RESUMO
The CXCL12 chemokine binds to the CXCR4 receptor and contributes to survival, proliferation, and migration of malignant cells. Recent reports indicate that breast cancer cells lacking expression of CXCL12 but exhibiting CXCR4 can metastasize to target organs that secrete CXCL12. We observed that Tamoxifen (Tam), similarly to 5-dAzaC, results in significantly increased levels of CXCL12 transcript and protein in MCF-7 breast cancer cells. Bisulfite sequencing suggests that Tam, similarly to 5-dAzaC, may increase CXCL12 expression via reduction in methylation of cytosine in the cytosine-guanosine (CpG) dinucleotide island of the CXCL12 promoter of MCF-7 cells. Our results, together with findings of other researches, may suggest that Tam epigenetically activates CXCL12 expression in breast cancer cells and can make these cells less susceptible to attraction by exogenous CXCL12 to metastasis sites.
Assuntos
Antineoplásicos Hormonais/farmacologia , Neoplasias da Mama/tratamento farmacológico , Quimiocina CXCL12/efeitos dos fármacos , Tamoxifeno/farmacologia , Antimetabólitos Antineoplásicos/farmacologia , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Quimiocina CXCL12/genética , Citosina/metabolismo , Metilação de DNA/efeitos dos fármacos , Decitabina , Epigênese Genética/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Análise de Sequência de DNA/métodosRESUMO
The effect of DNA methylation on CXCR4 expression has been demonstrated in pancreatic cancer and melanoma cells, but little is known about the effect of DNA methyltransferases 1 and 3 (DNMT1 and DNMT3B) on CXCR4 expression. Employing lentiviral vectors, we created stable RNA interference-mediated knockdown of DNMT1 and DNMT3B in AsPC1 pancreatic cancer cells. Using reverse transcription real-time quantitative PCR and flow cytometric analysis, we evaluated the increase in the expression of CXCR4 transcript and protein levels in these cells. Bisulfite sequencing analysis showed that the level of promoter demethylation appeared more effective in cells with knockdown of DNMT1 than in those with DNMT3B knockdown. Furthermore, the combined RNA interference knockdown of both DNMT1 and DNMT3B increased promoter demethylation, leading to a slight increase in CXCR4 expression. However, the demethylating agent 5-Aza-2'-deoxycytidine exhibited the strongest effect on promoter demethylation, which correlated with the highest production of CXCR4 transcript and protein in AsPC1 cells. Our results indicate that DNMT1 plays the main role in maintenance of methylation of CXCR4 promoter, while DNMT3B may function as an accessory DNA methyltransferase to modulate CXCR4 expression in AsPC1 cells.
