Mécanismes de transformation, sélection clonale et sous-clonale dans les néoplasmes myéloprolifératifs (JAK2V617F) après inactivation de TP53

Hematopoiesis is the production of all of the cellular components of blood. It occurs within the hematopoietic system, including organs and tissues such as the bone marrow, liver, and spleen. It begins early in the development of an embryo, well before birth, and continues all the life of an individual. During adult life, hematopoiesis occurs in the marrow of the long bones like femur and tibia, but mainly in the pelvis, cranium, vertebrae, and sternum. Hematopoiesis represents approximately 5% of total body mass in human healthy adult, 3% of total body mass in rat and 2% in dogs. In pathological conditions, the lymph nodes, liver, and spleen may regain their hematopoietic function that is called extramedullary hematopoiesis (EMH). Pathological hematopoiesis may show alterations in the number and morphology of different cell lineages in peripheral blood and mono- or multilineage cell changes in the bone marrow. Both normal and pathological hematopoiesis can be divided into two different systems, myeloid and lymphoid, according to the different lineages. Myeloid neoplasms include acute myeloid leukemia (AML) and related aggressive myeloid neoplasms, myeloproliferative neoplasms (MPN), mastocytosis, myelodysplastic syndromes (MDS), and MDS/MPN syndromes. JAK2V617F is one of the most common mutations expressed in myeloid neoplasms and more particularly in MPN where this mutation has been demonstrated as causative of the disorders. However, other genes can be abnormal in myeloid neoplasms. TP53 which encodes the tumor suppressor P53 is the most frequently mutated gene in cancers. Surprisingly, it is not one of the most mutated genes in myeloid neoplasms, however it has been suggested that TP53 could be associated to the evolution of MPN and MDS to acute leukemias. The aim of this work was first to better understand the clonal advantage of JAK2V617F mutated cells based on the hypothesis that instead of steady state JAK2V617F hematopoiesis the clonal advantage may be due to stressed JAK2V617F stem cells, and second to better delineate what is TP53 dependent or independent in MPN and verify if the TP53 inactivation can transform the MPN into AML.L'hématopoïèse est la production de tous les composants cellulaires du sang. Il se produit dans le système hématopoïétique, y compris les organes et les tissus tels que la moelle osseuse, le foie et la rate. Il commence tôt dans le développement d'un embryon, bien avant la naissance, et se poursuit toute la vie d'un individu. Au cours de la vie adulte, l'hématopoïèse se produit dans la moelle des os longs comme le fémur et le tibia, mais principalement dans le bassin, le crâne, les vertèbres et le sternum. L'hématopoïèse représente environ 5% de la masse corporelle totale chez l'homme adulte en bonne santé, 3% de la masse corporelle totale chez le rat et 2% chez le chien. Dans des conditions pathologiques, les ganglions lymphatiques, le foie et la rate peuvent retrouver leur fonction hématopoïétique appelée hématopoïèse extramédullaire (HEM). L'hématopoïèse pathologique peut montrer des altérations du nombre et de la morphologie des différentes lignées cellulaires dans le sang périphérique et des modifications cellulaires mono- ou multilignées dans la moelle osseuse. L'hématopoïèse normale et pathologique peut être divisée en deux systèmes différents, myéloïde et lymphoïde, selon les différentes lignées. Les néoplasmes myéloïdes comprennent la leucémie myéloïde aiguë (LMA) et les néoplasmes myéloïdes agressifs associés, les néoplasmes myéloprolifératifs (NPP), la mastocytose, les syndromes myélodysplasiques (SMD) et les syndromes MDS / MPN. JAK2V617F est l'une des mutations les plus courantes exprimées dans les néoplasmes myéloïdes et plus particulièrement dans le MPN où cette mutation a été démontrée comme causale des troubles. Cependant, d'autres gènes peuvent être anormaux dans les néoplasmes myéloïdes. TP53 qui code pour le suppresseur de tumeur P53 est le gène le plus fréquemment muté dans les cancers. Étonnamment, ce n'est pas l'un des gènes les plus mutés dans les néoplasmes myéloïdes, mais il a été suggéré que TP53 pourrait être associé à l'évolution du MPN et du SMD en leucémies aiguës. Le but de ce travail était d'abord de mieux comprendre l'avantage clonal des cellules mutées JAK2V617F en partant de l'hypothèse qu'au lieu de l'hématopoïèse de JAK2V617F à l'état d'équilibre, l'avantage clonal peut être dû à des cellules souches JAK2V617F stressées, et deuxièmement de mieux délimiter ce qui dépend de TP53 ou indépendant en MPN et vérifiez si l'inactivation du TP53 peut transformer le MPN en AML

