RESUMEN
Acute promyelocytic leukemia (APL) is characterized by a specific t(15;17) chromosomal translocation that yields the PML/RARA fusion gene. Clinically, besides chemotherapy, two drugs induce clinical remissions: retinoic acid (RA) and arsenic trioxide (As). Both agents directly target PML/RARA-mediated transcriptional repression and protein stability, inducing to various extent promyelocyte differentiation and clinical remission of APL patients. RA targets the RARA moiety of the fusion, whereas arsenic targets its PML part. PML/RARA expression in the mouse is sufficient to initiate APL. The RA-As association, which synergizes for PML/RARA degradation but not for differentiation, rapidly clears leukemia initiating cells (LIC), resulting in APL eradication in murine APL models, but also in several APL clinical trials. Cyclic AMP triggered PML/RARA phosphorylation also enhances RA-induced APL regression, PML/RARA degradation, and LIC clearance, raising new options for therapy-resistant patients. Although differentiation has a major role in debulking of the tumor, PML/RARA degradation seems to be the primary basis for APL eradication by the RA-As association. Oncoprotein degradation could be a general therapeutic strategy that may be extended beyond APL.
Asunto(s)
Leucemia Promielocítica Aguda/tratamiento farmacológico , Leucemia Promielocítica Aguda/metabolismo , Proteínas de Fusión Oncogénica/metabolismo , Animales , Sistemas de Liberación de Medicamentos , Sinergismo Farmacológico , Humanos , Leucemia Experimental/metabolismo , RatonesRESUMEN
Retinoic acid and arsenic trioxide target the protein stability and transcriptional repression activity of the fusion oncoprotein PML-RARA, resulting in regression of acute promyelocytic leukemia (APL). Phenotypically, retinoic acid induces differentiation of APL cells. Here we show that retinoic acid also triggers growth arrest of leukemia-initiating cells (LICs) ex vivo and their clearance in PML-RARA mouse APL in vivo. Retinoic acid treatment of mouse APLs expressing the fusion protein PLZF-RARA triggers full differentiation, but not LIC loss or disease remission, establishing that differentiation and LIC loss can be uncoupled. Although retinoic acid and arsenic synergize to clear LICs through cooperative PML-RARA degradation, this combination does not enhance differentiation. A cyclic AMP (cAMP)-dependent phosphorylation site in PML-RARA is crucial for retinoic acid-induced PML-RARA degradation and LIC clearance. Moreover, activation of cAMP signaling enhances LIC loss by retinoic acid, identifying cAMP as another potential APL therapy. Thus, whereas transcriptional activation of PML-RARA is likely to control differentiation, its catabolism triggers LIC eradication and long-term remission of mouse APL. Therapy-triggered degradation of oncoproteins could be a general strategy to eradicate cancer stem cells.
Asunto(s)
Leucemia Promielocítica Aguda/metabolismo , Proteínas de Fusión Oncogénica/metabolismo , Animales , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/metabolismo , Transformación Celular Neoplásica/patología , AMP Cíclico/farmacología , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Regulación Neoplásica de la Expresión Génica , Humanos , Leucemia Promielocítica Aguda/genética , Leucemia Promielocítica Aguda/patología , Ratones , Ratones Desnudos , Proteínas de Fusión Oncogénica/genética , Fosforilación , Serina/genética , Serina/metabolismo , Transducción de Señal , Tretinoina/farmacología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Leukemia-associated chimeric oncoproteins often act as transcriptional repressors, targeting promoters of master genes involved in hematopoiesis. We show that CRABPI (encoding cellular retinoic acid binding protein I) is a target of PLZF, which is fused to RARalpha by the t(11;17)(q23;q21) translocation associated with retinoic acid (RA)-resistant acute promyelocytic leukemia (APL). PLZF represses the CRABPI locus through propagation of chromatin condensation from a remote intronic binding element culminating in silencing of the promoter. Although the canonical, PLZF-RARalpha oncoprotein has no impact on PLZF-mediated repression, the reciprocal translocation product RARalpha-PLZF binds to this remote binding site, recruiting p300, inducing promoter hypomethylation and CRABPI gene up-regulation. In line with these observations, RA-resistant murine PLZF/RARalpha+RARalpha/PLZF APL blasts express much higher levels of CRABPI than standard RA-sensitive PML/RARalpha APL. RARalpha-PLZF confers RA resistance to a retinoid-sensitive acute myeloid leukemia (AML) cell line in a CRABPI-dependent fashion. This study supports an active role for PLZF and RARalpha-PLZF in leukemogenesis, identifies up-regulation of CRABPI as a mechanism contributing to retinoid resistance, and reveals the ability of the reciprocal fusion gene products to mediate distinct epigenetic effects contributing to the leukemic phenotype.
