RESUMEN
Less than a third of adults patients with acute myeloid leukemia (AML) are cured by current treatments, emphasizing the need for new approaches to therapy. We previously demonstrated that besides playing a role in drug-resistant leukemia cell lines, multidrug resistance protein 4 (MRP4/ABCC4) regulates leukemia cell proliferation and differentiation through the endogenous MRP4/ABCC4 substrate, cAMP. Here, we studied the role of MRP4/ABCC4 in tumor progression in a mouse xenograft model and in leukemic stem cells (LSCs) differentiation. We found a decrease in the mitotic index and an increase in the apoptotic index associated with the inhibition of tumor growth when mice were treated with rolipram (PDE4 inhibitor) and/or probenecid (MRPs inhibitor). Genetic silencing and pharmacologic inhibition of MRP4 reduced tumor growth. Furthermore, MRP4 knockdown induced cell cycle arrest and apoptosis in vivo. Interestingly, when LSC population was isolated, we observed that increased cAMP levels and MRP4/ABCC4 blockade resulted in LSCs differentiation. Taken together, our findings show that MRP4/ABCC4 has a relevant role in tumor growth and apoptosis and in the eradication of LSCs, providing the basis for a novel promising target in AML therapy.
Asunto(s)
Apoptosis/genética , Puntos de Control del Ciclo Celular/genética , Leucemia Mieloide Aguda/patología , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Células Madre Neoplásicas/citología , Animales , Apoptosis/efectos de los fármacos , Diferenciación Celular , Línea Celular Tumoral , Proliferación Celular/genética , AMP Cíclico/metabolismo , Ciclina D1/metabolismo , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/biosíntesis , Progresión de la Enfermedad , Femenino , Humanos , Leucemia Mieloide Aguda/tratamiento farmacológico , Ratones , Ratones Desnudos , Índice Mitótico , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/antagonistas & inhibidores , Trasplante de Neoplasias , Inhibidores de Fosfodiesterasa 4/farmacología , Interferencia de ARN , ARN Interferente Pequeño/biosíntesis , ARN Interferente Pequeño/genética , Rolipram/farmacología , Trasplante HeterólogoRESUMEN
Increased intracellular cAMP concentration plays a well established role in leukemic cell maturation. We previously reported that U937 cells stimulated by H2 receptor agonists, despite a robust increase in cAMP, fail to mature because of rapid H2 receptor desensitization and phosphodiesterase (PDE) activation. Here we show that intracellular cAMP levels not only in U937 cells but also in other acute myeloid leukemia cell lines are also regulated by multidrug resistance-associated proteins (MRPs), particularly MRP4. U937, HL-60, and KG-1a cells, exposed to amthamine (H2-receptor agonist), augmented intracellular cAMP concentration with a concomitant increase in the efflux. Extrusion of cAMP was ATP-dependent and probenecid-sensitive, supporting that the transport was MRP-mediated. Cells exposed to amthamine and the PDE4 inhibitor showed enhanced cAMP extrusion, but this response was inhibited by MRP blockade. Amthamine stimulation, combined with PDE4 and MRP inhibition, induced maximal cell arrest proliferation. Knockdown strategy by shRNA revealed that this process was mediated by MRP4. Furthermore, blockade by probenecid or MRP4 knockdown showed that increased intracellular cAMP levels induce maturation in U937 cells. These findings confirm the key role of intracellular cAMP levels in leukemic cell maturation and provide the first evidence that MRP4 may represent a new potential target for leukemia differentiation therapy.
Asunto(s)
AMP Cíclico/metabolismo , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patología , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Transducción de Señal/fisiología , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/fisiología , División Celular/efectos de los fármacos , División Celular/fisiología , Diseño de Fármacos , Células HL-60 , Humanos , Leucemia Mieloide Aguda/tratamiento farmacológico , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/antagonistas & inhibidores , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Inhibidores de Fosfodiesterasa 4/farmacología , Probenecid/farmacología , ARN Interferente Pequeño , Rolipram/farmacología , Transducción de Señal/efectos de los fármacos , Tiazoles/farmacología , Células U937RESUMEN
OBJECTIVE: To evaluate if routine antenatal screening for congenital syphilis (CS) was adequately implemented. DESIGN: Retrospective study. SETTING: Curaçao, St. Elisabeth Hospital. METHOD: From 1987-1991 16 infants were treated for congenital syphilis in the paediatric department of the St. Elisabeth Hospital. From hospital and lab records, syphilis serology of their mothers before and during pregnancy and at delivery were indexed as well as cord blood values. The response in case of positive syphilis serology was traced. RESULTS: During the evaluation period the congenital syphilis incidence was 1.1/1000 life born infants. 9 pregnant women avoided prenatal care. Despite positive syphilis serology in the 1st (1 patient) and 3rd trimester (4 patients) no action was undertaken. In 4 neonates with congenital syphilis no cord blood sample for screening was taken. On 2 occasions the cord blood RPR was false-negative. (Re)screening was not performed at delivery in 3 mothers although positive serology was found during pregnancy. CONCLUSION: Screening for congenital syphilis was not always applied. Insufficient action was noted if positive syphilis serology was detected. Intensification of screening for congenital syphilis in Curaçao is necessary especially for mothers with poor prenatal care. The need for immediate post partum screening for mother and child is stressed.