Mécanismes de transformation, sélection clonale et sous-clonale dans les néoplasmes myéloprolifératifs (JAK2V617F) après inactivation de TP53

Hematopoiesis is the production of all of the cellular components of blood. It occurs within the hematopoietic system, including organs and tissues such as the bone marrow, liver, and spleen. It begins early in the development of an embryo, well before birth, and continues all the life of an individual. During adult life, hematopoiesis occurs in the marrow of the long bones like femur and tibia, but mainly in the pelvis, cranium, vertebrae, and sternum. Hematopoiesis represents approximately 5% of total body mass in human healthy adult, 3% of total body mass in rat and 2% in dogs. In pathological conditions, the lymph nodes, liver, and spleen may regain their hematopoietic function that is called extramedullary hematopoiesis (EMH). Pathological hematopoiesis may show alterations in the number and morphology of different cell lineages in peripheral blood and mono- or multilineage cell changes in the bone marrow. Both normal and pathological hematopoiesis can be divided into two different systems, myeloid and lymphoid, according to the different lineages. Myeloid neoplasms include acute myeloid leukemia (AML) and related aggressive myeloid neoplasms, myeloproliferative neoplasms (MPN), mastocytosis, myelodysplastic syndromes (MDS), and MDS/MPN syndromes. JAK2V617F is one of the most common mutations expressed in myeloid neoplasms and more particularly in MPN where this mutation has been demonstrated as causative of the disorders. However, other genes can be abnormal in myeloid neoplasms. TP53 which encodes the tumor suppressor P53 is the most frequently mutated gene in cancers. Surprisingly, it is not one of the most mutated genes in myeloid neoplasms, however it has been suggested that TP53 could be associated to the evolution of MPN and MDS to acute leukemias. The aim of this work was first to better understand the clonal advantage of JAK2V617F mutated cells based on the hypothesis that instead of steady state JAK2V617F hematopoiesis the clonal advantage may be due to stressed JAK2V617F stem cells, and second to better delineate what is TP53 dependent or independent in MPN and verify if the TP53 inactivation can transform the MPN into AML.L'hématopoïèse est la production de tous les composants cellulaires du sang. Il se produit dans le système hématopoïétique, y compris les organes et les tissus tels que la moelle osseuse, le foie et la rate. Il commence tôt dans le développement d'un embryon, bien avant la naissance, et se poursuit toute la vie d'un individu. Au cours de la vie adulte, l'hématopoïèse se produit dans la moelle des os longs comme le fémur et le tibia, mais principalement dans le bassin, le crâne, les vertèbres et le sternum. L'hématopoïèse représente environ 5% de la masse corporelle totale chez l'homme adulte en bonne santé, 3% de la masse corporelle totale chez le rat et 2% chez le chien. Dans des conditions pathologiques, les ganglions lymphatiques, le foie et la rate peuvent retrouver leur fonction hématopoïétique appelée hématopoïèse extramédullaire (HEM). L'hématopoïèse pathologique peut montrer des altérations du nombre et de la morphologie des différentes lignées cellulaires dans le sang périphérique et des modifications cellulaires mono- ou multilignées dans la moelle osseuse. L'hématopoïèse normale et pathologique peut être divisée en deux systèmes différents, myéloïde et lymphoïde, selon les différentes lignées. Les néoplasmes myéloïdes comprennent la leucémie myéloïde aiguë (LMA) et les néoplasmes myéloïdes agressifs associés, les néoplasmes myéloprolifératifs (NPP), la mastocytose, les syndromes myélodysplasiques (SMD) et les syndromes MDS / MPN. JAK2V617F est l'une des mutations les plus courantes exprimées dans les néoplasmes myéloïdes et plus particulièrement dans le MPN où cette mutation a été démontrée comme causale des troubles. Cependant, d'autres gènes peuvent être anormaux dans les néoplasmes myéloïdes. TP53 qui code pour le suppresseur de tumeur P53 est le gène le plus fréquemment muté dans les cancers. Étonnamment, ce n'est pas l'un des gènes les plus mutés dans les néoplasmes myéloïdes, mais il a été suggéré que TP53 pourrait être associé à l'évolution du MPN et du SMD en leucémies aiguës. Le but de ce travail était d'abord de mieux comprendre l'avantage clonal des cellules mutées JAK2V617F en partant de l'hypothèse qu'au lieu de l'hématopoïèse de JAK2V617F à l'état d'équilibre, l'avantage clonal peut être dû à des cellules souches JAK2V617F stressées, et deuxièmement de mieux délimiter ce qui dépend de TP53 ou indépendant en MPN et vérifiez si l'inactivation du TP53 peut transformer le MPN en AML

