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1.
Nature ; 630(8015): 198-205, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38720074

RESUMO

Phosphoinositide-3-kinase-γ (PI3Kγ) is implicated as a target to repolarize tumour-associated macrophages and promote antitumour immune responses in solid cancers1-4. However, cancer cell-intrinsic roles of PI3Kγ are unclear. Here, by integrating unbiased genome-wide CRISPR interference screening with functional analyses across acute leukaemias, we define a selective dependency on the PI3Kγ complex in a high-risk subset that includes myeloid, lymphoid and dendritic lineages. This dependency is characterized by innate inflammatory signalling and activation of phosphoinositide 3-kinase regulatory subunit 5 (PIK3R5), which encodes a regulatory subunit of PI3Kγ5 and stabilizes the active enzymatic complex. We identify p21 (RAC1)-activated kinase 1 (PAK1) as a noncanonical substrate of PI3Kγ that mediates this cell-intrinsic dependency and find that dephosphorylation of PAK1 by PI3Kγ inhibition impairs mitochondrial oxidative phosphorylation. Treatment with the selective PI3Kγ inhibitor eganelisib is effective in leukaemias with activated PIK3R5. In addition, the combination of eganelisib and cytarabine prolongs survival over either agent alone, even in patient-derived leukaemia xenografts with low baseline PIK3R5 expression, as residual leukaemia cells after cytarabine treatment have elevated G protein-coupled purinergic receptor activity and PAK1 phosphorylation. Together, our study reveals a targetable dependency on PI3Kγ-PAK1 signalling that is amenable to near-term evaluation in patients with acute leukaemia.


Assuntos
Classe Ib de Fosfatidilinositol 3-Quinase , Leucemia , Transdução de Sinais , Quinases Ativadas por p21 , Animais , Humanos , Camundongos , Linhagem Celular , Classe Ib de Fosfatidilinositol 3-Quinase/genética , Classe Ib de Fosfatidilinositol 3-Quinase/metabolismo , Citarabina/farmacologia , Citarabina/uso terapêutico , Leucemia/tratamento farmacológico , Leucemia/enzimologia , Leucemia/genética , Leucemia/metabolismo , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Fosforilação Oxidativa/efeitos dos fármacos , Quinases Ativadas por p21/antagonistas & inibidores , Quinases Ativadas por p21/metabolismo , Fosforilação , Ensaios Antitumorais Modelo de Xenoenxerto
2.
Nature ; 618(7966): 834-841, 2023 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-37286599

RESUMO

Tumours most often arise from progression of precursor clones within a single anatomical niche. In the bone marrow, clonal progenitors can undergo malignant transformation to acute leukaemia, or differentiate into immune cells that contribute to disease pathology in peripheral tissues1-4. Outside the marrow, these clones are potentially exposed to a variety of tissue-specific mutational processes, although the consequences of this are unclear. Here we investigate the development of blastic plasmacytoid dendritic cell neoplasm (BPDCN)-an unusual form of acute leukaemia that often presents with malignant cells isolated to the skin5. Using tumour phylogenomics and single-cell transcriptomics with genotyping, we find that BPDCN arises from clonal (premalignant) haematopoietic precursors in the bone marrow. We observe that BPDCN skin tumours first develop at sun-exposed anatomical sites and are distinguished by clonally expanded mutations induced by ultraviolet (UV) radiation. A reconstruction of tumour phylogenies reveals that UV damage can precede the acquisition of alterations associated with malignant transformation, implicating sun exposure of plasmacytoid dendritic cells or committed precursors during BPDCN pathogenesis. Functionally, we find that loss-of-function mutations in Tet2, the most common premalignant alteration in BPDCN, confer resistance to UV-induced cell death in plasmacytoid, but not conventional, dendritic cells, suggesting a context-dependent tumour-suppressive role for TET2. These findings demonstrate how tissue-specific environmental exposures at distant anatomical sites can shape the evolution of premalignant clones to disseminated cancer.


