RESUMEN
Epigenetic proteins containing YEATS domains (YD) are an emerging target class in drug discovery. Described herein are the discovery and characterization efforts associated with PFI-6, a new chemical probe for the YD of MLLT1 (ENL/YEATS1) and MLLT3 (AF9/YEATS3). For hit identification, fragment-like mimetics of endogenous YD ligands (crotonylated histone-containing proteins), were synthesized via parallel medicinal chemistry (PMC) and screened for MLLT1 binding. Subsequent SAR studies led to iterative MLLT1/3 binding and selectivity improvements, culminating in the discovery of PFI-6. PFI-6 demonstrates good affinity and selectivity for MLLT1/3 vs. other human YD proteins (YEATS2/4) and engages MLLT3 in cells. Small-molecule X-ray co-crystal structures of two molecules, including PFI-6, bound to the YD of MLLT1/3 are also described. PFI-6 may be a useful tool molecule to better understand the biological effects associated with modulation of MLLT1/3.
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Histonas , Proteínas Nucleares , Humanos , Proteínas Nucleares/metabolismo , Histonas/metabolismo , Dominios Proteicos , Descubrimiento de Drogas , Proteínas de Neoplasias/metabolismo , Factores de Transcripción/metabolismoRESUMEN
This is a retrospective study comparing the effectiveness of umbilical cord blood transplantation (UCBT) and chemotherapy for patients in the first complete remission period for acute myeloid leukemia with KMT2A-MLLT3 rearrangements. A total of 22 patients were included, all of whom achieved first complete remission (CR1) through 1-2 rounds of induction chemotherapy, excluding patients with an early relapse. Twelve patients were treated with UCBT, and 10 patients were treated with chemotherapy after 2 to 4 courses of consolidation therapy. The 3-year overall survival (OS) of the UCBT group was 71.3% (95% CI, 34.4-89.8%), and that of the chemotherapy group was 10% (95% CI, 5.89-37.3%). The OS of the UCBT group was significantly higher than that of the chemotherapy group (P = 0.003). The disease-free survival (DFS) of the UCBT group was 60.8% (95% CI, 25.0-83.6%), which was significantly higher than the 10% (95% CI, 5.72-35.8%) of the chemotherapy group (P = 0.003). The relapse rate of the UCBT group was 23.6% (95% CI, 0-46.8%), and that of the chemotherapy group was 85.4% (95% CI, 35.8-98.4%), which was significantly higher than that of the UCBT group (P < 0.001). The non-relapse mortality (NRM) rate in the UCBT group was 19.8% (95% CI, 0-41.3%), and that in the chemotherapy group was 0.0%. The NRM rate in the UCBT group was higher than that in the chemotherapy group, but there was no significant difference between the two groups (P = 0.272). Two patients in the UCBT group relapsed, two died of acute and chronic GVHD, and one patient developed chronic GVHD 140 days after UCBT and is still alive, so the GVHD-free/relapse-free survival (GRFS) was 50% (95% CI, 17.2-76.1%). AML patients with KMT2A-MLLT3 rearrangements who receive chemotherapy as their consolidation therapy after CR1 have a very poor prognosis. UCBT can overcome the poor prognosis and significantly improve survival, and the GRFS for these patients is very good. We suggest that UCBT is a better choice than chemotherapy for KMT2A-MLLT3 patients.
