RESUMEN
Translocations of Meningioma-1 (MN1) occur in a subset of acute myeloid leukemias (AML) and result in high expression of MN1, either as a full-length protein, or as a fusion protein that includes most of the N-terminus of MN1. High levels of MN1 correlate with poor prognosis. When overexpressed in murine hematopoietic progenitors, MN1 causes an aggressive AML characterized by an aberrant myeloid precursor-like gene expression program that shares features of KMT2A-rearranged (KMT2A-r) leukemia, including high levels of Hoxa and Meis1 gene expression. Compounds that target a critical KMT2A-Menin interaction have proven effective in KMT2A-r leukemia. Here, we demonstrate that Menin (Men1) is also critical for the self-renewal of MN1-driven AML through the maintenance of a distinct gene expression program. Genetic inactivation of Men1 led to a decrease in the number of functional leukemia-initiating cells. Pharmacologic inhibition of the KMT2A-Menin interaction decreased colony-forming activity, induced differentiation programs in MN1-driven murine leukemia and decreased leukemic burden in a human AML xenograft carrying an MN1-ETV6 translocation. Collectively, these results nominate Menin inhibition as a promising therapeutic strategy in MN1-driven leukemia.
Asunto(s)
Leucemia Mieloide Aguda/genética , Proteínas Proto-Oncogénicas/genética , Transactivadores/genética , Proteínas Supresoras de Tumor/genética , Animales , Línea Celular Tumoral , Regulación Leucémica de la Expresión Génica/genética , Células HEK293 , N-Metiltransferasa de Histona-Lisina/genética , Humanos , Ratones , Ratones NoqueadosRESUMEN
Ewing's sarcoma (ES) is a rare and highly malignant cancer that grows in the bones or surrounding tissues mostly affecting adolescents and young adults. A chimeric fusion between the RNA binding protein EWS and the ETS family transcription factor FLI1 (EWS-FLI1), which is generated from a chromosomal translocation, is implicated in driving most ES cases by modulation of transcription and alternative splicing. The small-molecule YK-4-279 inhibits EWS-FLI1 function and induces apoptosis in ES cells. We aimed to identify both the underlying mechanism of the drug and potential combination therapies that might enhance its antitumor activity. We tested 69 anticancer drugs in combination with YK-4-279 and found that vinca alkaloids exhibited synergy with YK-4-279 in five ES cell lines. The combination of YK-4-279 and vincristine reduced tumor burden and increased survival in mice bearing ES xenografts. We determined that independent drug-induced events converged to cause this synergistic therapeutic effect. YK-4-279 rapidly induced G2-M arrest, increased the abundance of cyclin B1, and decreased EWS-FLI1-mediated generation of microtubule-associated proteins, which rendered cells more susceptible to microtubule depolymerization by vincristine. YK-4-279 reduced the expression of the EWS-FLI1 target gene encoding the ubiquitin ligase UBE2C, which, in part, contributed to the increase in cyclin B1. YK-4-279 also increased the abundance of proapoptotic isoforms of MCL1 and BCL2, presumably through inhibition of alternative splicing by EWS-FLI1, thus promoting cell death in response to vincristine. Thus, a combination of vincristine and YK-4-279 might be therapeutically effective in ES patients.
Asunto(s)
Resistencia a Antineoplásicos/efectos de los fármacos , Puntos de Control de la Fase G2 del Ciclo Celular/efectos de los fármacos , Indoles/farmacología , Puntos de Control de la Fase M del Ciclo Celular/efectos de los fármacos , Proteínas de Fusión Oncogénica/antagonistas & inhibidores , Proteína Proto-Oncogénica c-fli-1/antagonistas & inhibidores , Proteína EWS de Unión a ARN/antagonistas & inhibidores , Sarcoma de Ewing/tratamiento farmacológico , Vincristina/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/genética , Línea Celular Tumoral , Ciclina B1/genética , Ciclina B1/metabolismo , Resistencia a Antineoplásicos/genética , Puntos de Control de la Fase G2 del Ciclo Celular/genética , Humanos , Puntos de Control de la Fase M del Ciclo Celular/genética , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/genética , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteínas de Fusión Oncogénica/genética , Proteínas de Fusión Oncogénica/metabolismo , Proteína Proto-Oncogénica c-fli-1/genética , Proteína Proto-Oncogénica c-fli-1/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteína EWS de Unión a ARN/genética , Proteína EWS de Unión a ARN/metabolismo , Sarcoma de Ewing/genética , Sarcoma de Ewing/metabolismo , Sarcoma de Ewing/patología , Enzimas Ubiquitina-Conjugadoras/genética , Enzimas Ubiquitina-Conjugadoras/metabolismoRESUMEN