Assuntos
Transformação Celular Neoplásica , Células Dendríticas , Leucemia Mieloide Aguda , Neoplasias Cutâneas , Pele , Raios Ultravioleta , Humanos , Células da Medula Óssea/metabolismo , Células da Medula Óssea/patologia , Células da Medula Óssea/efeitos da radiação , Morte Celular/efeitos da radiação , Linhagem da Célula/genética , Linhagem da Célula/efeitos da radiação , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Transformação Celular Neoplásica/efeitos da radiação , Células Clonais/metabolismo , Células Clonais/patologia , Células Clonais/efeitos da radiação , Células Dendríticas/metabolismo , Células Dendríticas/patologia , Células Dendríticas/efeitos da radiação , Leucemia Mieloide Aguda/etiologia , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Mutação/efeitos da radiação , Especificidade de Órgãos , Análise da Expressão Gênica de Célula Única , Neoplasias Cutâneas/etiologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Raios Ultravioleta/efeitos adversos , Pele/patologia , Pele/efeitos da radiação
3.
Blood ; 131(20): 2223-2234, 2018 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-29555646

RESUMO

Despite the well-established cell-intrinsic role of epigenetic factors in normal and malignant hematopoiesis, their cell-extrinsic role remains largely unexplored. Herein we investigated the hematopoietic impact of inactivating Ezh2, a key component of polycomb repressive complex 2 (PRC2), in the fetal liver (FL) vascular niche. Hematopoietic specific (Vav-iCre) Ezh2 inactivation enhanced FL hematopoietic stem cell (HSC) expansion with normal FL erythropoiesis. In contrast, endothelium (Tie2-Cre) targeted Ezh2 inactivation resulted in embryonic lethality with severe anemia at embryonic day 13.5 despite normal emergence of functional HSCs. Ezh2-deficient FL endothelium overexpressed Mmp9, which cell-extrinsically depleted the membrane-bound form of Kit ligand (mKitL), an essential hematopoietic cytokine, in FL. Furthermore, Mmp9 inhibition in vitro restored mKitL expression along with the erythropoiesis supporting capacity of FL endothelial cells. These data establish that Ezh2 is intrinsically dispensable for FL HSCs and provides proof of principle that modulation of epigenetic regulators in niche components can exert a marked cell-extrinsic impact.


Assuntos
Células Endoteliais/citologia , Células Endoteliais/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Feto , Hematopoese Extramedular , Fígado/fisiologia , Anemia/genética , Anemia/metabolismo , Animais , Biomarcadores , Células Cultivadas , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Imunofluorescência , Expressão Gênica , Inativação Gênica , Hematopoese Extramedular/genética , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Imuno-Histoquímica , Metaloproteinase 9 da Matriz/genética , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Fenótipo , Receptor TIE-2/genética , Receptor TIE-2/metabolismo , Fator de Células-Tronco/metabolismo
4.
Haematologica ; 104(11): 2215-2224, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-30975913

RESUMO

Somatic mutations in acute myeloid leukemia are acquired sequentially and hierarchically. First, pre-leukemic mutations, such as t(8;21) that encodes AML1-ETO, are acquired within the hematopoietic stem cell (HSC) compartment, while signaling pathway mutations, including KRAS activating mutations, are late events acquired during transformation of leukemic progenitor cells and are rarely detectable in HSC. This raises the possibility that signaling pathway mutations are detrimental to clonal expansion of pre-leukemic HSC. To address this hypothesis, we used conditional genetics to introduce Aml1-ETO and K-RasG12D into murine HSC, either individually or in combination. In the absence of activated Ras, Aml1-ETO-expressing HSC conferred a competitive advantage. However, activated K-Ras had a marked detrimental effect on Aml1-ETO-expressing HSC, leading to loss of both phenotypic and functional HSC. Cell cycle analysis revealed a loss of quiescence in HSC co-expressing Aml1-ETO and K-RasG12D, accompanied by an enrichment in E2F and Myc target gene expression and depletion of HSC self-renewal-associated gene expression. These findings provide a mechanistic basis for the observed absence of KRAS signaling mutations in the pre-malignant HSC compartment.


Assuntos
Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Mutação , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteína 1 Parceira de Translocação de RUNX1/genética , Proteína 1 Parceira de Translocação de RUNX1/metabolismo , Animais , Proliferação de Células/genética , Expressão Gênica , Perfilação da Expressão Gênica , Células-Tronco Hematopoéticas/patologia , Humanos , Camundongos , Camundongos Transgênicos , Modelos Animais , Modelos Biológicos , Lesões Pré-Cancerosas/genética , Lesões Pré-Cancerosas/metabolismo
5.
Trends Cancer ; 9(12): 987-988, 2023 12.
Artigo em Inglês | MEDLINE | ID: mdl-37783616

RESUMO

Dendritic cells play an important role in anticancer immunity by exposing T cells to tumor-associated antigens. In a recent study, Zhao et al. show that BCL2 inhibition improves the ability of dendritic cells to present antigen to T cells and activate their antitumor cytotoxicity.


