RESUMEN
Polyalanine repeat expansion diseases are hypothesized to result from unequal chromosomal recombination, yet mechanistic studies are lacking. We identified two de novo cases of hand-foot-genital syndrome (HFGS) associated with polyalanine expansions in HOXA13 that afforded rare opportunities to investigate the mechanism. The first patient with HFGS was heterozygous for a de novo nine codon polyalanine expansion. Haplotype investigation showed that the expansion arose on the maternally inherited chromosome but not through unequal crossing over between homologs, leaving unequal sister chromatid exchange during mitosis or meiosis or slipped mispairing as possible explanations. The asymptomatic father of the second patient with HFGS was mosaic for a six codon polyalanine expansion. Multiple tissue PCR and clonal analysis of paternal fibroblasts showed only expansion/WT and WT/WT clones, and haplotype data showed that two unaffected offspring inherited the same paternal allele without the expansion, supporting a postzygotic origin. Absence of the contracted allele in the mosaic father does not support sister chromatid exchange in the origin of the expansion. Mosaicism for HOXA13 polyalanine expansions may be associated with a normal phenotype, making examination of parental DNA essential in apparently de novo HFGS cases to predict accurate recurrence risks. We could not find an example in the literature where unequal sister chromatid exchange has been proven for any polyalanine expansion, suggesting that the principal mechanism for polyalanine expansions (and contractions) is slipped mispairing without repair or that the true frequency of unequal sister chromatid exchange involving these repeats is low.
Asunto(s)
Anomalías Múltiples/genética , Expansión de las Repeticiones de ADN/genética , Deformidades Congénitas del Pie/genética , Deformidades Congénitas de la Mano/genética , Proteínas de Homeodominio/genética , Anomalías Urogenitales/genética , Adolescente , Adulto , Femenino , Humanos , Recién Nacido , Masculino , Mutación , Péptidos , FenotipoRESUMEN
Receptor tyrosine kinases (RTKs) play important roles in the control of fundamental cellular processes, influencing the balance between cell proliferation and death. RTKs have emerged as molecular targets for the treatment of various cancers. Green tea and its polyphenolic compounds, the catechins, exhibit chemopreventive and chemotherapeutic properties in many human cancer cell types, as well as in various carcinogenicity models in vivo. Epidemiological studies are somewhat less convincing, but some positive correlations have been observed. The tea catechins, including (-)-epigallocatechin-3-gallate (EGCG), have pleiotropic effects on cellular proteins and signaling pathways. This review focuses on the ability of the tea constituents to suppress RTK signaling, and summarizes the mechanisms by which EGCG and other catechins might exert their protective effects towards dysregulated RTKs in cancer cells. The findings are discussed in the context of ongoing clinical trials with RTK inhibitors, and the possibility for drug/nutrient interactions enhancing therapeutic efficacy.
Asunto(s)
Anticarcinógenos/farmacología , Catequina/farmacología , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Té/metabolismo , Animales , Anticarcinógenos/uso terapéutico , Catequina/uso terapéutico , Humanos , Neoplasias/tratamiento farmacológico , Proteínas Tirosina Quinasas Receptoras/metabolismoRESUMEN
The Met receptor tyrosine kinase is deregulated in a variety of cancers and is correlated with advanced stage and poor prognosis. Thus, Met has been identified as an attractive candidate for targeted therapy. We compared the tea polyphenol (-)-epigallocatechin-3-gallate (EGCG) and a specific Met inhibitor, SU11274, as suppressing agents of Met signaling in HCT116 human colon cancer cells. Treatment with hepatocyte growth factor increased phospho-Met levels, and this was inhibited in a concentration-dependent manner by EGCG and SU11274 (IC(50) 3.0 vs. 0.05muM, respectively). Downstream activation of Erk and Akt signaling pathways also was suppressed. Both compounds at a concentration of 5muM lowered cell viability and proliferation, with EGCG being more effective than SU11274, and the invasion of colon cancer cells in Matrigel assays was strongly inhibited. These findings are discussed in the context of the pleiotropic effects of tea catechins, their tissue metabolite levels, and the potential to inhibit colon cancer metastasis and invasion.