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Sangre Fetal/trasplante , N-Metiltransferasa de Histona-Lisina/genética , Leucemia Mieloide Aguda/terapia , Proteína de la Leucemia Mieloide-Linfoide/genética , Proteínas Nucleares/genética , Adolescente , Adulto , Niño , Preescolar , Quimioterapia de Consolidación , Supervivencia sin Enfermedad , Femenino , Reordenamiento Génico , Enfermedad Injerto contra Huésped , Humanos , Quimioterapia de Inducción , Leucemia Mieloide Aguda/diagnóstico , Leucemia Mieloide Aguda/genética , Masculino , Persona de Mediana Edad , Pronóstico , Adulto JovenRESUMEN
Leukemias derived from the MLL-AF9 rearrangement rely on dysfunctional transcriptional networks. ZNF521, a transcription co-factor implicated in the control of hematopoiesis, has been proposed to sustain leukemic transformation in collaboration with other oncogenes. Here, we demonstrate that ZNF521 mRNA levels correlate with specific genetic aberrations: in particular, the highest expression is observed in AMLs bearing MLL rearrangements, while the lowest is detected in AMLs with FLT3-ITD, NPM1, or CEBPα double mutations. In cord blood-derived CD34+ cells, enforced expression of ZNF521 provides a significant proliferative advantage and enhances MLL-AF9 effects on the induction of proliferation and the expansion of leukemic progenitor cells. Transcriptome analysis of primary CD34+ cultures displayed subsets of genes up-regulated by MLL-AF9 or ZNF521 single transgene overexpression as well as in MLL-AF9/ZNF521 combinations, at either the early or late time points of an in vitro leukemogenesis model. The silencing of ZNF521 in the MLL-AF9 + THP-1 cell line coherently results in an impairment of growth and clonogenicity, recapitulating the effects observed in primary cells. Taken together, these results underscore a role for ZNF521 in sustaining the self-renewal of the immature AML compartment, most likely through the perturbation of the gene expression landscape, which ultimately favors the expansion of MLL-AF9-transformed leukemic clones.
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Biomarcadores de Tumor/metabolismo , Proteínas de Unión al ADN/metabolismo , Regulación Neoplásica de la Expresión Génica , Células Madre Hematopoyéticas/patología , Leucemia Mieloide Aguda/patología , Proteína de la Leucemia Mieloide-Linfoide/metabolismo , Proteínas de Fusión Oncogénica/metabolismo , Apoptosis , Biomarcadores de Tumor/genética , Proliferación Celular , Proteínas de Unión al ADN/genética , Células Madre Hematopoyéticas/metabolismo , Humanos , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Proteína de la Leucemia Mieloide-Linfoide/genética , Nucleofosmina , Proteínas de Fusión Oncogénica/genética , Pronóstico , Tasa de Supervivencia , Células Tumorales CultivadasRESUMEN
BACKGROUND: DNA copy number variations (CNVs) are a hallmark of cancer, and the current study aimed to demonstrate the profile of the CNVs for oral cavity squamous cell carcinoma (OSCC) and elucidate the clinicopathological associations and molecular mechanisms of a potential marker derived from CNVs, mixed-lineage leukemia translocated to chromosome 3 protein (MLLT3), in OSCC carcinogenesis. MATERIALS AND METHODS: CNVs in 37 OSCC tissue specimens were analyzed using a high-resolution microarray, the OncoScan array. Gene expression was analyzed by real-time polymerase chain reaction in 127 OSCC and normal tissue samples. Cell function assays included cell cycle, migration, invasion and chromatin immunoprecipitation assays. RESULTS: We found a novel copy number amplified region, chromosome 9p, encompassing MLLT3 via the comparison of our data set with six other OSCC genome-wide CNV data sets. MLLT3 overexpression was associated with poorer overall survival in patients with OSCC (p = .048). MLLT3 knockdown reduced cell migration and invasion. The reduced invasion ability in MLLT3-knockdown cells was rescued with double knockdown of MLLT3 and CBP/p300-interacting transactivator with ED rich carboxy-terminal domain 4 (CITED4; 21.0% vs. 61.5%). Knockdown of MLLT3 impaired disruptor of telomeric silencing-1-like (Dot1L)-associated hypermethylation in the promoter of the tumor suppressor, CITED4 (p < .001), and hence dysregulated HIF-1α-mediated genes (TWIST, MMP1, MMP2, VIM, and CDH1) in OSCC cells. CONCLUSION: We identified unique CNVs in tumors of Taiwanese patients with OSCC. Notably, MLLT3 overexpression is related to the poorer prognosis of patients with OSCC and is required for Dot1L-mediated transcriptional repression of CITED4, leading to dysregulation of HIF-1α-mediated genes. IMPLICATIONS FOR PRACTICE: This article reports unique copy number variations in oral cavity squamous cell carcinoma (OSCC) tumors of Taiwanese patients. Notably, MLLT3 overexpression is related to the poorer prognosis of patients with OSCC and is required for Dot1L-mediated transcriptional repression of CITED4, leading to dysregulation of HIF-1α-mediated genes.
