RESUMEN
Essential for cell survival, the 90 kD Heat Shock Proteins (HSP90) are molecular chaperons required for conformational stabilization and trafficking of numerous client proteins. Functional HSP90 is required for the stability of AKT, a serine-threonine kinase phosphorylated in response to growth factor stimulation. AKT plays a crucial regulatory role in differentiation, cell cycle, transcription, translation, metabolism and apoptosis. Acute promyelocytic leukaemia (APL) is characterized by the presence of the promyelocytic leukaemia/retinoic acid receptor alpha (PML/RARA) fusion protein, which deregulates expression of several genes involved in differentiation and apoptosis. Here, we report inhibition of HSP90AA1 and HSP90AB1 isomer transcription in blasts isolated from patients with APL, associated with reduction of HSP90 protein expression and loss of control on AKT protein phosphorylation. We show that in vitro treatment of PML/RARA expressing cells with all-trans retinoic acid (ATRA) up-regulates HSP90 expression and stabilizes AKT. Addition of the HSP90-inhibitor 17-(allylamino)-17-demethoxygeldanamycin in combination with ATRA, blocks upregulation of AKT protein, indicating that HSP90 is necessary for ATRA action on AKT. This is the first report proving that expression of HSP90 isomers are directly and differentially repressed by PML/RARA, with critical results on cellular homeostasis of target proteins, such as AKT, in APL blasts.
Asunto(s)
Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Leucemia Promielocítica Aguda/tratamiento farmacológico , Proteínas de Fusión Oncogénica/metabolismo , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Benzoquinonas/farmacología , Células HEK293 , Proteínas HSP90 de Choque Térmico/biosíntesis , Proteínas HSP90 de Choque Térmico/genética , Histonas/genética , Histonas/metabolismo , Humanos , Lactamas Macrocíclicas/farmacología , Leucemia Promielocítica Aguda/patología , Proteínas de Fusión Oncogénica/biosíntesis , Proteínas de Fusión Oncogénica/genética , Regiones Promotoras Genéticas , Proteína de la Leucemia Promielocítica/biosíntesis , Proteína de la Leucemia Promielocítica/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Receptor alfa de Ácido Retinoico/biosíntesis , Receptor alfa de Ácido Retinoico/genética , Transfección , Tretinoina/farmacología , Células Tumorales Cultivadas , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Overexpression, polymorphisms, and mutations of the WT1 gene have been reported in several human tumors including acute myeloid leukemia (AML) and variably correlated with prognosis. Acute promyelocytic leukemia (APL) represents the AML subset disclosing higher WT1 expression levels; however, no WT1 studies specifically focused on APL have been conducted. We screened for the presence of mutations, SNP rs16754, and expression levels of WT1 gene in 103 adult patients with newly diagnosed APL. Fms-like tyrosine kinase (FLT3) mutations were analyzed as well. WT1 mutations were identified in four (4 %) patients. At least one copy of the minor SNP rs16754 allele (WT1(AG) or WT1(GG)) was detected in 30 (29 %) patients. Six patients (6 %) were homozygous for the minor allele (WT1(GG)) and this genotype was associated with higher WT1 mRNA copies (p = 0.018). FLT3 mutations were found in 37 % of patients and correlated with high WT1 mRNA expression (p = 0.004). Patients heterozygous or homozygous for the minor allele and patients homozygous for major (WT1(AA)) allele did not differ in terms of presenting features. In adult APL, WT1 gene mutational and polymorphic profile shows similarities with pediatric AML rather than with adult AML.
Asunto(s)
Regulación Leucémica de la Expresión Génica , Leucemia Promielocítica Aguda/genética , Leucemia Promielocítica Aguda/fisiopatología , Mutación , Proteínas de Neoplasias/genética , Polimorfismo de Nucleótido Simple , Proteínas WT1/genética , Adulto , Codón sin Sentido , Estudios de Cohortes , Mutación del Sistema de Lectura , Estudios de Asociación Genética , Heterocigoto , Homocigoto , Humanos , Leucemia Promielocítica Aguda/metabolismo , Mutación Missense , Proteínas de Neoplasias/metabolismo , ARN Mensajero/metabolismo , Recurrencia , Ciudad de Roma , España , Análisis de Supervivencia , Proteínas WT1/metabolismoRESUMEN
Acute promyelocitic leukemia (APL) is characterized by the pathognomonic presence in leukemic blasts of the hybrid protein PML/RARA, that acts as a transcriptional repressor impairing the expression of genes that are critical to myeloid differentiation. Here, we show that primary blasts from APL patients express lower levels of the oncosuppressor protein PTEN, as compared to blast cells from other AML subtypes or normal bone marrow, and demonstrate that PML-RARA directly inhibits PTEN expression. We show that All-Trans Retinoic Acid (ATRA) triggers in APL cells an active chromatin status at the core regulatory region of the PTEN promoter, that allows the binding of the myeloid-regulating transcription factor PU.1, and, in turn, the transcriptional induction of PTEN. ATRA, via PML/RARA degradation, also promotes PTEN nuclear re-localization and decreases expression of the PTEN target Aurora A kinase. In conclusion, our findings support the notion that PTEN is one of the primary targets of PML/RARA in APL.