RESUMEN
Phosphoinositide 3-kinase (PI3K)/protein kinase B/Akt and Ras/mitogen-activated protein kinase pathways are often constitutively activated in melanoma and have thus been considered as promising drug targets. Exposure of melanoma cells to NVP-BAG956, NVP-BBD130, and NVP-BEZ235, a series of novel, potent, and stable dual PI3K/mammalian target of rapamycin (mTOR) inhibitors, resulted in complete G1 growth arrest, reduction of cyclin D1, and increased levels of p27(KIP1), but negligible apoptosis. In contrast, treatment of melanoma with the pan-class I PI3K inhibitor ZSTK474 or the mTORC1 inhibitor rapamycin resulted only in minor reduction of cell proliferation. In a syngeneic B16 mouse melanoma tumor model, orally administered NVP-BBD130 and NVP-BEZ235 efficiently attenuated tumor growth at primary and lymph node metastatic sites with no obvious toxicity. Metastatic melanoma in inhibitor-treated mice displayed reduced numbers of proliferating and significantly smaller tumor cells. In addition, neovascularization was blocked and tumoral necrosis increased when compared with vehicle-treated mice. In conclusion, compounds targeting PI3K and mTOR simultaneously were advantageous to attenuate melanoma growth and they develop their potential by targeting tumor growth directly, and indirectly via their interference with angiogenesis. Based on the above results, NVP-BEZ235, which has entered phase I/II clinical trials in patients with advanced solid tumors, has a potential in metastatic melanoma therapy.
Asunto(s)
Imidazoles/farmacología , Melanoma Experimental/tratamiento farmacológico , Inhibidores de las Quinasa Fosfoinosítidos-3 , Proteínas Quinasas/metabolismo , Quinolinas/farmacología , Sirolimus/farmacología , Triazinas/farmacología , Proteínas Quinasas Dependientes de 3-Fosfoinosítido , Administración Oral , Animales , Antibióticos Antineoplásicos/farmacología , Apoptosis/fisiología , Ciclo Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Sinergismo Farmacológico , Femenino , Proteína Forkhead Box O1 , Factores de Transcripción Forkhead/metabolismo , Humanos , Immunoblotting , Técnicas para Inmunoenzimas , Melanoma Experimental/patología , Ratones , Ratones Endogámicos C57BL , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Transporte de Proteínas , Serina-Treonina Quinasas TOR , Células Tumorales CultivadasRESUMEN
Phosphoinositides (PIs) play fundamental roles as signalling molecules in numerous cellular processes. Direct analysis of PIs is typically accomplished by metabolic labelling with (3)H-inositol or inorganic (32)P followed by deacylation, ion-exchange chromatography and flow scintillation detection. This analysis is laborious, time-consuming, and involves massive amounts of radioactivity. To overcome these limitations we established a robust, non-radioactive LC-ESI-MS assay for the separation and analysis of deacylated PIs that allows discrimination of all isomers without the need for radioactive labelling. We applied the method to various cell types to study the PI levels upon specific stimulation.