Asunto(s)
Cromosomas Humanos Par 11/genética , Cromosomas Humanos Par 17/genética , Factores de Transcripción de Tipo Kruppel/metabolismo , Leucemia Promielocítica Aguda/genética , Leucemia Promielocítica Aguda/metabolismo , Receptores de Ácido Retinoico/metabolismo , Secuencia de Bases , Sitios de Unión , Línea Celular , Cromatina/genética , Metilación de ADN , Progresión de la Enfermedad , Resistencia a Antineoplásicos/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica , Humanos , Leucemia Promielocítica Aguda/patología , Datos de Secuencia Molecular , Proteína de la Leucemia Promielocítica con Dedos de Zinc , Receptores de Ácido Retinoico/genética , Receptor alfa de Ácido Retinoico , Retinoides/farmacología , Regulación hacia ArribaRESUMEN
Calcineurin is a calcium-activated serine/threonine phosphatase critical to a number of developmental processes in the cardiovascular, nervous and immune systems. In the T-cell lineage, calcineurin activation is important for pre-T-cell receptor (TCR) signaling, TCR-mediated positive selection of thymocytes into mature T cells, and many aspects of the immune response. The critical role of calcineurin in the immune response is underscored by the fact that calcineurin inhibitors, such as cyclosporin A (CsA) and FK506, are powerful immunosuppressants in wide clinical use. We observed sustained calcineurin activation in human B- and T-cell lymphomas and in all mouse models of lymphoid malignancies analyzed. In intracellular NOTCH1 (ICN1)- and TEL-JAK2-induced T-cell lymphoblastic leukemia, two mouse models relevant to human malignancies, in vivo inhibition of calcineurin activity by CsA or FK506 induced apoptosis of leukemic cells and rapid tumor clearance, and substantially prolonged mouse survival. In contrast, ectopic expression of a constitutively activated mutant of calcineurin favored leukemia progression. Moreover, CsA treatment induced apoptosis in human lymphoma and leukemia cell lines. Thus, calcineurin activation is critical for the maintenance of the leukemic phenotype in vivo, identifying this pathway as a relevant therapeutic target in lymphoid malignancies.
Asunto(s)
Antineoplásicos/farmacología , Calcineurina/metabolismo , Leucemia-Linfoma de Células T del Adulto/tratamiento farmacológico , Leucemia-Linfoma de Células T del Adulto/enzimología , Animales , Inhibidores de la Calcineurina , Línea Celular Tumoral , Ciclosporina/farmacología , Modelos Animales de Enfermedad , Activación Enzimática/efectos de los fármacos , Humanos , Leucemia-Linfoma de Células T del Adulto/patología , Linfoma de Células B/tratamiento farmacológico , Linfoma de Células B/enzimología , Linfoma de Células B/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Proteínas de Fusión Oncogénica/deficiencia , Proteínas de Fusión Oncogénica/genética , Receptor Notch1/fisiología , Tacrolimus/farmacologíaRESUMEN
Differentiation therapy for acute myeloid leukemia uses transcriptional modulators to reprogram cancer cells. The most relevant clinical example is acute promyelocytic leukemia (APL), which responds dramatically to either retinoic acid (RA) or arsenic trioxide (As(2)O(3)). In many myeloid leukemia cell lines, cyclic adenosine monophosphate (cAMP) triggers growth arrest, cell death, or differentiation, often in synergy with RA. Nevertheless, the toxicity of cAMP derivatives and lack of suitable models has hampered trials designed to assess the in vivo relevance of theses observations. We show that, in an APL cell line, cAMP analogs blocked cell growth and unraveled As(2)O(3)-triggered differentiation. Similarly, in RA-sensitive or RA-resistant mouse models of APL, continuous infusions of 8-chloro-cyclic adenosine monophosphate (8-Cl-cAMP) triggered major growth arrest, greatly enhanced both spontaneous and RA- or As(2)O(3)-induced differentiation and accelerated the restoration of normal hematopoiesis. Theophylline, a well-tolerated phosphodiesterase inhibitor which stabilizes endogenous cAMP, also impaired APL growth and enhanced spontaneous or As(2)O(3)-triggered cell differentiation in vivo. Accordingly, in an APL patient resistant to combined RA-As(2)O(3) therapy, theophylline induced blast clearance and restored normal hematopoiesis. Taken together, these results demonstrate that in vivo activation of cAMP signaling contributes to APL clearance, independently of its RA-sensitivity, thus raising hopes that other myeloid leukemias may benefit from this therapeutic approach.