RESUMO
Plasmacytoid dendritic cells (pDCs) are the principal natural type I interferon-producing dendritic cells. Neoplastic expansion of pDCs and pDC precursors leads to blastic plasmacytoid dendritic cell neoplasm (BPDCN), and clonal expansion of mature pDCs has been described in chronic myelomonocytic leukemia. The role of pDC expansion in acute myeloid leukemia (AML) is poorly studied. Here, we characterize patients with AML with pDC expansion (pDC-AML), which we observe in â¼5% of AML cases. pDC-AMLs often possess cross-lineage antigen expression and have adverse risk stratification with poor outcome. RUNX1 mutations are the most common somatic alterations in pDC-AML (>70%) and are much more common than in AML without pDC expansion and BPDCN. We demonstrate that pDCs are clonally related to, as well as originate from, leukemic blasts in pDC-AML. We further demonstrate that leukemic blasts from RUNX1-mutated AML upregulate a pDC transcriptional program, poising the cells toward pDC differentiation and expansion. Finally, tagraxofusp, a targeted therapy directed to CD123, reduces leukemic burden and eliminates pDCs in a patient-derived xenograft model. In conclusion, pDC-AML is characterized by a high frequency of RUNX1 mutations and increased expression of a pDC transcriptional program. CD123 targeting represents a potential treatment approach for pDC-AML.
Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core/genética , Células Dendríticas/patologia , Leucemia Mieloide Aguda/genética , Adulto , Idoso , Crise Blástica/genética , Crise Blástica/patologia , Células Dendríticas/metabolismo , Feminino , Humanos , Leucemia Mieloide Aguda/patologia , Masculino , Pessoa de Meia-Idade , MutaçãoRESUMO
Cancer stem cells have been strongly linked to resistance and relapse in many malignancies. However, purifying them from within the bulk tumor has been challenging, so their precise genetic and functional characteristics are not well defined. The side population assay exploits the ability of some cells to efflux Hoechst dye via ATP-binding cassette transporters. Stem cells have increased expression of these transporters and this assay has been shown to enrich for stem cells in various tissues and cancers. This study identifies the side population within a zebrafish model of acute lymphoblastic leukemia and correlates the frequency of side population cells with the frequency of leukemia stem cells (more precisely referred to as leukemia-propagating cells within our transplantation model). In addition, the side population within the leukemia evolves with serial transplantation, increasing in tandem with leukemia-propagating cell frequency over subsequent generations. Sorted side population cells from these tumors are enriched for leukemia-propagating cells and have enhanced engraftment compared to sorted non-side population cells when transplanted into syngeneic recipients. RNA-sequencing analysis of sorted side population cells compared to non-side population cells identified a shared expression profile within the side population and pathway analysis yielded Wnt-signaling as the most overrepresented. Gene set enrichment analysis showed that stem cell differentiation and canonical Wnt-signaling were significantly upregulated in the side population. Overall, these results demonstrate that the side population in zebrafish acute lymphoblastic leukemia significantly enriches for leukemia-propagating cells and identifies the Wnt pathway as a likely genetic driver of leukemia stem cell fate.
Assuntos
Biomarcadores Tumorais/análise , Diferenciação Celular , Transformação Celular Neoplásica/patologia , Células-Tronco Neoplásicas/patologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Células da Side Population/patologia , Via de Sinalização Wnt , Animais , Transformação Celular Neoplásica/metabolismo , Humanos , Células-Tronco Neoplásicas/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Células da Side Population/metabolismo , Células Tumorais Cultivadas , Peixe-ZebraRESUMO
Clonal hematopoiesis (CH) is a common premalignant state in the blood and confers an increased risk of blood cancers and all-cause mortality. Identification of therapeutic targets in CH has been hindered by the lack of an ex vivo platform amenable for studying primary hematopoietic stem and progenitor cells (HSPCs). Here, we utilize an ex vivo co-culture system of HSPCs with bone marrow endothelial cells to perform CRISPR/Cas9 screens in mutant HSPCs. Our data reveal that loss of the histone demethylase family members Kdm3b and Jmjd1c specifically reduces the fitness of Idh2- and Tet2-mutant HSPCs. Kdm3b loss in mutant cells leads to decreased expression of critical cytokine receptors including Mpl, rendering mutant HSPCs preferentially susceptible to inhibition of downstream JAK2 signaling. Our study nominates an epigenetic regulator and an epigenetically regulated receptor signaling pathway as genotype-specific therapeutic targets and provides a scalable platform to identify genetic dependencies in mutant HSPCs.
