dependent classes of CML patients revealed a significant enrichment of Reactive Oxygen Species (ROS) Pathway. Our data provide the first evidence for a link between SOD2 expression and genetic instability in CML. Consequently, SOD2 mRNA levels should be analyzed in prospective studies as patients with low SOD2 expression could be more prone to develop a mutator phenotype under TKI therapies

Abstracts of the First International Conference on Advances in Electrical and Computer Engineering 2023

This book presents extended abstracts of the selected contributions to the First International Conference on Advances in Electrical and Computer Engineering (ICAECE'2023), held on 15-16 May 2023 by the Faculty of Science and Technology, Department of Electrical Engineering, University of Echahid Cheikh Larbi Tebessi, Tebessa-Algeria. ICAECE'2023 was delivered in-person and virtually and was open for researchers, engineers, academics, and industrial professionals from around the world interested in new trends and advances in current topics of Electrical and Computer Engineering. Conference Title: First International Conference on Advances in Electrical and Computer Engineering 2023Conference Acronym: ICAECE'2023Conference Date: 15-16 May 2023Conference Venue: University of Echahid Cheikh Larbi Tebessi, Tebessa-AlgeriaConference Organizer: Faculty of Science and Technology, Department of Electrical Engineering, University of Echahid Cheikh Larbi Tebessi, Tebessa-Algeri