Asunto(s)
Catequina/análogos & derivados , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/metabolismo , Proteínas Proto-Oncogénicas c-met/antagonistas & inhibidores , Receptores de Factores de Crecimiento/antagonistas & inhibidores , Antineoplásicos/farmacología , Catequina/farmacología , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Neoplasias del Colon/patología , Células HCT116 , Factor de Crecimiento de Hepatocito/farmacología , Humanos , Indoles/farmacología , Invasividad Neoplásica/prevención & control , Fosfatidilinositol 3-Quinasas/metabolismo , Piperazinas/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Transducción de Señal/efectos de los fármacos , Sulfonamidas/farmacología , Té/químicaRESUMEN
Most cancer deaths result from spread of the primary tumor to distant sites (metastasis). MET is an important protein for metastasis in multiple tumor types. Here we report on the ability of tea catechins to suppress MET activation in human colon cancer cells and propose a mechanism by which they might compete for the kinase domain of the MET protein.
Asunto(s)
Antineoplásicos/farmacología , Camellia sinensis , Catequina/farmacología , Flavonoides/farmacología , Modelos Moleculares , Fenoles/farmacología , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Receptores de Factores de Crecimiento/antagonistas & inhibidores , Antineoplásicos/química , Catequina/química , Línea Celular Tumoral , Neoplasias del Colon , Ensayos de Selección de Medicamentos Antitumorales , Ácido Gálico/química , Humanos , Cinética , Polifenoles , Proteínas Proto-Oncogénicas/química , Proteínas Proto-Oncogénicas c-met , Receptores de Factores de Crecimiento/química , Relación Estructura-ActividadRESUMEN
Colorectal cancer is the second leading cause of cancer-related deaths in the U.S. Met, the receptor for hepatocyte growth factor (HGF), is over-expressed in colon tumors and is associated with poor prognosis. Recently, the green tea polyphenol (-)-epigallocatechin gallate (EGCG) was reported to suppress Met activation in breast cancer cells. However, the possible confounding effect of hydrogen peroxide (H(2)O(2)), produced when EGCG is added to cell culture media, was not assessed. In the present study, the human colon cancer cell lines HCT116 and HT29 were used to examine the relationships between Met activation, EGCG treatment, and H(2)O(2) generation. At concentrations of 0.5, 1, and 5 microM, EGCG suppressed markedly the activation of Met in the presence of HGF. Concentrations of 10muM EGCG and below generated low amounts of H(2)O(2) (<1.5 microM), whereas higher H(2)O(2) concentrations (>5 microM) were required to directly increase the phosphorylation of Met. Moreover, suppression of Met activation by EGCG occurred in the presence or absence of catalase, suggesting that such effects were not an 'artifact' of H(2)O(2) generated from EGCG in cell culture media. We conclude that EGCG might be a beneficial therapeutic agent in the colon, inhibiting Met signaling and helping to attenuate tumor spread/metastasis, independent of H(2)O(2)-related mechanisms.
Asunto(s)
Antineoplásicos/farmacología , Antioxidantes/farmacología , Catequina/análogos & derivados , Neoplasias del Colon/metabolismo , Peróxido de Hidrógeno/metabolismo , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Receptores de Factores de Crecimiento/antagonistas & inhibidores , Catequina/farmacología , Línea Celular Tumoral , Humanos , Proteínas Proto-Oncogénicas c-metRESUMEN
Organic chemistry has made possible the synthesis of molecules that expand on Nature's genetic alphabet. Using the previously described nonstandard DNA base pair constructed from isoguanine and 5-methylisocytosine, we report a highly specific and sensitive method that allows for the fast and specific quantitation of genetic sequences in a closed tube format. During PCR amplification, enzymatic site-specific incorporation of a quencher covalently linked to isoguanine allows for the simultaneous detection and identification of multiple targets. The specificity of method is then established by analysis of thermal denaturation or melting of the amplicons. The appropriate functions of all reactions are further verified by incorporation of an independent target into the reaction mixture. We report that the method is sensitive down to the single copy level, and specificity is demonstrated by multiplexed end-point genotypic analysis of four targets simultaneously using four separate fluorescent reporters. The method is general enough for quantitative and qualitative analysis of both RNA and DNA using previously developed primer sets. Though the method described employs the commonly used PCR, the enzymatic incorporation of reporter groups into DNA site-specifically should find broad utility throughout molecular biology.