Mécanismes de transformation, sélection clonale et sous-clonale dans les néoplasmes myéloprolifératifs (JAK2V617F) après inactivation de TP53

L'hématopoïèse est la production de tous les composants cellulaires du sang. Il se produit dans le système hématopoïétique, y compris les organes et les tissus tels que la moelle osseuse, le foie et la rate. Il commence tôt dans le développement d'un embryon, bien avant la naissance, et se poursuit toute la vie d'un individu. Au cours de la vie adulte, l'hématopoïèse se produit dans la moelle des os longs comme le fémur et le tibia, mais principalement dans le bassin, le crâne, les vertèbres et le sternum. L'hématopoïèse représente environ 5% de la masse corporelle totale chez l'homme adulte en bonne santé, 3% de la masse corporelle totale chez le rat et 2% chez le chien. Dans des conditions pathologiques, les ganglions lymphatiques, le foie et la rate peuvent retrouver leur fonction hématopoïétique appelée hématopoïèse extramédullaire (HEM). L'hématopoïèse pathologique peut montrer des altérations du nombre et de la morphologie des différentes lignées cellulaires dans le sang périphérique et des modifications cellulaires mono- ou multilignées dans la moelle osseuse. L'hématopoïèse normale et pathologique peut être divisée en deux systèmes différents, myéloïde et lymphoïde, selon les différentes lignées. Les néoplasmes myéloïdes comprennent la leucémie myéloïde aiguë (LMA) et les néoplasmes myéloïdes agressifs associés, les néoplasmes myéloprolifératifs (NPP), la mastocytose, les syndromes myélodysplasiques (SMD) et les syndromes MDS / MPN. JAK2V617F est l'une des mutations les plus courantes exprimées dans les néoplasmes myéloïdes et plus particulièrement dans le MPN où cette mutation a été démontrée comme causale des troubles. Cependant, d'autres gènes peuvent être anormaux dans les néoplasmes myéloïdes. TP53 qui code pour le suppresseur de tumeur P53 est le gène le plus fréquemment muté dans les cancers. Étonnamment, ce n'est pas l'un des gènes les plus mutés dans les néoplasmes myéloïdes, mais il a été suggéré que TP53 pourrait être associé à l'évolution du MPN et du SMD en leucémies aiguës. Le but de ce travail était d'abord de mieux comprendre l'avantage clonal des cellules mutées JAK2V617F en partant de l'hypothèse qu'au lieu de l'hématopoïèse de JAK2V617F à l'état d'équilibre, l'avantage clonal peut être dû à des cellules souches JAK2V617F stressées, et deuxièmement de mieux délimiter ce qui dépend de TP53 ou indépendant en MPN et vérifiez si l'inactivation du TP53 peut transformer le MPN en AML.Hematopoiesis is the production of all of the cellular components of blood. It occurs within the hematopoietic system, including organs and tissues such as the bone marrow, liver, and spleen. It begins early in the development of an embryo, well before birth, and continues all the life of an individual. During adult life, hematopoiesis occurs in the marrow of the long bones like femur and tibia, but mainly in the pelvis, cranium, vertebrae, and sternum. Hematopoiesis represents approximately 5% of total body mass in human healthy adult, 3% of total body mass in rat and 2% in dogs. In pathological conditions, the lymph nodes, liver, and spleen may regain their hematopoietic function that is called extramedullary hematopoiesis (EMH). Pathological hematopoiesis may show alterations in the number and morphology of different cell lineages in peripheral blood and mono- or multilineage cell changes in the bone marrow. Both normal and pathological hematopoiesis can be divided into two different systems, myeloid and lymphoid, according to the different lineages. Myeloid neoplasms include acute myeloid leukemia (AML) and related aggressive myeloid neoplasms, myeloproliferative neoplasms (MPN), mastocytosis, myelodysplastic syndromes (MDS), and MDS/MPN syndromes. JAK2V617F is one of the most common mutations expressed in myeloid neoplasms and more particularly in MPN where this mutation has been demonstrated as causative of the disorders. However, other genes can be abnormal in myeloid neoplasms. TP53 which encodes the tumor suppressor P53 is the most frequently mutated gene in cancers. Surprisingly, it is not one of the most mutated genes in myeloid neoplasms, however it has been suggested that TP53 could be associated to the evolution of MPN and MDS to acute leukemias. The aim of this work was first to better understand the clonal advantage of JAK2V617F mutated cells based on the hypothesis that instead of steady state JAK2V617F hematopoiesis the clonal advantage may be due to stressed JAK2V617F stem cells, and second to better delineate what is TP53 dependent or independent in MPN and verify if the TP53 inactivation can transform the MPN into AML