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Variaciones en el Número de Copia de ADN , Neoplasias de la Boca/genética , Proteínas Nucleares/genética , Carcinoma de Células Escamosas de Cabeza y Cuello/genética , Línea Celular Tumoral , Movimiento Celular/genética , Femenino , Humanos , Masculino , Persona de Mediana Edad , Neoplasias de la Boca/patología , Invasividad Neoplásica , Análisis de Secuencia por Matrices de Oligonucleótidos , Carcinoma de Células Escamosas de Cabeza y Cuello/patología , TransfecciónRESUMEN
YEATS domain (YD) containing proteins are an emerging class of epigenetic targets in drug discovery. Dysregulation of these modified lysine-binding proteins has been linked to the onset and progression of cancers. We herein report the discovery and characterisation of the first small-molecule chemical probe, SGC-iMLLT, for the YD of MLLT1 (ENL/YEATS1) and MLLT3 (AF9/YEATS3). SGC-iMLLT is a potent and selective inhibitor of MLLT1/3-histone interactions. Excellent selectivity over other human YD proteins (YEATS2/4) and bromodomains was observed. Furthermore, our probe displays cellular target engagement of MLLT1 and MLLT3. The first small-molecule X-ray co-crystal structures with the MLLT1 YD are also reported. This first-in-class probe molecule can be used to understand MLLT1/3-associated biology and the therapeutic potential of small-molecule YD inhibitors.
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Proteínas de Neoplasias/antagonistas & inhibidores , Proteínas de Neoplasias/química , Proteínas Nucleares/antagonistas & inhibidores , Proteínas Nucleares/química , Bibliotecas de Moléculas Pequeñas/química , Factores de Transcripción/antagonistas & inhibidores , Factores de Transcripción/química , Cristalografía por Rayos X , Histonas/metabolismo , Humanos , Simulación del Acoplamiento Molecular , Proteínas de Neoplasias/metabolismo , Proteínas Nucleares/metabolismo , Dominios Proteicos , Mapas de Interacción de Proteínas/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/farmacología , Factores de Transcripción/metabolismoRESUMEN
Pediatric acute myeloid leukemia (AML) is a highly heterogeneous disease, presenting cytogenetic and molecular abnormalities which turned out to be critical prognostic factors. Ploidy changes as gain or loss of individual chromosomes are rare in AML, occurring only in about 1-2% of the affected children. Hyperdiploid karyotypes are exceedingly rare in infants less than 12 months of age. In this age group, structural rearrangements involving the KMT2A gene occur in about 58% of the cases. Among them, the translocation t(9;11)(p22;q23), KMT2A-MLLT3, is the most common abnormality accounting for approximately 22% of KMT2A rearrangements in infant AML cases. Here, we describe a 7- month-old girl with a history of fever and severe diarrhea, and a physical examination remarkable for pallor and hepatosplenomegaly. A novel complex hyperdiploid karyotype 53,XX,+X,+6,t(9;11)(p21.3;q23.3),+der(9)t(9;11)(p21.3;q23.3),dup(13)(q31q34),+14,+19,+21,+22 was characterized by high-resolution molecular cytogenetic approaches. Fluorescence in situ hybridization, multiplex-FISH, and multicolor chromosome banding were applied, revealing 2 reverse MLLT3-KMT2A fusions and a duplication of the GAS6 oncogene. Our work suggests that molecular cytogenetic studies are crucial for the planning of a proper strategy for risk therapy in AML infants with hyperdiploid karyotypes.