DOT1L is a protein methyltransferase involved in the development and maintenance of MLL-rearranged (MLL-r) leukemia through its ectopic methylation of histones associated with well-characterized leukemic genes. Pinometostat (EPZ-5676), a selective inhibitor of DOT1L, is in clinical development in relapsed/refractory acute leukemia patients harboring rearrangements of the MLL gene. The observation of responses and subsequent relapses in the adult trial treating MLL-r patients motivated preclinical investigations into potential mechanisms of pinometostat treatment-emergent resistance (TER) in cell lines confirmed to have MLL-r. TER was achieved in five MLL-r cell lines, KOPN-8, MOLM-13, MV4-11, NOMO-1, and SEM. Two of the cell lines, KOPN-8 and NOMO-1, were thoroughly characterized to understand the mechanisms involved in pinometostat resistance. Unlike many other targeted therapies, resistance does not appear to be achieved through drug-induced selection of mutations of the target itself. Instead, we identified both drug efflux transporter dependent and independent mechanisms of resistance to pinometostat. In KOPN-8 TER cells, increased expression of the drug efflux transporter ABCB1 (P-glycoprotein, MDR1) was the primary mechanism of drug resistance. In contrast, resistance in NOMO-1 cells occurs through a mechanism other than upregulation of a specific efflux pump. RNA-seq analysis performed on both parental and resistant KOPN-8 and NOMO-1 cell lines supported two unique candidate pathway mechanisms that may explain the pinometostat resistance observed in these cell lines. These results are the first demonstration of TER models of the DOT1L inhibitor pinometostat and may provide useful tools for investigating clinical resistance. Mol Cancer Ther; 16(8); 1669-79. ©2017 AACR.
Asunto(s)
Bencimidazoles/uso terapéutico , Resistencia a Antineoplásicos , Reordenamiento Génico , N-Metiltransferasa de Histona-Lisina/genética , Leucemia/tratamiento farmacológico , Leucemia/genética , Proteína de la Leucemia Mieloide-Linfoide/genética , Subfamilia B de Transportador de Casetes de Unión a ATP/genética , Subfamilia B de Transportador de Casetes de Unión a ATP/metabolismo , Bencimidazoles/farmacología , Biomarcadores de Tumor/metabolismo , Línea Celular Tumoral , Resistencia a Antineoplásicos/efectos de los fármacos , Regulación Leucémica de la Expresión Génica/efectos de los fármacos , Histonas/metabolismo , Humanos , Lisina/metabolismo , Metilación , Modelos Biológicos , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
Transient potential receptor melastatin-2 (TRPM2) is a nonselective cationic, Ca(2+)-permeable transmembrane pore that is preferentially expressed in cells of the myeloid lineage and modulates signaling pathways converging into NF-kB. This is of potential interest for acute myeloid leukemia (AML) therapy, as NF-κB signaling is emerging as a key pathway, mediating drug resistance and leukemia-initiating cell survival in AML. Inhibition of NF-κB signaling has been found to be synergistic with chemotherapy. TRPM2 is overexpressed in AML compared with normal bone marrow, with the highest levels in the FAB M3-6 subtypes. To determine the effect of TRPM2 depletions in a defined genetic model, we established MLL-AF9-driven AML on a Trpm2(-/-) genetic background. Trpm2(-/-) MLL-AF9 leukemias displayed reduced NF-κB phosphorylation as well as nuclear translocation. In vivo, primary and secondary recipients of Trpm2(-/-) MLL-AF9 leukemias exhibit increased latency compared with recipients of wild-type leukemia cells. However, the difference in latency was small and was lost in tertiary transplants. The effect of loss of Trpm2 in a BCR-ABL/NUP98-HOXA9 fusion model was even smaller. Given reports that loss or inhibition of TRPM2 enhanced killing by DNA-damaging agents in neuroblastoma, breast cancer, and prostate cancer cell lines, we exposed Trpm2(-/-) and Trpm2(wt) primary MLL-AF9 leukemias to doxorubicin, cytarabine, and etoposide, but found no difference in IC50 values. The in vitro response to decitabine was also unaffected. In summary, Trpm2 does not seem to play a major role in myeloid leukemogenesis. Additionally, loss of Trpm2 does not augment the cytotoxicity of standard AML chemotherapeutic agents.