Insights into the Potential Mechanisms of JAK2V617F Somatic Mutation Contributing Distinct Phenotypes in Myeloproliferative Neoplasms

Myeloproliferative neoplasms (MPN) are a group of blood cancers in which the bone marrow (BM) produces an overabundance of erythrocyte, white blood cells, or platelets. Philadelphia chromosome-negative MPN has three subtypes, including polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF). The over proliferation of blood cells is often associated with somatic mutations, such as JAK2, CALR, and MPL. JAK2V617F is present in 95% of PV and 50–60% of ET and PMF. Based on current molecular dynamics simulations of full JAK2 and the crystal structure of individual domains, it suggests that JAK2 maintains basal activity through self-inhibition, whereas other domains and linkers directly/indirectly enhance this self-inhibited state. Nevertheless, the JAK2V617F mutation is not the only determinant of MPN phenotype, as many normal individuals carry the JAK2V617F mutation without a disease phenotype. Here we review the major MPN phenotypes, JAK-STAT pathways, and mechanisms of development based on structural biology, while also describing the impact of other contributing factors such as gene mutation allele burden, JAK-STAT-related signaling pathways, epigenetic modifications, immune responses, and lifestyle on different MPN phenotypes. The cross-linking of these elements constitutes a complex network of interactions and generates differences in individual and cellular contexts that determine the phenotypic development of MPN

Etoposide, an anticancer drug involved in therapy-related secondary leukemia: Enzymes at play

International audienceEtoposide is a semi-synthetic glycoside derivative of podophyllotoxin, also known as VP-16. It is a widely used anticancer medicine in clinics. Unfortunately, high doses or long-term etoposide treatment can induce therapyrelated leukemia. The mechanism by which etoposide induces secondary hematopoietic malignancies is still unclear. In this article, we review the potential mechanisms of etoposide induced therapy-related leukemia. Etoposide related leukemogenesis is known to depend on reactive oxidative metabolites of etoposide, notably etoposide quinone, which interacts with cellular proteins such as topoisomerases II (TOP2), CREB-binding protein (CREBBP), and T-Cell Protein Tyrosine Phosphatase (TCPTP). CYP3A4 and CYP3A5 metabolize etoposide to etoposide catechol, which readily oxidizes to etoposide quinone. As a poison of TOP2 enzymes, etoposide and its metabolites induce DNA double-stranded breaks (DSB), and the accumulation of DSB triggers cell apoptosis. If the cell survives, the DSB gives rise to the likelihood of faulty DNA repair events. The gene translocation could occur in mixed-lineage leukemia (MLL) gene, which is well-known in leukemogenesis. Recently, studies have revealed that etoposide metabolites, especially etoposide quinone, can covalently bind to cysteines residues of CREBBP and TCPTP enzymes, . This leads to enzyme inhibition and further affects histone acetylation and phosphorylation of the JAK-STAT pathway, thus putatively altering the proliferation and differentiation of hematopoietic stem cells (HSC). In brief, current studies suggest that etoposide and its metabolites contribute to etoposide therapy-related leukemia through TOP2 mediated DSB and impairs specific enzyme activity, such as CREBBP and TCPTP

Combined Biological and Modeling Approach of Hematopoiesis: From Native to Stressed Erythropoiesis