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Duplicación Cromosómica , Análisis Citogenético/métodos , Diploidia , Péptidos y Proteínas de Señalización Intercelular/genética , Cariotipo , Leucemia Mieloide Aguda/genética , Proteínas Nucleares/genética , Oncogenes , Femenino , Reordenamiento Génico , N-Metiltransferasa de Histona-Lisina/genética , Humanos , Lactante , Proteína de la Leucemia Mieloide-Linfoide/genética , Translocación GenéticaAsunto(s)
Linfoma de Burkitt , N-Metiltransferasa de Histona-Lisina , Proteína de la Leucemia Mieloide-Linfoide , Proteínas de Fusión Oncogénica , Leucemia-Linfoma Linfoblástico de Células Precursoras , Enfermedad Aguda , Linfocitos B , N-Metiltransferasa de Histona-Lisina/genética , Humanos , Proteína de la Leucemia Mieloide-Linfoide/genética , Proteínas Nucleares , Proteínas de Fusión Oncogénica/genéticaRESUMEN
OBJECTIVES: Patients with acute myeloid leukaemia (AML) of the monocytic lineage often lack molecular markers for minimal residual disease (MRD) monitoring. The MLL-MLLT3 fusion transcript found in patients with AML harbouring t(9;11) is amenable to RT-qPCR quantification but because of the heterogeneity of translocation break points, the MLL-MLLT3 fusion gene is a challenging target. We hypothesised that MRD monitoring using MLL-MLLT3 as a RT-qPCR marker is feasible in the majority of patients with t(9;11)-positive AML. METHODS: Using a locked nucleic acid probe, we developed a sensitive RT-qPCR assay for quantification of the most common break point region of the MLL-MLLT3 fusion gene. Five paediatric patients with t(9;11)-positive AML were monitored using the MLL-MLLT3 assay. RESULTS: A total of 43 bone marrow (BM) and 52 Peripheral blood (PB) samples were collected from diagnosis until follow-up. Two patients relapsed, and both were MRD positive in BM after first induction course. A total of three relapses occurred, and they were detected by RT-qPCR 3 wks before haematological relapse was diagnosed. CONCLUSION: This MLL-MLLT3 RT-qPCR assay could be useful in MRD monitoring of a group of patients with AML who often lack reliable MRD markers.
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Biomarcadores de Tumor/genética , Leucemia Mieloide Aguda/diagnóstico , Neoplasia Residual/diagnóstico , Proteínas de Fusión Oncogénica/genética , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Antineoplásicos/uso terapéutico , Médula Ósea/química , Niño , Preescolar , Cromosomas Humanos Par 11 , Cromosomas Humanos Par 9 , Sondas de ADN/química , Sondas de ADN/genética , Humanos , Lactante , Recién Nacido , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/genética , Leucocitos Mononucleares/química , Neoplasia Residual/tratamiento farmacológico , Neoplasia Residual/genética , Oligonucleótidos/química , Oligonucleótidos/genética , Recurrencia , Inducción de Remisión , Translocación GenéticaRESUMEN
INTRODUCTION: AML patients with KMT2A-MLLT3 and other 11q23 abnormalities belong to the intermediate and high-risk level groups, respectively. Whether the poor prognostic value of Ecotropic Viral Integration site-1 (EVI1) overexpression suits either the subtypes of KMT2A-MLLT3 or Non-KMT2A-MLLT3 AML patients (intermediate and high risk group) needs to be further investigated. METHODS: We retrospectively analyzed the clinical characteristics of 166 KMT2A-r and KMT2A-PTD AML patients. RESULTS: For the Non-KMT2A-MLLT3 group, patients in the EVI1-high subgroup had shorter OS and DFS and higher CIR than those in the EVI1-low subgroup (p = .027, p = .018, and p = .020, respectively). Additionally, both KMT2A-MLLT3 and Non-KMT2A-MLLT3 patients who received chemotherapy alone had poorer prognosis than patients who also received allogeneic hematopoietic stem cell transplant (allo-HSCT) regardless of their EVI1 expression level (all p < .001). For transplanted patients with KMT2A-MLLT3 or Non-KMT2A-MLLT3 rearrangement, the EVI1-high subgroup had worse prognosis than the EVI1-low subgroup (all p < .05). The 2-year CIR of the KMT2A-MLLT3 and Non-KMT2A-MLLT3 groups with high EVI1 expression was high (52% and 49.6%, respectively). However, for patients with low EVI1 expression, the 2-year CIR of transplanted patients with KMT2A-MLLT3 and Non-KMT2A-MLLT3 was relatively low. CONCLUSIONS: Our study showed that for the Non-KMT2A-MLLT3 group, the EVI1-high group had shorter OS and DFS than the EVI1-low group. High EVI1 expression showed an adverse effect in AML with KMT2A rearrangement in different risk stratification subtypes. For the EVI1-high patients with non-KMT2A-MLLT3 rearrangement, other novel regimens towards relapse should be taken into consideration.