Asunto(s)
Transformación Celular Neoplásica/genética , Resistencia a Antineoplásicos/genética , Leucemia Mieloide Aguda/genética , Canales Catiónicos TRPM/genética , Animales , Transformación Celular Neoplásica/metabolismo , Modelos Animales de Enfermedad , Regulación Leucémica de la Expresión Génica , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/mortalidad , Leucemia Mieloide Aguda/patología , Ratones , Ratones Noqueados , FN-kappa B/metabolismo , Fosforilación , Pronóstico , Canales Catiónicos TRPM/metabolismoRESUMEN
Meningioma-1 (MN1) overexpression is frequently observed in patients with acute myeloid leukemia (AML) and is predictive of poor prognosis. In murine models, forced expression of MN1 in hematopoietic progenitors induces an aggressive myeloid leukemia that is strictly dependent on a defined gene expression program in the cell of origin, which includes the homeobox genes Hoxa9 and Meis1 as key components. Here, we have shown that this program is controlled by two histone methyltransferases, MLL1 and DOT1L, as deletion of either Mll1 or Dot1l in MN1-expressing cells abrogated the cell of origin-derived gene expression program, including the expression of Hoxa cluster genes. In murine models, genetic inactivation of either Mll1 or Dot1l impaired MN1-mediated leukemogenesis. We determined that HOXA9 and MEIS1 are coexpressed with MN1 in a subset of clinical MN1hi leukemia, and human MN1hi/HOXA9hi leukemias were sensitive to pharmacologic inhibition of DOT1L. Together, these data point to DOT1L as a potential therapeutic target in MN1hi AML. In addition, our findings suggest that epigenetic modulation of the interplay between an oncogenic lesion and its cooperating developmental program has therapeutic potential in AML.
Asunto(s)
N-Metiltransferasa de Histona-Lisina/metabolismo , Leucemia Mieloide Aguda/metabolismo , Metiltransferasas/metabolismo , Proteína de la Leucemia Mieloide-Linfoide/metabolismo , Proteínas Oncogénicas/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Animales , Femenino , N-Metiltransferasa de Histona-Lisina/genética , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patología , Masculino , Metiltransferasas/genética , Ratones , Ratones Noqueados , Proteína 1 del Sitio de Integración Viral Ecotrópica Mieloide , Proteína de la Leucemia Mieloide-Linfoide/genética , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Proteínas Oncogénicas/genética , Transactivadores , Proteínas Supresoras de Tumor/genéticaRESUMEN
Early T cell precursor acute lymphoblastic leukemia (ETP-ALL) is an aggressive subtype of ALL distinguished by stem-cell-associated and myeloid transcriptional programs. Inactivating alterations of Polycomb repressive complex 2 components are frequent in human ETP-ALL, but their functional role is largely undefined. We have studied the involvement of Ezh2 in a murine model of NRASQ61K-driven leukemia that recapitulates phenotypic and transcriptional features of ETP-ALL. Homozygous inactivation of Ezh2 cooperated with oncogenic NRASQ61K to accelerate leukemia onset. Inactivation of Ezh2 accentuated expression of genes highly expressed in human ETP-ALL and in normal murine early thymic progenitors. Moreover, we found that Ezh2 contributes to the silencing of stem-cell- and early-progenitor-cell-associated genes. Loss of Ezh2 also resulted in increased activation of STAT3 by tyrosine 705 phosphorylation. Our data mechanistically link Ezh2 inactivation to stem-cell-associated transcriptional programs and increased growth/survival signaling, features that convey an adverse prognosis in patients.
Asunto(s)
Proteína Potenciadora del Homólogo Zeste 2/genética , Regulación Leucémica de la Expresión Génica , Genes ras , Complejo Represivo Polycomb 2/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Factor de Transcripción STAT3/genética , Animales , Modelos Animales de Enfermedad , Proteína Potenciadora del Homólogo Zeste 2/deficiencia , Histonas/genética , Histonas/metabolismo , Humanos , Interleucina-6/genética , Interleucina-6/metabolismo , Janus Quinasa 1/genética , Janus Quinasa 1/metabolismo , Ratones , Ratones Transgénicos , Fosforilación , Complejo Represivo Polycomb 2/deficiencia , Complejo Represivo Polycomb 2/metabolismo , Células Precursoras de Linfocitos T/metabolismo , Células Precursoras de Linfocitos T/patología , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patología , Receptores de Interleucina-6/genética , Receptores de Interleucina-6/metabolismo , Factor de Transcripción STAT3/metabolismo , Transducción de Señal , Transcripción GenéticaRESUMEN
Genome scale sequencing in patients with cancer has revealed a lower frequency of genetic aberrations in hematologic disorders compared with most other malignancies, suggesting a prominent role for epigenetic mechanisms. In parallel, epigenetic modifiers that are altered in cancer play critical roles in normal hematopoietic development, influencing both self-renewal of hematopoietic stem cells and differentiation into the different lineages. In this review, we aim to compare the role of several key DNA or histone modifying enzymes and complexes in normal development and hematopoietic malignancies, including DNMT3A, TET2, IDH1, IDH2, MLL1, MLL4, DOT1L, PRC1/2 and WSHC1/NSD2/MMSET. Insights into their biological mechanisms led to the development of therapies designed to target mutant IDH1 and IDH2, DOT1L in MLL-rearranged leukemias and EZH2 in several cancer types including lymphomas. Inhibitors for these enzymes are currently in clinical trials.
