RESUMEN
The 14-3-3/c-Abl protein-protein interaction (PPI) is related to carcinogenesis and in particular to pathogenesis of chronic myeloid leukemia (CML). Previous studies have demonstrated that molecules able to disrupt this interaction improve the nuclear translocation of c-Abl, inducing apoptosis in leukemia cells. Through an X-ray crystallography screening program, we have identified two phosphate-containing compounds, inosine monophosphate (IMP) and pyridoxal phosphate (PLP), as binders of human 14-3-3σ, by targeting the protein amphipathic groove. Interestingly, they also act as weak inhibitors of the 14-3-3/c-Abl PPI, demonstrated by NMR, SPR, and FP data. A 37-compound library of PLP and IMP analogues was investigated using a FP assay, leading to the identification of three further molecules acting as weak inhibitors of the 14-3-3/c-Abl complex formation. The antiproliferative activity of IMP, PLP, and the three derivatives was tested against K-562 cells, showing that the parent compounds had the most pronounced effect on tumor cells. PLP and IMP were also effective in promoting the c-Abl nuclear translocation in c-Abl overexpressing cells. Further, these compounds demonstrated low cytotoxicity on human Hs27 fibroblasts. In conclusion, our data suggest that 14-3-3σ targeting compounds represent promising hits for further development of drugs against c-Abl-dependent cancers.
Asunto(s)
Proteínas 14-3-3/antagonistas & inhibidores , Exorribonucleasas/antagonistas & inhibidores , Organofosfatos/farmacología , Unión Proteica/efectos de los fármacos , Proteínas Proto-Oncogénicas c-abl/antagonistas & inhibidores , Bibliotecas de Moléculas Pequeñas/farmacología , Proteínas 14-3-3/química , Proteínas 14-3-3/metabolismo , Secuencia de Aminoácidos , Antineoplásicos/metabolismo , Antineoplásicos/farmacología , Antineoplásicos/toxicidad , Núcleo Celular/metabolismo , Cristalografía por Rayos X , Ensayos de Selección de Medicamentos Antitumorales , Exorribonucleasas/química , Exorribonucleasas/metabolismo , Humanos , Inosina Monofosfato/metabolismo , Inosina Monofosfato/farmacología , Inosina Monofosfato/toxicidad , Células K562 , Organofosfatos/metabolismo , Organofosfatos/toxicidad , Proteínas Proto-Oncogénicas c-abl/metabolismo , Fosfato de Piridoxal/metabolismo , Fosfato de Piridoxal/farmacología , Fosfato de Piridoxal/toxicidad , Alineación de Secuencia , Bibliotecas de Moléculas Pequeñas/toxicidadRESUMEN
14-3-3 are regulatory proteins that through protein-protein interactions (PPI) with numerous binding partners could be involved in several human diseases, including cancer, neurodegenerative disorders, and pathogens infections. Following our research interest in the development of 14-3-3 PPI inhibitors, here we exploited the privileged 4-aminoantipyrine scaffold in the design and synthesis of some derivatives endowed with antiproliferative activity against K-562 cells, and capable of binding to recombinant 14-3-3σ as evidenced by NMR spectroscopy. The binding mode was further explored by molecular modelling, while coupling confocal microscopy with intensitometric analysis showed that compound 1 was able to promote the nuclear translocation of c-Abl at low micromolar concentrations. Overall, 1 is chemically stable compared to parent 14-3-3 PPI inhibitors, and thus emerged as a confirmed hit for further development.