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Leucemia Mieloide Aguda , Proteína del Locus del Complejo MDS1 y EV11 , Humanos , Reordenamiento Génico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/terapia , Proteína del Locus del Complejo MDS1 y EV11/genética , Proteína del Locus del Complejo MDS1 y EV11/uso terapéutico , Pronóstico , Estudios Retrospectivos , Medición de Riesgo , Integración Viral , Trasplante de Células Madre Hematopoyéticas/métodos , Expresión GénicaRESUMEN
We present a patient with acute megakaryoblastic leukemia (AMKL) harboring KMT2A-MLLT3 that converted to T cell acute lymphoblastic leukemia (T-ALL) at her fourth relapse. A 4-year-old girl developed AMKL with multiple swollen lymph nodes. She exhibited several recurrences in the bone marrow and died of septic shock after her fourth relapse. Bone marrow cells at the initial diagnosis and at all four relapses had the same KMT2A-MLLT3 fusion transcript. She also developed a somatic mutation (c.7177C > T p.Q2393X) of NOTCH1 at the fourth relapse. This sequential phenotypic and cytogenetic study may yield valuable insights into the mechanism of AMKL to T-ALL lineage switch and possible implications for treatment selection.
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Trasplante de Células Madre Hematopoyéticas , Leucemia Megacarioblástica Aguda , Leucemia-Linfoma Linfoblástico de Células T Precursoras , Niño , Preescolar , Femenino , Humanos , Leucemia Megacarioblástica Aguda/genética , Leucemia Megacarioblástica Aguda/terapia , Proteínas Nucleares/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/terapia , Recurrencia , Linfocitos TRESUMEN
AF9 (MLLT3) and ENL (MLLT1) are members of the YEATS family (named after the five proteins first shown to contain this domain: Yaf9, ENL, AF9, Taf14, Sas5) defined by the presence of a YEATS domain. The YEATS domain is an epigenetic reader that binds to acetylated and crotonylated lysines, unlike the bromodomain which can only bind to acetylated lysines. All members of this family have been shown to be components of various complexes with roles in chromatin remodeling, histone modification, histone variant deposition, and transcriptional regulation. MLLT3 is a critical regulator of hematopoiesis with a role in maintaining the hematopoietic stem or progenitor cell (HSPC) population. Approximately 10% of acute myeloid leukemia (AML) and acute lymphocytic leukemia (ALL) patients harbor a translocation involving MLL (mixed lineage leukemia). In the context of MLL fusion patients with AML and ALL, MLL-AF9 and MLL-ENL fusions are observed in 34 and 31% of the patients, respectively. The intrinsically disordered C-terminal domain of MLLT3 (AHD, ANC1 homology domain) undergoes coupled binding and folding upon interaction with partner proteins AF4, DOT1L, BCOR, and CBX8. Backbone dynamics studies of the complexes suggest a role for dynamics in function. Inhibitors of the interaction of the intrinsically disordered AHD with partner proteins have been described, highlighting the feasibility of targeting intrinsically disordered regions. MLLT1 undergoes phase separation to enhance recruitment of the super elongation complex (SEC) and drive transcription. Mutations in MLLT1 observed in Wilms tumor patients enhance phase separation and transcription to drive an aberrant gene expression program.
