RESUMO
DNA hypomethylation may activate oncogene transcription, thus promoting carcinogenesis and tumor development. S-adenosylmethionine (SAM) is a methyl donor in numerous methylation reactions and acts as an inhibitor of intracellular demethylase activity, which results in hypermethylation of DNA. The main objectives of this study were to determine whether DNA hypomethylation correlated with vascular endothelial growth factor-C (VEGF-C) expression, and the effect of SAM on VEGF-C methylation and gastric cancer growth inhibition. VEGF-C expression was assayed by Western blotting and RT-qPCR in gastric cancer cells, and by immunohistochemistry in tumor xenografts. VEGF-C methylation was assayed by bisulfite DNA sequencing. The effect of SAM on cell apoptosis was assayed by flow cytometry analyses and its effect on cancer growth was assessed in nude mice. The VEGF-C promoters of MGC-803, BGC-823, and SGC-7901 gastric cancer cells, which normally express VEGF-C, were nearly unmethylated. After SAM treatment, the VEGF-C promoters in these cells were highly methylated and VEGF-C expression was downregulated. SAM also significantly inhibited tumor growth in vitro and in vivo. DNA methylation regulates expression of VEGF-C. SAM can effectively induce VEGF-C methylation, reduce the expression of VEGF-C, and inhibit tumor growth. SAM has potential as a drug therapy to silence oncogenes and block the progression of gastric cancer.
Assuntos
Animais , Humanos , Masculino , Antineoplásicos/farmacologia , Metilação de DNA/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , S-Adenosilmetionina/farmacologia , Neoplasias Gástricas/tratamento farmacológico , Fator C de Crescimento do Endotélio Vascular/metabolismo , Apoptose/efeitos dos fármacos , Western Blotting , Linhagem Celular Tumoral , Carcinogênese/efeitos dos fármacos , Metilação de DNA/genética , Citometria de Fluxo , Regulação Neoplásica da Expressão Gênica/fisiologia , Xenoenxertos/efeitos dos fármacos , Imuno-Histoquímica , Camundongos Nus , Oncogenes/efeitos dos fármacos , Regiões Promotoras Genéticas/efeitos dos fármacos , Reação em Cadeia da Polimerase em Tempo Real , RNA Mensageiro/análise , Neoplasias Gástricas/metabolismo , Fator C de Crescimento do Endotélio Vascular/efeitos dos fármacos , Fator C de Crescimento do Endotélio Vascular/genéticaRESUMO
Effect of a potent methylation inhibitor oxidized adenosine (Adox), and a universal methyl group donor S-adenosyl-L-methionine (AdoMet) on trehalose metabolism was studied in two haploids of S. cerevisiae of mating types MATalpha, met3 (6460 -8D) and MATa, leu2, ura3, his4 (8534 -10A). Trehalose level decreased in presence of Adox in both strains. Both neutral trehalase (NT) and trehalose-6-phosphate (tre-6-p) synthase activities increased in presence of Adox in -8D strain. Decrease in trehalose level in -8D thus could not be explained in the light of increased tre-6-p synthase activity; however, it could be correlated with increased NT activity. In strain -10A, NT activity was reduced in presence of Adox while tre-6-p synthase activity increased. Enzyme activity profiles in -10A thus do not explain the reduced trehalose level on Adox treatment. Effect of AdoMet was not very prominent in either strain, though in -8D a small increase in trehalose level was seen on treatment. Intracellular AdoMet level of untreated cells of -10A was seen to be almost six times higher than that of -8D. Further, AdoMet treatment caused increase in its level compared to untreated cells, suggesting AdoMet uptake. No effect of either compound was seen on acid trehalase (AT) activity in any strain. The results suggest that there was a possible effect of demethylation on trehalose metabolism (particularly in the synthetic direction) in both strains, though effect of methylation was not very prominent, the reason for which is not very clear.