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Multistage hematopoietic stem cell regulation in the mouse: A combined biological and mathematical approach

International audienceWe have reconciled steady-state and stress hematopoiesis in a single mathematicalmodel based on murine in vivo experiments and with a focus on hematopoieticstem and progenitor cells. A phenylhydrazine stress was first applied tomice. A reduced cell number in each progenitor compartment was evidenced duringthe next 7 days through a drastic level of differentiation without proliferation,followed by a huge proliferative response in all compartments including longtermhematopoietic stem cells, before a return to normal levels. Data analysisled to the addition to the 6-compartment model, of time-dependent regulationthat depended indirectly on the compartment sizes. The resulting model wasfinely calibrated using a stochastic optimization algorithm and could reproducebiological data in silico when applied to different stress conditions (bleeding,chemotherapy, HSC depletion). In conclusion, our multi-step and time-dependentmodel of immature hematopoiesis provides new avenues to a better understandingof both normal and pathological hematopoiesis

Genomic and functional impact of Trp53 inactivation in JAK2V617F myeloproliferative neoplasms

International audienceAbstract Classical myeloproliferative neoplasms (MPNs) are characterized by the proliferation of myeloid cells and the risk of transformation into myelofibrosis or acute myeloid leukemia (AML) and TP53 mutations in MPN patients are linked to AML. However, JAK2V617F has been reported to impact the TP53 response to DNA damage, suggesting potential overlapping role of TP53 inactivation in MPN. We established a mouse model showing that JAK2V617F/Vav-Cre/Trp53 −/− mice displayed a similar phenotype to JAK2V617F/Vav-Cre mice, but their proliferation was outcompeted in competitive grafts. RNA-Seq revealed that half of the genes affected by JAK2V617F were affected by p53-inactivation, including the interferon pathway. To validate this finding, mice were repopulated with a mixture of wild-type and JAK2V617F (or JAK2V617F/Vav-Cre/Trp53 −/− ) cells and treated with pegylated interferonα. JAK2V617F-reconstituted mice entered complete hematological remission, while JAK2V617F/Vav-Cre /Trp53 −/− -reconstituted mice did not, confirming that p53 loss induced interferon-α resistance. KEGG and Gene Ontology analyses of common deregulated genes showed that these genes were mainly implicated in cytokine response, proliferation, and leukemia evolution, illustrating that in this mouse model, the development of MPN is not affected by TP53 inactivation. Taken together, our results show that many genetic modifications induced by JAK2V617F are influenced by TP53, the MPN phenotype may not be. Trp53 loss alone is insufficient to induce rapid leukemic transformation in steady-state hematopoiesis in JAK2V617F MPN, and Trp53 loss may contribute to interferon resistance in MPN

Human CREBBP acetyltransferase is impaired by etoposide quinone, an oxidative and leukemogenic metabolite of the anticancer drug etoposide through modification of redox-sensitive zinc-finger cysteine residues

International audienceEtoposide is an extensively prescribed anticancer drug that, unfortunately, causes therapy-related leukemia. The mechanisms by which etoposide induces secondary hematopoietic malignancies are poorly documented. However, etoposide-related leukemogenesis is known to depend on oxidative metabolites of etoposide, notably etoposide quinone, that can react with protein cysteine residues such as in topoisomerases II. CREBBP is a major histone acetyltransferase that functions mainly as a transcriptional co-activator. This epigenetic enzyme is considered as a tumor suppressor that plays a major role in hematopoiesis. Genetic alterations affecting CREBBP activity are highly common in hematopoietic malignancies. We report here that CREBBP is impaired by etoposide quinone. Molecular and kinetic analyses show that this inhibition occurs through the rapid and covalent (kinhib = 16.102 M-1. s-1) adduction of etoposide quinone with redox sensitive cysteine residues within the RING and PHD Zn2+-fingers of CREBBP catalytic core leading to subsequent release of Zn2+. In agreement with these findings, experiments conducted in cells and in mice treated with etoposide showed irreversible inhibition of endogenous CREBBP activity and decreased H3K18 and H3K27 acetylation. As shown for topoisomerases II, our work thus suggests that the leukemogenic metabolite etoposide quinone can impair the epigenetic CREBBP acetyltransferase through reaction with redox sensitive cysteine residues