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Hematopoyesis/fisiología , Neoplasias Renales/genética , Proteínas de Neoplasias/fisiología , Proteínas Nucleares/fisiología , Factores de Transcripción/fisiología , Tumor de Wilms/genética , Fusión Génica , Humanos , Proteínas Intrínsecamente Desordenadas/química , Proteínas Intrínsecamente Desordenadas/genética , Proteínas Intrínsecamente Desordenadas/metabolismo , Leucemia Bifenotípica Aguda/genética , Mutación , Proteínas de Neoplasias/química , Proteínas Nucleares/química , Dominios y Motivos de Interacción de Proteínas , Factores de Transcripción/químicaRESUMEN
The association between mature B-cell phenotype and KMT2A rearrangements in acute lymphoblastic leukemia is a very rare finding. It identifies a group of patients with similar clinical and biological characteristics that clearly differs from the entity B-cell lymphoblastic leukemia/lymphoma with t(v;11q23)/KMT2A-rearranged, which typically presents an immature pro B-cell phenotype. We describe the clinical-biological characteristics and disease outcome of three pediatric ALL patients with these features treated at our institution, and review 28 cases described in the literature. Most cases occur in children under 2 years-old, presenting a mature B-cell phenotype that uniformly expresses cytoplasmic and surface IgM with lambda light chain restriction, with heterogeneous co-expression of immaturity antigens. Patients do not have MYC rearrangements and all show KMT2A abnormalities, with 76% presenting t(9;11)(p21;q23)/MLLT3-KMT2A. These patients have an unfavorable clinical outcome and a 48% relapse rate. In-depth knowledge of this disease entity is needed to improve outcome.
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Leucemia-Linfoma Linfoblástico de Células Precursoras , Linfocitos B , Humanos , Lactante , FenotipoRESUMEN
BACKGROUND/AIM: Fusion of histone-lysine N-methyltransferase 2A gene (KMT2A) with the Rho guanine nucleotide exchange factor 12 gene (ARHGEF12), both located in 11q23, was reported in some leukemic patients. We report a KMT2A-ARHGEF12 fusion occurring during treatment of a pediatric acute myeloid leukemia (AML) with topoisomerase II inhibitors leading to a secondary acute lymphoblastic leukemia (ALL). MATERIALS AND METHODS: Multiple genetic analyses were performed on bone marrow cells of a girl initially diagnosed with AML. RESULTS: At the time of diagnosis with AML, the t(9;11)(p21;q23)/KMT2A-MLLT3 genetic abnormality was found. After chemotherapy resulting in AML clinical remission, a 2 Mb deletion in 11q23 was found generating a KMT2A-ARHGEF12 fusion gene. When the patient later developed B lineage ALL, a t(14;19)(q32;q13), loss of one chromosome 9, and KMT2A-ARHGEF12 were detected. CONCLUSION: The patient sequentially developed AML and ALL with three leukemia-specific genomic abnormalities in her bone marrow cells, two of which were KMT2A-rearrangements.