Assuntos
Neoplasias , Humanos , Neoplasias/tratamento farmacológico , Antígenos de Neoplasias , Linfócitos T , Células Dendríticas , Proteínas Proto-Oncogênicas c-bcl-2
6.
bioRxiv ; 2023 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-38328043

RESUMO

Phosphoinositide 3-kinase gamma (PI3Kγ) is implicated as a target to repolarize tumor-associated macrophages and promote anti-tumor immune responses in solid cancers. However, cancer cell-intrinsic roles of PI3Kγ are unclear. Here, by integrating unbiased genome-wide CRISPR interference screening with functional analyses across acute leukemias, we define a selective dependency on the PI3Kγ complex in a high-risk subset that includes myeloid, lymphoid, and dendritic lineages. This dependency is characterized by innate inflammatory signaling and activation of phosphoinositide 3-kinase regulatory subunit 5 ( PIK3R5 ), which encodes a regulatory subunit of PI3Kγ and stabilizes the active enzymatic complex. Mechanistically, we identify p21 (RAC1) activated kinase 1 (PAK1) as a noncanonical substrate of PI3Kγ that mediates this cell-intrinsic dependency independently of Akt kinase. PI3Kγ inhibition dephosphorylates PAK1, activates a transcriptional network of NFκB-related tumor suppressor genes, and impairs mitochondrial oxidative phosphorylation. We find that treatment with the selective PI3Kγ inhibitor eganelisib is effective in leukemias with activated PIK3R5 , either at baseline or by exogenous inflammatory stimulation. Notably, the combination of eganelisib and cytarabine prolongs survival over either agent alone, even in patient-derived leukemia xenografts with low baseline PIK3R5 expression, as residual leukemia cells after cytarabine treatment have elevated G protein-coupled purinergic receptor activity and PAK1 phosphorylation. Taken together, our study reveals a targetable dependency on PI3Kγ/PAK1 signaling that is amenable to near-term evaluation in patients with acute leukemia.

7.
Cancer Discov ; 12(2): 522-541, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34615655

RESUMO

Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is an aggressive leukemia of plasmacytoid dendritic cells (pDC). BPDCN occurs at least three times more frequently in men than in women, but the reasons for this sex bias are unknown. Here, studying genomics of primary BPDCN and modeling disease-associated mutations, we link acquired alterations in RNA splicing to abnormal pDC development and inflammatory response through Toll-like receptors. Loss-of-function mutations in ZRSR2, an X chromosome gene encoding a splicing factor, are enriched in BPDCN, and nearly all mutations occur in males. ZRSR2 mutation impairs pDC activation and apoptosis after inflammatory stimuli, associated with intron retention and inability to upregulate the transcription factor IRF7. In vivo, BPDCN-associated mutations promote pDC expansion and signatures of decreased activation. These data support a model in which male-biased mutations in hematopoietic progenitors alter pDC function and confer protection from apoptosis, which may impair immunity and predispose to leukemic transformation. SIGNIFICANCE: Sex bias in cancer is well recognized, but the underlying mechanisms are incompletely defined. We connect X chromosome mutations in ZRSR2 to an extremely male-predominant leukemia. Aberrant RNA splicing induced by ZRSR2 mutation impairs dendritic cell inflammatory signaling, interferon production, and apoptosis, revealing a sex- and lineage-related tumor suppressor pathway.This article is highlighted in the In This Issue feature, p. 275.


