RESUMO
Polyphenols are natural compounds widely present in fruits and vegetables, which have antimutagenic and anticancer properties. The aim of the present study was to determine the anticancer effect of a polyphenol-rich Aronia melanocarpa juice (AMJ) containing 7.15 g/L of polyphenols in the acute lymphoblastic leukemia Jurkat cell line, and, if so, to clarify the underlying mechanism and to identify the active polyphenols involved. AMJ inhibited cell proliferation, which was associated with cell cycle arrest in G(2)/M phase, and caused the induction of apoptosis. These effects were associated with an upregulation of the expression of tumor suppressor p73 and active caspase 3, and a downregulation of the expression of cyclin B1 and the epigenetic integrator UHRF1. AMJ significantly increased the formation of reactive oxygen species (ROS), decreased the mitochondrial membrane potential and caused the release of cytochrome c into the cytoplasm. Treatment with intracellular ROS scavengers prevented the AMJ-induced apoptosis and upregulation of the expression of p73 and active caspase 3. The fractionation of the AMJ and the use of identified isolated compounds indicated that the anticancer activity was associated predominantly with chlorogenic acids, some cyanidin glycosides, and derivatives of quercetin. AMJ treatment also induced apoptosis of different human lymphoblastic leukemia cells (HSB-2, Molt-4 and CCRF-CEM). In addition, AMJ exerted a strong pro-apoptotic effect in human primary lymphoblastic leukemia cells but not in human normal primary T-lymphocytes. Thus, the present findings indicate that AMJ exhibits strong anticancer activity through a redox-sensitive mechanism in the p53-deficient Jurkat cells and that this effect involves several types of polyphenols. They further suggest that AMJ has chemotherapeutic properties against acute lymphoblastic leukemia by selectively targeting lymphoblast-derived tumor cells.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Caspase 3/metabolismo , Proteínas de Ligação a DNA/metabolismo , Leucemia/metabolismo , Proteínas Nucleares/metabolismo , Photinia/química , Extratos Vegetais/farmacologia , Proteínas Supressoras de Tumor/metabolismo , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Citocromos c/metabolismo , Proteínas de Ligação a DNA/genética , Humanos , Células Jurkat , Linfócitos/efeitos dos fármacos , Linfócitos/imunologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Proteínas Nucleares/genética , Oxirredução , Polifenóis/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Proteína Tumoral p73 , Proteínas Supressoras de Tumor/genética , Ubiquitina-Proteína LigasesRESUMO
Nuclear receptors (NRs) are ligand-inducible transcription factors that regulate a plethora of cell biological phenomena, thus orchestrating complex events like development, organ homeostasis, immune function, and reproduction. Due to their regulatory potential, NRs are major drug targets for a variety of diseases, including cancer and metabolic diseases, and had a major societal impact following the development of contraceptives and abortifacients. Not surprisingly in view of this medical and societal importance, a large amount of diverse NR ligands have been generated and the corresponding structural and functional analyses have provided a deep insight into the molecular basis of ligand action. What we have learnt is that ligands regulate, via allosteric conformational changes, the ability of NRs to interact with different sets of coregulators which in turn recruit enzymatically active complexes, the workhorses of the ligand-induced epigenetic and transcription-regulatory events. Thus, ligands essentially direct the communication of a given NR with its intracellular environment at the chromatin and extragenomic level to modulate gene programs directly at the chromatin level or via less well-understood extranuclear actions. Here we will review our current structural and mechanistic insight into the functionalities of subsets of retinoid and rexinoid ligands that act generically as antagonists but follow different mechanistic principles, resulting in "classical" or neutral antagonism, or inverse agonism. In addition, we describe the chemical features and guidelines for the synthesis of retinoids/rexinoids that exert specific functions and we provide protocols for a number of experimental approaches that are useful for studies of the agonistic and antagonistic features of NR ligands.
Assuntos
Desenho de Fármacos , Agonismo Inverso de Drogas , Receptores do Ácido Retinoico/antagonistas & inibidores , Receptores do Ácido Retinoico/metabolismo , Animais , Linhagem Celular , Avaliação Pré-Clínica de Medicamentos/métodos , Polarização de Fluorescência/métodos , Humanos , Ligantes , Modelos Moleculares , Peptídeo Hidrolases/metabolismo , Preparações Farmacêuticas/síntese química , Receptores do Ácido Retinoico/química , Receptores do Ácido Retinoico/genética , Retinoides/agonistas , Retinoides/antagonistas & inibidores , Retinoides/síntese química , Retinoides/química , Espectrometria de Massas por Ionização por Electrospray/métodos , Transfecção/métodosRESUMO
Amphioxus, the closest living invertebrate relative of the vertebrates, has a notochord, segmental axial musculature, pharyngeal gill slits and dorsal hollow nerve cord, but lacks neural crest. In amphioxus, as in vertebrates, exogenous retinoic acid (RA) posteriorizes the embryo. The mouth and gill slits never form, AmphiPax1, which is normally downregulated where gill slits form, remains upregulated and AmphiHox1 expression shifts anteriorly in the nerve cord. To dissect the role of RA signaling in patterning chordate embryos, we have cloned the single retinoic acid receptor (AmphiRAR), retinoid X receptor (AmphiRXR) and an orphan receptor (AmphiTR2/4) from amphioxus. AmphiTR2/4 inhibits AmphiRAR-AmphiRXR-mediated transactivation in the presence of RA by competing for DR5 or IR7 retinoic acid response elements (RAREs). The 5' untranslated region of AmphiTR2/4 contains an IR7 element, suggesting possible auto- and RA-regulation. The patterns of AmphiTR2/4 and AmphiRAR expression during embryogenesis are largely complementary: AmphiTR2/4 is strongly expressed in the cerebral vesicle (homologous to the diencephalon plus anterior midbrain), while AmphiRAR expression is high in the equivalent of the hindbrain and spinal cord. Similarly, while AmphiTR2/4 is expressed most strongly in the anterior and posterior thirds of the endoderm, the highest AmphiRAR expression is in the middle third. Expression of AmphiRAR is upregulated by exogenous RA and completely downregulated by the RA antagonist BMS009. Moreover, BMS009 expands the pharynx posteriorly; the first three gill slit primordia are elongated and shifted posteriorly, but do not penetrate, and additional, non-penetrating gill slit primordia are induced. Thus, in an organism without neural crest, initiation and penetration of gill slits appear to be separate events mediated by distinct levels of RA signaling in the pharyngeal endoderm. Although these compounds have little effect on levels of AmphiTR2/4 expression, RA shifts pharyngeal expression of AmphiTR2/4 anteriorly, while BMS009 extends it posteriorly. Collectively, our results suggest a model for anteroposterior patterning of the amphioxus nerve cord and pharynx, which is probably applicable to vertebrates as well, in which a low anterior level of AmphiRAR (caused, at least in part, by competitive inhibition by AmphiTR2/4) is necessary for patterning the forebrain and formation of gill slits, the posterior extent of both being set by a sharp increase in the level of AmphiRAR. Supplemental data available on-line