RESUMO
Gain-of-function mutations activating JAK/STAT signaling are seen in the majority of patients with myeloproliferative neoplasms (MPN), most commonly JAK2V617F. Although clinically approved JAK inhibitors improve symptoms and outcomes in MPNs, remissions are rare, and mutant allele burden does not substantively change with chronic therapy. We hypothesized this is due to limitations of current JAK inhibitors to potently and specifically abrogate mutant JAK2 signaling. We therefore developed a conditionally inducible mouse model allowing for sequential activation, and then inactivation, of Jak2V617F from its endogenous locus using a combined Dre-rox/Cre-lox dual-recombinase system. Jak2V617F deletion abrogates MPN features, induces depletion of mutant-specific hematopoietic stem/progenitor cells, and extends overall survival to an extent not observed with pharmacologic JAK inhibition, including when cooccurring with somatic Tet2 loss. Our data suggest JAK2V617F represents the best therapeutic target in MPNs and demonstrate the therapeutic relevance of a dual-recombinase system to assess mutant-specific oncogenic dependencies in vivo. SIGNIFICANCE: Current JAK inhibitors to treat myeloproliferative neoplasms are ineffective at eradicating mutant cells. We developed an endogenously expressed Jak2V617F dual-recombinase knock-in/knock-out model to investigate Jak2V617F oncogenic reversion in vivo. Jak2V617F deletion abrogates MPN features and depletes disease-sustaining MPN stem cells, suggesting improved Jak2V617F targeting offers the potential for greater therapeutic efficacy. See related commentary by Celik and Challen, p. 701. This article is featured in Selected Articles from This Issue, p. 695.
Assuntos
Janus Quinase 2 , Transtornos Mieloproliferativos , Animais , Humanos , Camundongos , Modelos Animais de Doenças , Células-Tronco Hematopoéticas/metabolismo , Janus Quinase 2/genética , Janus Quinase 2/metabolismo , Mutação , Transtornos Mieloproliferativos/genética , Transtornos Mieloproliferativos/tratamento farmacológico , Transdução de SinaisRESUMO
Targeted therapies have come to play an increasingly important role in cancer therapy over the past two decades. This success has been made possible in large part by technological advances in sequencing, which have greatly advanced our understanding of the mutational landscape of human cancer and the genetic drivers present in individual tumors. We are rapidly discovering a growing number of mutations that occur in targetable pathways, and thus tumor genetic testing has become an important component in the choice of appropriate therapies. Targeted therapy has dramatically transformed treatment outcomes and disease prognosis in some settings, whereas in other oncologic contexts, targeted approaches have yet to demonstrate considerable clinical efficacy. In this Review, we summarize the current knowledge of targetable mutations that occur in a range of cancers, including hematologic malignancies and solid tumors such as non-small cell lung cancer and breast cancer. We outline seminal examples of druggable mutations and targeting modalities and address the clinical and research challenges that must be overcome to maximize therapeutic benefit.
Assuntos
Neoplasias da Mama , Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/terapia , Feminino , Humanos , Neoplasias Pulmonares/genética , Mutação , PrognósticoRESUMO
Somatic mutations in cancer genes have been detected in clonal expansions across healthy human tissue, including in clonal hematopoiesis. However, because mutated and wild-type cells are admixed, we have limited ability to link genotypes with phenotypes. To overcome this limitation, we leveraged multi-modality single-cell sequencing, capturing genotype, transcriptomes and methylomes in progenitors from individuals with DNMT3A R882 mutated clonal hematopoiesis. DNMT3A mutations result in myeloid over lymphoid bias, and an expansion of immature myeloid progenitors primed toward megakaryocytic-erythroid fate, with dysregulated expression of lineage and leukemia stem cell markers. Mutated DNMT3A leads to preferential hypomethylation of polycomb repressive complex 2 targets and a specific CpG flanking motif. Notably, the hypomethylation motif is enriched in binding motifs of key hematopoietic transcription factors, serving as a potential mechanistic link between DNMT3A mutations and aberrant transcriptional phenotypes. Thus, single-cell multi-omics paves the road to defining the downstream consequences of mutations that drive clonal mosaicism.
Assuntos
Hematopoiese Clonal , DNA (Citosina-5-)-Metiltransferases , DNA Metiltransferase 3A/genética , DNA (Citosina-5-)-Metiltransferases/genética , Metilases de Modificação do DNA/genética , Hematopoese/genética , Humanos , Mutação , Complexo Repressor Polycomb 2/genéticaRESUMO
Heterogeneous cell populations, from either healthy or malignant tissues, may contain a population of cells characterized by a differential ability to efflux the DNA-binding dye Hoechst 33342. This "side population" of cells can be identified using flow cytometric methods after the Hoechst 33342 dye is excited by an ultraviolet (UV) laser. The side population of many cell types contains stem- or progenitor-like cells. However, not all cell types have an identifiable side population. Danio rerio, zebrafish, have a robust in vivo model of T-cell acute lymphoblastic leukemia (T-ALL), but whether these zebrafish T-ALLs have a side population is unknown. The method described here outlines how to isolate the side population cells in zebrafish T-ALL. To begin, the T-ALL in zebrafish is generated via the microinjection of tol2 plasmids into one-cell stage embryos. Once the tumors have grown to a stage at which they expand into more than half of the animal's body, the T-ALL cells can be harvested. The cells are then stained with Hoechst 33342 and examined by flow cytometry for side population cells. This method has broad applications in zebrafish T-ALL research. While there are no known cell surface markers in zebrafish that confirm whether these side population cells are cancer stem cell-like, in vivo functional transplantation assays are possible. Furthermore, single-cell transcriptomics could be applied to identify the genetic features of these side population cells.