Assuntos
Células Dendríticas/metabolismo , Transtornos Mieloproliferativos/genética , Ribonucleoproteínas/genética , Apoptose , Feminino , Identidade de Gênero , Humanos , Masculino , Mutação
8.
Nat Commun ; 12(1): 7019, 2021 12 02.
Artigo em Inglês | MEDLINE | ID: mdl-34857757

RESUMO

Yolk sac (YS) hematopoiesis is critical for the survival of the embryo and a major source of tissue-resident macrophages that persist into adulthood. Yet, the transcriptional and epigenetic regulation of YS hematopoiesis remains poorly characterized. Here we report that the epigenetic regulator Ezh2 is essential for YS hematopoiesis but dispensable for subsequent aorta-gonad-mesonephros (AGM) blood development. Loss of EZH2 activity in hemogenic endothelium (HE) leads to the generation of phenotypically intact but functionally deficient erythro-myeloid progenitors (EMPs), while the generation of primitive erythroid cells is not affected. EZH2 activity is critical for the generation of functional EMPs at the onset of the endothelial-to-hematopoietic transition but subsequently dispensable. We identify a lack of Wnt signaling downregulation as the primary reason for the production of non-functional EMPs. Together, our findings demonstrate a critical and stage-specific role of Ezh2 in modulating Wnt signaling during the generation of EMPs from YS HE.


Assuntos
Proteína Potenciadora do Homólogo 2 de Zeste/genética , Células Eritroides/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Células-Tronco Embrionárias Murinas/metabolismo , Células Progenitoras Mieloides/metabolismo , Proteínas de Transporte Vesicular/genética , Saco Vitelino/metabolismo , Animais , Diferenciação Celular , Embrião de Mamíferos , Proteína Potenciadora do Homólogo 2 de Zeste/deficiência , Epigênese Genética , Células Eritroides/citologia , Feminino , Feto , Genes Reporter , Hematopoese/genética , Fígado/citologia , Fígado/crescimento & desenvolvimento , Fígado/metabolismo , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Células-Tronco Embrionárias Murinas/citologia , Células Progenitoras Mieloides/patologia , Cultura Primária de Células , Proteínas de Transporte Vesicular/metabolismo , Via de Sinalização Wnt , Saco Vitelino/citologia , Saco Vitelino/crescimento & desenvolvimento , Proteína Vermelha Fluorescente
9.
J Exp Med ; 218(2)2021 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-33416891

RESUMO

Juvenile myelomonocytic leukemia (JMML) is a poor-prognosis childhood leukemia usually caused by RAS-pathway mutations. The cellular hierarchy in JMML is poorly characterized, including the identity of leukemia stem cells (LSCs). FACS and single-cell RNA sequencing reveal marked heterogeneity of JMML hematopoietic stem/progenitor cells (HSPCs), including an aberrant Lin-CD34+CD38-CD90+CD45RA+ population. Single-cell HSPC index-sorting and clonogenic assays show that (1) all somatic mutations can be backtracked to the phenotypic HSC compartment, with RAS-pathway mutations as a "first hit," (2) mutations are acquired with both linear and branching patterns of clonal evolution, and (3) mutant HSPCs are present after allogeneic HSC transplant before molecular/clinical evidence of relapse. Stem cell assays reveal interpatient heterogeneity of JMML LSCs, which are present in, but not confined to, the phenotypic HSC compartment. RNA sequencing of JMML LSC reveals up-regulation of stem cell and fetal genes (HLF, MEIS1, CNN3, VNN2, and HMGA2) and candidate therapeutic targets/biomarkers (MTOR, SLC2A1, and CD96), paving the way for LSC-directed disease monitoring and therapy in this disease.


Assuntos
Células-Tronco Hematopoéticas/patologia , Leucemia Mielomonocítica Juvenil/patologia , Animais , Biomarcadores Tumorais/genética , Linhagem Celular , Feminino , Humanos , Leucemia Mielomonocítica Juvenil/genética , Masculino , Camundongos , Mutação/genética , Células-Tronco Neoplásicas/patologia , Transdução de Sinais/genética , Regulação para Cima/genética
10.
Cancer Cell ; 33(2): 274-291.e8, 2018 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-29438697

RESUMO

Lympho-myeloid restricted early thymic progenitors (ETPs) are postulated to be the cell of origin for ETP leukemias, a therapy-resistant leukemia associated with frequent co-occurrence of EZH2 and RUNX1 inactivating mutations, and constitutively activating signaling pathway mutations. In a mouse model, we demonstrate that Ezh2 and Runx1 inactivation targeted to early lymphoid progenitors causes a marked expansion of pre-leukemic ETPs, showing transcriptional signatures characteristic of ETP leukemia. Addition of a RAS-signaling pathway mutation (Flt3-ITD) results in an aggressive leukemia co-expressing myeloid and lymphoid genes, which can be established and propagated in vivo by the expanded ETPs. Both mouse and human ETP leukemias show sensitivity to BET inhibition in vitro and in vivo, which reverses aberrant gene expression induced by Ezh2 inactivation.


Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core/genética , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Leucemia Mieloide Aguda/genética , Mutação/genética , Animais , Regulação Leucêmica da Expressão Gênica , Camundongos Knockout , Células Mieloides/metabolismo , Transdução de Sinais/genética , Células-Tronco
11.
Nat Med ; 23(6): 692-702, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28504724

RESUMO

Recent advances in single-cell transcriptomics are ideally placed to unravel intratumoral heterogeneity and selective resistance of cancer stem cell (SC) subpopulations to molecularly targeted cancer therapies. However, current single-cell RNA-sequencing approaches lack the sensitivity required to reliably detect somatic mutations. We developed a method that combines high-sensitivity mutation detection with whole-transcriptome analysis of the same single cell. We applied this technique to analyze more than 2,000 SCs from patients with chronic myeloid leukemia (CML) throughout the disease course, revealing heterogeneity of CML-SCs, including the identification of a subgroup of CML-SCs with a distinct molecular signature that selectively persisted during prolonged therapy. Analysis of nonleukemic SCs from patients with CML also provided new insights into cell-extrinsic disruption of hematopoiesis in CML associated with clinical outcome. Furthermore, we used this single-cell approach to identify a blast-crisis-specific SC population, which was also present in a subclone of CML-SCs during the chronic phase in a patient who subsequently developed blast crisis. This approach, which might be broadly applied to any malignancy, illustrates how single-cell analysis can identify subpopulations of therapy-resistant SCs that are not apparent through cell-population analysis.


Assuntos
Crise Blástica/genética , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Células-Tronco Neoplásicas/metabolismo , Análise de Célula Única , Adulto , Idoso , Imunoprecipitação da Cromatina , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Feminino , Citometria de Fluxo , Biblioteca Gênica , Genes abl/genética , Humanos , Hibridização in Situ Fluorescente , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Análise de Sequência de RNA , Transcriptoma , Adulto Jovem
12.
J Exp Med ; 214(7): 2005-2021, 2017 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-28637883

RESUMO

Although previous studies suggested that the expression of FMS-like tyrosine kinase 3 (Flt3) initiates downstream of mouse hematopoietic stem cells (HSCs), FLT3 internal tandem duplications (FLT3 ITDs) have recently been suggested to intrinsically suppress HSCs. Herein, single-cell interrogation found Flt3 mRNA expression to be absent in the large majority of phenotypic HSCs, with a strong negative correlation between Flt3 and HSC-associated gene expression. Flt3-ITD knock-in mice showed reduced numbers of phenotypic HSCs, with an even more severe loss of long-term repopulating HSCs, likely reflecting the presence of non-HSCs within the phenotypic HSC compartment. Competitive transplantation experiments established that Flt3-ITD compromises HSCs through an extrinsically mediated mechanism of disrupting HSC-supporting bone marrow stromal cells, with reduced numbers of endothelial and mesenchymal stromal cells showing increased inflammation-associated gene expression. Tumor necrosis factor (TNF), a cell-extrinsic potent negative regulator of HSCs, was overexpressed in bone marrow niche cells from FLT3-ITD mice, and anti-TNF treatment partially rescued the HSC phenotype. These findings, which establish that Flt3-ITD-driven myeloproliferation results in cell-extrinsic suppression of the normal HSC reservoir, are of relevance for several aspects of acute myeloid leukemia biology.


Assuntos
Proliferação de Células/genética , Células-Tronco Hematopoéticas/metabolismo , Mutação , Nicho de Células-Tronco/genética , Tirosina Quinase 3 Semelhante a fms/genética , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Transplante de Medula Óssea , Células Cultivadas , Etanercepte/farmacologia , Perfilação da Expressão Gênica/métodos , Células-Tronco Hematopoéticas/efeitos dos fármacos , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Célula Única/métodos , Sequências de Repetição em Tandem/genética , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo
13.
Oncoscience ; 5(11-12): 271-272, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30652110
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