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Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamiento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Niño , Cromosomas Humanos Par 11 , Femenino , Eliminación de Gen , Fusión Génica , Factores de Intercambio de Guanina Nucleótido/genética , N-Metiltransferasa de Histona-Lisina/genética , Humanos , Leucemia Mieloide Aguda/patología , Proteína de la Leucemia Mieloide-Linfoide/genética , Proteínas Nucleares/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologíaRESUMEN
Eleven-nineteen leukemia (ENL) contains an epigenetic reader domain (YEATS domain) that recognizes lysine acylation on histone 3 and facilitates transcription initiation and elongation through its interactions with the super elongation complex (SEC) and the histone methyl transferase DOT1L. Although it has been known for its role as a fusion protein in mixed lineage leukemia (MLL), overexpression of native ENL, and thus dysregulation of downstream genes in acute myeloid leukemia (AML), has recently been implicated as a driver of disease that is reliant on the epigenetic reader activity of the YEATS domain. We developed a peptide displacement assay (histone 3 tail with acylated lysine) and screened a small-molecule library totaling more than 24,000 compounds for their propensity to disrupt the YEATS domain-histone peptide binding. Among these, we identified a first-in-class dual inhibitor of ENL ( Kd = 745 ± 45 nM) and its paralog AF9 ( Kd = 523 ± 53 nM) and performed "SAR by catalog" with the aim of starting the development of a chemical probe for ENL.
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Descubrimiento de Drogas , Factores de Elongación Transcripcional/antagonistas & inhibidores , Factores de Elongación Transcripcional/química , Fenómenos Biofísicos , Evaluación Preclínica de Medicamentos , Células HEK293 , Histonas/metabolismo , Humanos , Concentración 50 Inhibidora , Péptidos/metabolismo , Dominios Proteicos , Relación Estructura-ActividadRESUMEN
Osteosarcoma was the most common malignant tumor derived from bone/soft tissues and afflicted mainly the adolescents. Surgical removing assisted with chemotherapy/radiotherapy was the major therapeutics regardless of high mortality. MLLT3 was one of the most common partners fused with mixed lineage leukemia gene and caused leukemia. But its role in osteosarcoma was not previously reported. In this study, we found that MLLT3 was expressed abundantly in osteosarcoma tissues and cell lines including U-2OS, MG-63, Saos-2 and HOS. And by Kaplan-Meier analysis, MLLT3 expression was shown to be clinically correlated with survival of osteosarcoma patients. The 5-year survival rate was 64.3% for patients with low MLLT3 expression (n=14) while it was 36.4% for patients with high MLLT3 expression (n=11). The difference was significant (p=0.026). When MLLT3 was knocked-down, the proliferation ability decreased about 38% in HOS cells and 46% in MG-63 cells. But the migration as well as the invasive ability was not affected. In addition, the expression level of ß-catenin was comparable in HOS cells with/without knockdown of MLLT3. The specific inhibitor LGK-974 against Wnt/ß-catenin signaling showed little additive effect with MLLT3 knockdown on growth ability of HOS cells. But the level of c-Jun, BCL-2, Akt and c-Myc was significantly decreased in HOS cells with MLLT3 knockdown, which indicated that MLLT3 may at least partially regulate JNK signaling in HOS cells. In conclusion, we proved MLLT3 as an oncogene in osteosarcoma through partially regulating the JNK signaling pathway. And MLLT3 may be a promising target for treatment of patients with osteosarcoma.
RESUMEN
Epidemiological data have linked birth control formulations to an increased risk of infant acute leukemia involving MLL rearrangements. Reverse transcription polymerase chain reaction (RT-PCR) studies showed that 10 nM estradiol enhanced MLL transcription in addition to its common translocation partners, MLLT2 (AF4) and MLLT3 (AF9). The same concentration of estradiol triggered MLL and MLLT3 co-localization without affecting the interaction of genes located on the same chromosomes. Estradiol also stimulated the generation of MLL-MLLT3 fusion transcripts as seen by RT-PCR. RNAi knockdown of activation-induced cytidine deaminase (AICDA) suppressed the induction of MLL-MLLT3 fusion transcript formation observed with estradiol. Additionally, chromatin immunoprecipitation (ChIP) analysis showed estradiol dependent localization of AICDA in MLL intron 11, upstream of a hotspot for both DNA cleavage and rearrangement, but not downstream within intron 12. Combined, these studies show that levels of estradiol consistent with that observed during pregnancy have the potential to initiate MLL fusions through an AICDA-mediated mechanism.